Abstract

Auxin concentration gradients, established by polar transport of auxin, are essential for the establishment and maintenance of polar growth and morphological patterning. Three families of cellular transport proteins, PIN-formed (PIN), P-glycoprotein (ABCB/PGP), and AUXIN RESISTANT 1/LIKE AUX1 (AUX1/LAX), can independently and co-ordinately transport auxin in plants. Regulation of these proteins involves intricate and co-ordinated cellular processes, including protein–protein interactions, vesicular trafficking, protein phosphorylation, ubiquitination, and stabilization of the transporter complexes on the plasma membrane.

Introduction

The phytohormone indole-3-acetic acid (IAA), or auxin, plays an essential role in embryogenesis (Friml et al., 2003), cell division and elongation (Campanoni and Nick, 2005), vascular tissue differentiation (Mattsson et al., 2003), phyllotactic patterning (Bainbridge et al., 2008), lateral root formation (Dubrovsky et al., 2008), phototropism (Kimura and Kagawa, 2006), gravitropism (Palme et al., 2006), and other physiological processes. Although auxin was the first phytohormone to be discovered (Went, 1927), the molecular mechanisms underlying its transport and perception have only been elucidated in the past decade (Kepinski, 2007; Delker et al., 2008).

IAA is synthesized in the shoot, particularly by leaf primordia and young leaves, and transported to the root through vascular and bundle sheath tissues (Ljung et al., 2005; Bandyopadhyay et al., 2007). The synthesis, transport, and catabolism of IAA is tightly regulated by both transcriptional and post-transcriptional processes that are co-ordinately regulated via the ubiquitination of AUX/IAA repressor proteins by the SCFTIR1/AFB mechanism followed by proteolytic degradation (Quint and Gray, 2006). Additional post-transcriptional mechanisms further regulate auxin transport. This review focuses on the role of post-translational mechanisms that regulate auxin transport processes by modifying, activating, redistributing, or degrading auxin transport proteins or protein complexes.

Polar auxin transport

Auxin is polarly transported from cell to cell in a process that involves chemiosmotically-driven export to and uptake from the apoplast. As IAA is a weak organic acid (pKa=4.75), it exhibits a ∼5:1 distribution of anionic (IAA): protonated (IAAH) speciation in the acidic (pH ∼5.5) apoplast. IAA can thus enter the cell by either lipophilic diffusion of IAAH or by anionic uptake via H+-IAA symporters. The latter process is required as the diffusive flux of IAAH across the plasma membrane is thought to be an order of magnitude slower than that of carrier-mediated IAA translocation, and diffusion alone cannot account for auxin fluxes that naturally occur in plants (Kramer and Bennett, 2006). In contrast, the cellular efflux of IAA requires protein mediation, as IAA is almost exclusively anionic in the cytoplasm (pH ∼7.0) and cannot diffuse across the membrane on its own. The localization and activity of auxin transport complexes are thus crucial in establishing the polarity of auxin transport.

The concentration gradient created by directional movement of auxin is fundamental to the establishment of plant axial polarity, organ patterning, and morphological adaptation to the environment (De Smet and Jurgens, 2007). From the first cell division in plant embryogenesis, auxin is preferentially accumulated in the zygotic apical cell where it functions as an important determinant of that cell's proembryonic fate (Friml et al., 2003). Vascular differentiation also coincides with auxin accumulation in preprocambial cells (Mattsson et al., 2003). In the root, acropetal transport (base to apex) of auxin (Blakeslee et al., 2005a) within the stele is responsible for the initiation of lateral root primordia from pericycle cells that can respond to auxin activation (Casimiro et al., 2001; Bhalerao et al., 2002; Wu et al., 2007). In Arabidopsis, the priming of pericycle cells for auxin responsiveness occurs in the basal region of the root meristem and is controlled by a periodic shift in the basipetal (apex to base) redistribution of auxin through the lateral root cap and epidermal cell files, resulting in an alternating pattern of regularly spaced lateral roots (De Smet et al., 2007). In gravitropic root bending, asymmetric changes in the basipetal (root apex to base) transport stream caused by alteration in the gravitational vector leads to differential root elongation and bending in the direction of the gravitational vector (Palme et al., 2006). Similarly, phototropic bending in hypocotyls is thought to result from asymmetric accumulation of auxin in cells distal to the site of illumination resulting in asymmetric growth and bending of the hypocotyl toward light (Kimura and Kagawa, 2006).

Classes of auxin transport proteins

Auxin transport proteins have been identified and, to date, have been grouped in three families: AUXIN RESISTANT 1/LIKE AUX1 (AUX1/LAX) uptake symporters, PIN-FORMED (PIN) efflux carriers, and P-GLYCOPROTEIN (MDR/PGP/ABCB) efflux/conditional transporters.

AUX1/LAX uptake symporters

AUX1 was originally identified in a genetic screen for Arabidopsis mutants that exhibited auxin-resistant root growth (Bennett et al., 1996). The AUX1 gene encodes a transmembrane protein similar to amino acid permeases. AUX1 participates in loading of shoot auxin into the phloem for long-distance transport toward the root tip and in the basipetal transport of auxin out of the lateral root cap at the root apex (Swarup et al., 2002, 2004). The ability of AUX1 to mediate auxin influx has been demonstrated in planta and by expression in heterologous systems. Treatment with the membrane-permeable artificial auxin 1-naphthaleneacetic acid (1-NAA) was shown to rescue the agravitropic phenotype of the aux1 mutant, which is also resistant to the weakly permeate and poorly transported auxin herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) (Marchant et al., 1999). When expressed in Xenopus oocytes, AUX1 was shown to function as a high-affinity auxin uptake carrier protein (Yang et al., 2006; Kerr and Bennett, 2007).

AUX1 is localized to the lower end of the cells in the lateral root cap, epidermal cells below the elongation zone, columella, and protophloem (Kleine-Vehn et al., 2006). The three members of the Like-AUX1 (LAX) family are functional analogues of AUX1, and appear to function in tissue-specific auxin uptake (Swarup et al., 2004). Quadruple aux/lax mutants exhibit aberrant formation of leaf primordia and reduced polar PIN localization consistent with altered auxin flux (Bainbridge et al., 2008). LAX3 functions in the early stages of lateral root formation (de Billy et al., 2001; Swarup et al., 2008).

PIN efflux carriers

The Arabidopsis pin-formed1 (pin1) mutant exhibits defects in vascular patterning, organogenesis, and phyllotaxis (Galweiler et al., 1998; Reinhardt, 2005). PIN proteins belong to the unique auxin efflux facilitator family found in plants and some fungi and are predicted to have 10–12 transmembrane domains (Galweiler et al., 1998; Blakeslee et al., 2007). Among the eight members of the PIN family in Arabidopsis, five have been experimentally shown to function as auxin efflux carrier proteins when expressed in Arabidopsis, tobacco BY-2, human HeLa, and/or yeast cell cultures (Petrasek et al., 2006; Blakeslee et al., 2007). PIN-mediated efflux in these heterologous systems was partially inhibited by the auxin efflux inhibitor 1-naphthylphthalamic acid (NPA), although, in all cases when it was used, NPA strongly inhibited background auxin efflux in cells not expressing recombinant PIN proteins, especially in plant cell systems (Petrasek et al., 2006).

Subcellular localization of PIN proteins maps with the directionality of auxin transport vectors, especially in embryonic development and organogenesis (Benkova et al., 2003; Blilou et al., 2005). In the central vascular cylinder, PIN1 is basally localized in the xylem parenchyma and participates in the transport of auxin along the embryonic axis from the shoot to the root tip. PIN2 exhibits a basal (bottom) and lateral localization in root cortical cells and apical (top) localization in root epidermal cells consistent with its apparent role in redirection and reflux of auxin at the root tip (Chen et al., 1998; Luschnig et al., 1998; Muller et al., 1998). PIN3 exhibits an apolar orientation in root columella cells, but relocalizes in the direction of auxin movement upon gravistimulation (Friml et al., 2002b). PIN3 is also localized to the inner surface of hypocotyl bundle sheath cells where it appears to function in the redirection of auxin back into the vascular cylinder. PIN3 is also found in epidermal cells, and the pin3 mutant shows shorter epidermal cells in light-grown hypocotyls, thought to be caused by a defect in cell elongation (Friml et al., 2002b). As mutational analysis indicates that PIN3 functions in tropic responses, activation of its transport activity near sites of illumination could be expected to accelerate auxin movement out of tissues on the illuminated (non-bending side) of the hypocotyl.

Other PIN proteins exhibit primarily apolar subcellular localizations but still contribute to directional auxin movement. PIN4 exhibits mixed polar and apolar localizations in the provascular quiescent centre and daughter cells and functions in root meristem patterning (Friml et al., 2002a). PIN7 is abundant in epidermal tissues where it exhibits a non-polar localization (Blakeslee et al., 2007), and is also involved in the establishment of the apical–basal axis, particularly in the hypophysis (Friml et al., 2003). Despite their seemingly discrete expression patterns and functions, some redundancy is found among PIN family members as compensatory expression of some PIN genes was observed in pin1 and pin2 mutants, and ectopic expression of these PIN homologues was sufficient to rescue the auxin transport phenotype of the mutants (Vieten et al., 2005). One member of the PIN family, PIN5, is particularly intriguing, as it lacks a variable central domain common to other characterized PIN proteins that, in the case of PIN1 has been shown to mediate protein–protein interactions (Blakeslee et al., 2007). Analysis of PIN5 auxin transport activity (if any) will help determine whether the central variable domain of the PIN proteins plays a functional role in transport or has a primarily regulatory function as has been proposed.

Some evidence suggests that auxin efflux proteins also mediate intercellular auxin movement from mildly alkaline organelles to the neutral cytosol. Anionic auxin accumulation within compartments with a pH >7.0 is predicted by chemiosmotic models, and the essential auxin binding protein ABP1 is relatively abundant in the mildly alkaline endoplasmic reticulum (Timpte, 2001). Further, the toxic effects of the artificial auxin 2,4-dichlorophenoxyacetic acid (2,4-D) are associated with its accumulation in the endoplasmic reticulum (Dharmasiri et al., 2006). As compared to IAA, 2,4-D is poorly transported by efflux carrier proteins, one of the uncharacterized PIN transporters, such as PIN5 or PIN8, and/or one or more uncharacterized ABCB transporter may mediate auxin efflux from the ER.

ABCB efflux transporters

A third class of auxin transporters are phospho-glycoproteins (PGPs) that belong to the ABCB subgroup of the ATP-Binding-Cassette (ABC) transporter superfamily. The best known member of the ABCB subgroup is the human ABCB1 protein which has been extensively studied for its role in increased resistance to chemotherapeutic agents resulting from its overexpression in cancer cells (Luckie et al., 2003). However, the use of the multidrug resistance (MDR) nomenclature for this subgroup of proteins has been discontinued as the majority of family members appear to exhibit a higher degree of transport substrate specificity than mammalian ABCB1 (Verrier et al., 2008). In Arabidopsis thaliana, the 21 members of ABCB subgroup exhibit both distinct and overlapping expression patterns throughout all stages of plant growth and development (Blakeslee et al., 2005b). The best characterized members of Arabidopsis ABCB proteins are the auxin transporters ABCB1, ABCB4, and ABCB19. Multiple reports have catalogued PIN and AUX/LAX gene expression and protein localization (Blakeslee et al., 2005a, b; Tanaka et al., 2006; Zazimalova et al., 2007). By contrast, a current summary of ABCB auxin transporter gene expression and ABCB protein distribution is lacking in the literature. A brief summary of the expression patterns of ABCB auxin transporter genes is provided in Table 1 and a summary of protein localization is shown in Fig. 1.

Table 1.

Summary of microarray data indicating ABCB1, ABCB4, and ABCB19 gene expression (from Genevestigator, https://www.genevestigator.ethz.ch/)

Anatomy
 
ABCB1
 
ABCB4
 
ABCB19
 
Mean  SE Mean  SE Mean  SE 
graphic 0 Callus 5538 ± 406 4427 ± 368 2037 ± 132 
1 Cell suspension 5270 ± 294 3766 ± 256 807 ± 107 
2 Seedling 2889 ± 53 1646 ± 72 2485 ± 50 
    21 Cotyledons 2186 ± 163 186 ± 24 1786 ± 227 
    22 Hypocotyl 5951 ± 922 1343 ± 330 1769 ± 381 
    23 Radicle 4265 ± 265 2324 ± 92 3111 ± 106 
3 Inflorescence 4538 ± 185 366 ± 46 2794 ± 134 
    31 Flower 4562 ± 235 144 ± 25 3348 ± 231 
        311 Carpel 5241 ± 449 50 ± 4205 ± 822 
            3111 Ovary 3926 ± 157 57 ± 2223 ± 264 
            3112 Stigma 7374 ± 642 42 ± 12 619 ± 105 
        312 Petal 4601 ± 698 46 ± 14 5024 ± 1004 
        313 Sepal 3231 ± 331 883 ± 113 921 ± 334 
        314 Stamen 1256 ± 321 44 ± 12 1561 ± 511 
            3141 Pollen 53 ± 27 ± 14 274 ± 13 
        315 Pedicel 8765 ± 134 139 ± 18 4159 ± 120 
    32 Silique 5351 ± 554 202 ± 40 1889 ± 246 
    33 Seed 1641 ± 108 885 ± 174 1655 ± 251 
    34 Stem 8620 ± 508 502 ± 76 2068 ± 259 
    35 Node 10654 ± 134 475 ± 21 1588 ± 59 
    36 Shoot apex 5026 ± 229 100 ± 4827 ± 311 
    37 Cauline leaf 2588 ± 141 540 ± 105 747 ± 26 
4 Rosette 2497 ± 41 708 ± 58 1413 ± 32 
    41 Juvenile leaf 2492 ± 130 1267 ± 335 1253 ± 66 
    42 Adult leaf 1999 ± 50 540 ± 43 957 ± 42 
    43 Petiole 3512 ± 421 143 ± 15 4356 ± 227 
    44 Senescent leaf 1609 ± 62 1138 ± 69 254 ± 11 
    45 Hypocotyl 3250 ± 252 2293 ± 296 424 ± 80 
        451 Xylem 2667 ± 74 3225 ± 93 180 ± 18 
        452 Cork 4544 ± 215 1544 ± 119 585 ± 66 
5 Roots 4245 ± 81 4187 ± 183 3197 ± 82 
    52 Lateral root 4685 ± 392 2841 ± 803 1496 ± 391 
    53 Root tip 2913 ± 191 1791 ± 361 6225 ± 886 
    54 Elongation zone 4845 ± 179 4119 ± 696 5012 ± 1084 
    55 Root hair zone 5197 ± 563 8427 ± 1175 4868 ± 555 
    56 Endodermis 4094 ± 774 4560 ± 279 5503 ± 571 
    57 Endodermis+cortex 3338 ± 146 2266 ± 569 7226 ± 148 
    58 Epid. atrichoblasts 4458 ± 413 2864 ± 85 3511 ± 415 
    59 Lateral root cap 4985 ± 344 3151 ± 787 4154 ± 469 
    60 Stele 4351 ± 98 2303 ± 101 5567 ± 435 
Anatomy
 
ABCB1
 
ABCB4
 
ABCB19
 
Mean  SE Mean  SE Mean  SE 
graphic 0 Callus 5538 ± 406 4427 ± 368 2037 ± 132 
1 Cell suspension 5270 ± 294 3766 ± 256 807 ± 107 
2 Seedling 2889 ± 53 1646 ± 72 2485 ± 50 
    21 Cotyledons 2186 ± 163 186 ± 24 1786 ± 227 
    22 Hypocotyl 5951 ± 922 1343 ± 330 1769 ± 381 
    23 Radicle 4265 ± 265 2324 ± 92 3111 ± 106 
3 Inflorescence 4538 ± 185 366 ± 46 2794 ± 134 
    31 Flower 4562 ± 235 144 ± 25 3348 ± 231 
        311 Carpel 5241 ± 449 50 ± 4205 ± 822 
            3111 Ovary 3926 ± 157 57 ± 2223 ± 264 
            3112 Stigma 7374 ± 642 42 ± 12 619 ± 105 
        312 Petal 4601 ± 698 46 ± 14 5024 ± 1004 
        313 Sepal 3231 ± 331 883 ± 113 921 ± 334 
        314 Stamen 1256 ± 321 44 ± 12 1561 ± 511 
            3141 Pollen 53 ± 27 ± 14 274 ± 13 
        315 Pedicel 8765 ± 134 139 ± 18 4159 ± 120 
    32 Silique 5351 ± 554 202 ± 40 1889 ± 246 
    33 Seed 1641 ± 108 885 ± 174 1655 ± 251 
    34 Stem 8620 ± 508 502 ± 76 2068 ± 259 
    35 Node 10654 ± 134 475 ± 21 1588 ± 59 
    36 Shoot apex 5026 ± 229 100 ± 4827 ± 311 
    37 Cauline leaf 2588 ± 141 540 ± 105 747 ± 26 
4 Rosette 2497 ± 41 708 ± 58 1413 ± 32 
    41 Juvenile leaf 2492 ± 130 1267 ± 335 1253 ± 66 
    42 Adult leaf 1999 ± 50 540 ± 43 957 ± 42 
    43 Petiole 3512 ± 421 143 ± 15 4356 ± 227 
    44 Senescent leaf 1609 ± 62 1138 ± 69 254 ± 11 
    45 Hypocotyl 3250 ± 252 2293 ± 296 424 ± 80 
        451 Xylem 2667 ± 74 3225 ± 93 180 ± 18 
        452 Cork 4544 ± 215 1544 ± 119 585 ± 66 
5 Roots 4245 ± 81 4187 ± 183 3197 ± 82 
    52 Lateral root 4685 ± 392 2841 ± 803 1496 ± 391 
    53 Root tip 2913 ± 191 1791 ± 361 6225 ± 886 
    54 Elongation zone 4845 ± 179 4119 ± 696 5012 ± 1084 
    55 Root hair zone 5197 ± 563 8427 ± 1175 4868 ± 555 
    56 Endodermis 4094 ± 774 4560 ± 279 5503 ± 571 
    57 Endodermis+cortex 3338 ± 146 2266 ± 569 7226 ± 148 
    58 Epid. atrichoblasts 4458 ± 413 2864 ± 85 3511 ± 415 
    59 Lateral root cap 4985 ± 344 3151 ± 787 4154 ± 469 
    60 Stele 4351 ± 98 2303 ± 101 5567 ± 435 

The highest expression is shown in bold. Data presented are means and standard errors of normalized Affymetrix expression values.

Fig. 1.

Localization of ABCB1, ABCB4, and ABCB19 proteins in Arabidopsis roots. Shown are ProABCB1:ABCB1-GFP (transgenic lines courtesy from Dr Jiri Friml), ProABCB4:ABCB4-GFP (transgenic lines courtesy from Dr Misuk Cho), and ProABCB19:ABCB19-GFP (transgenic lines courtesy from Dr Jiri Friml). Bar=25 μm.

Fig. 1.

Localization of ABCB1, ABCB4, and ABCB19 proteins in Arabidopsis roots. Shown are ProABCB1:ABCB1-GFP (transgenic lines courtesy from Dr Jiri Friml), ProABCB4:ABCB4-GFP (transgenic lines courtesy from Dr Misuk Cho), and ProABCB19:ABCB19-GFP (transgenic lines courtesy from Dr Jiri Friml). Bar=25 μm.

The involvement of ABCB proteins in auxin transport was first suggested by Sidler et al., (1998) when expression levels of PGP1/ABCB1 in Arabidopsis were found to regulate hypocotyl elongation in a light-dependent manner (Sidler et al., 1998). ABCB1 was subsequently shown to function co-ordinately with PGP19/MDR1/ABCB19 in mediating polar auxin transport in Arabidopsis (Noh et al., 2001). The sequence of ABCB19 is highly similar to that of ABCB1 (Verrier et al., 2008). Arabidopsis abcb1 and abcb19 mutants exhibit reductions in both growth and root basipetal auxin transport with the most pronounced reductions seen in the double abcb1 abcb19 mutant. Polar auxin transport is reduced ∼70% in abcb1 abcb19, while pin1 exhibits a ∼30% reduction (Blakeslee et al., 2007), but abcb mutants show none of the defects in organogenesis that are seen in pin1 (Noh et al., 2001). This suggests that ABCBs primarily regulate long-distance auxin transport and localized loading of auxin into the transport system and do not function in establishing the basal vectorial auxin flows that function in organogenesis (Bandyopadhyay et al., 2007; Blakeslee et al., 2007; Bailly et al., 2008). This interpretation of ABCB auxin transport function was confirmed when loss of function of ABCB1 orthologues were found to underlie the dwarf phenotypes of the agriculturally-important brachytic2/zmabcb1 maize and dwarf3/sbabcb1 sorghum mutants (Multani et al., 2003).

Arabidopsis abcb19 also exhibits exaggerated phototropic and gravitropic responses (Noh et al., 2001, 2003; Lin and Wang, 2005; Lewis et al., 2007; Wu et al., 2007). In addition, it has recently been shown that the expression level of ABCB19 is suppressed upon activation of the phytochrome and cryptochrome photoreceptors in response to the red and blue light, respectively (Nagashima et al., 2008). These results point to an ABCB19 function in the repression of the differential growth of the light- and gravity-stimulated hypocotyl (Noh et al., 2003; Nagashima et al., 2008).

A direct role for ABCB1 and ABCB19 in cellular efflux was demonstrated when increased auxin retention was observed in mesophyll protoplasts from Arabidopsis abcb1 and abcb19 mutants (Geisler et al., 2005). Further, as was seen with PIN proteins, heterologous expression of ABCB1 and ABCB19 in HeLa and/or yeast cells resulted in enhanced auxin efflux that was inhibited by NPA (Geisler et al., 2005; Bouchard et al., 2006). However, unlike what was seen with PIN expression, the efflux mediated by the ABCBs in mammalian cells was insensitive to inhibitors of mammalian organic anion transporters, suggesting that ABCB-mediated auxin export did not involve activation of an endogenous transport activity (Geisler et al., 2005; Petrasek et al., 2006).

ABCB4 functions in the movement of auxin away from the root tip and appears to function primarily in the regulated export of auxin out of the elongation zone (Santelia et al., 2005; Terasaka et al., 2005; Cho et al., 2007; Lewis et al., 2007). ABCB4 exhibits structural similarity to the berberine uptake transporter CjMDR1/CjABCB1 from the medicinal plant Coptis japonica (Shitan et al., 2003) and, to a lesser extent, the ABCB14 malate uptake transporter from Arabidopsis guard cells (Lee et al., 2008), but exhibits dissimilarity in putative substrate binding sites (H Yang and A Murphy, unpublished data). Some experimental evidence indicates that ABCB4 functions in auxin uptake. When expressed in mammalian cells, ABCB4 activates a net increase in auxin retention (Terasaka et al., 2005), and heterologous expression of ABCB4 in the yeast IAA-sensitive yap1 mutant led to enhanced growth sensitivity to IAA (Santelia et al., 2005). However, NPA treatment of mammalian cells expressing ABCB4 activated efflux to levels equivalent to that seen in NPA-treated cells expressing Arabidopsis ABCB1 or ABCB19 (Terasaka et al., 2005). Recently, ABCB4 expression has been found to confer auxin efflux activity in root hairs and tobacco suspension cells (Cho et al., 2007). These results suggest that the directionality of ABCB4-mediated transport is regulated by plant-specific modulators (Cho et al., 2007). However, it is also possible that ABCB4 export is directly activated by threshold levels of transport substrates as is the case with human ABCB1 (Kimura et al., 2007). An examination of published results suggests that auxin efflux activated by ABCB4 is more evident in experiments utilizing longer time periods for assays, while activation of uptake is seen with lower concentrations in shorter time periods. As such, ABCB4 might best be referred to as a conditional auxin efflux/uptake transporter. Structural modelling comparisons of ABCB4 with ABCB1, ABCB19, and the ABCB14 malate uptake transporter (H Yang and A Murphy, unpublished results) suggest that unique structure/sequence variations in ABCB4 underlie its conditional activity.

ABCB4 appears to function primarily in the accumulation of auxin in the elongation zone as well as efflux from root trichoblast cells. Mutations in ABCB4 exhibit decreased linear growth (Terasaka et al., 2005), increased initial rates of root bending (Lewis et al., 2007), and altered root hair formation (Santelia et al., 2005; Cho et al., 2007) that are dependent on growth conditions. All of these functions are consistent with ABCB4 expression patterns, ABCB4 protein distribution and subcellular localization, and auxin transport profiles of abcb4 mutants.

Regulation of auxin transport proteins by protein–protein interactions

Activation of ABCB proteins by TWD1/FKBP42

Multiple lines of evidence suggest that there are at least two distinct NPA-binding sites in Arabidopsis membranes. A high affinity site associated with the inhibition of auxin transport at the plasma membrane is associated with an integral membrane protein, and possibly, an associated peripheral protein (Murphy et al., 2002). A second, low affinity NPA binding site is thought to be a membrane anchored or peripheral amidase (Murphy et al., 2002). Other sites of NPA action have been associated with membrane trafficking events, but require such high concentrations of NPA to be visualized that they can be regarded as non-specific (Geldner et al., 2001). NPA affinity chromatography was initially used to isolate the ABCB1, 4, and 19 proteins (Murphy et al., 2002; Geisler et al., 2003; Terasaka et al., 2005). An FKBP immunophilin-like protein, TWD1/FKBP42 was copurified with the ABCBs (Murphy et al., 2002).

Subsequent studies have established that the C-terminal domains of ABCB1 and ABCB19 interact with FKBP42 and that the phenotypes of abcb1 abcb19 resemble those of the twd1 mutant (Geisler et al., 2003). Based on sequence prediction, FKBP42 was proposed to be a glycophosphatidyl inositol (GPI)-anchored protein (Geisler et al., 2003). However, no GPI moiety was detected when TWD1 was biochemically analysed (Murphy et al., 2002; Granzin et al., 2006), and subsequent structural characterizations are inconsistent with the presence of a GPI anchor (Eckhoff et al., 2005). Further, the abundance of FKBP42 is very low compared to ABCB1 and ABCB19 (Bailly et al., 2008), suggesting that FKBP42 functions in activating ABCB membrane complexes, rather than anchoring complex formation. FKBP42 has been proposed to induce conformational changes in ABCB1 and ABCB19 (Geisler et al., 2003; Bouchard et al., 2006; Bailly et al., 2008). ABCB1–FKBP42 interactions have been shown to be sensitive to both NPA and flavonoid inhibitors of auxin transport, which appear to interact with multiple sites of action in the respective proteins (Bailly et al., 2008). Conformational changes are involved in the regulation of mammalian ABCB (Ambudkar et al., 2006), although little is known about the protein interactions that initiate these changes.

Consistent with this proposed function, loss of FKBP42 conferred resistance to the pharmacological agent gravacin, which is an inhibitor of ABCB19 activity (Rojas-Pierce et al., 2007). Gravacin was originally identified as an inhibitor of gravitropic bending in hypocotyls and was subsequently found to interfere with subcellular targeting of vacuolar marker proteins (Surpin et al., 2005). Wild-type Arabidopsis seedlings treated with gravacin resulted in reductions of auxin transport that were similar to those seen in abcb19, while gravacin treatment of abcb19 mutants resulted in nominal further reductions in auxin transport (Rojas-Pierce et al., 2007). A screen for mutants that are resistant to gravacin resulted in the identification of abcb/pgp19-4 which harbours a point mutation in the C-terminal domain of ABCB19 (E1174K) (Rojas-Pierce et al., 2007).

Treatment with gravacin did not alter the gravitropic response of twd1, and microsomes derived from twd1 showed reduced binding to gravacin (Rojas-Pierce et al., 2007; Bailly et al., 2008). However, the immunolocalization of ABCB19 was unchanged in twd1 mutants compared to wild type (Titapiwatanakun et al., 2008), suggesting that FKBP42 is more likely to function in activation, rather than localization of ABCB1/19 to the plasma membrane.

PIN-ABCB interactions

Although both ABCB19 and PIN1 can function as independent auxin efflux transporters, these proteins can interact and co-ordinately transport auxin (Bandyopadhyay et al., 2007; Blakeslee et al., 2007). Co-ordinated, but independent functions for PINs and ABCB1/19 are particularly evident in embryonic development and lateral root formation. However, physical interaction between ABCB19 and PIN1 in post-embryonic growth is suggested by positive results in subcellular colocalization, coimmunoprecipitation, and yeast two-hybrid interaction analyses (Blakeslee et al., 2007). Moreover, functional interactions between ABCB19 and PIN1 are supported by apparently synergistic phenotypes of pin1 abcb19 mutants and enhanced auxin efflux activity, inhibitor sensitivity, and substrate specificity of HeLa cells co-expressing ABCB19 and PIN1. As was the case with FKBP42/TWD1, interactions of ABCB19 with PIN1 are mediated by the C-terminal domain of the protein, and gravacin effectively interferes with the enhanced auxin transport mediated by ABCB19 and PIN1 coexpression (Rojas-Pierce et al., 2007).

Some evidence of interactions between PIN1 and ABCB1 are suggested by co-immunoprecipitation studies and by increases in auxin efflux when the two proteins are co-expressed in heterologous systems (Blakeslee et al., 2007). However, PIN1–ABCB1 interactions appear to be indirect, as no evidence of protein interactions are seen in yeast two-hybrid assays (Blakeslee et al., 2007). By contrast, interactions of PIN2 with ABCB1 may be more robust, as coexpression of PIN2 and ABCB1 in yeast has synergistic effects on auxin transport and pin2 pgp1 pgp19 mutants exhibit severely agravitropic growth phenotypes (Blakeslee et al., 2007).

Regulation of auxin transporters by auxin levels and fluxes

Auxin has been proposed to ‘canalize’ it own transport by reorienting transport components (Sachs, 1981), presumably by altering the subcellular localization of auxin transport proteins or by regulating their transport activity. The direction of auxin flow has been shown to influence polar PIN localization in a cell type-specific manner (Sauer et al., 2006). For instance, in graviresponding root columella cells, PIN3 is disproportionately oriented on the lower side of cells in the path of auxin destined to accumulate in the epidermal cells of the distal elongation zone (Friml et al., 2002b). Similarly, expression of an apically-localized chimeric PIN1 protein under the control of the PIN2 promoter was able to suppress the root agravitropic phenotype of the pin2 mutant, while comparable localization of basally-localized PIN1 protein failed to produce the same result (Wisniewska et al., 2006). To date, most studies have focused on easily visualized changes in PIN protein localization in response to altered auxin fluxes. However, it is unlikely that PIN proteins are only regulated by localized auxin accumulations. Numerous studies have shown that PIN expression and PIN protein abundance are altered by changes in auxin levels or transport (Peer et al., 2004; Vieten et al., 2005). It is likely that protein phosphorylation and turnover of PINs play an important role in auxin-dependent regulation of PIN function as well.

Expression of ABCB1, ABCB4, and ABCB19 is up-regulated by auxin application (Noh et al., 2001; Geisler et al., 2005; Terasaka et al., 2005). The promoter sequence of ABCB1 includes auxin responsive motifs and the time-course of β-glucuronidase (GUS) expression directed by ABCB1 promoter (ABCB1pro:GUS) at the shoot and root apices in response to auxin treatment indicate that ABCB1 expression responds rapidly to auxin treatment (Geisler et al., 2005). By contrast, ABCB4 appears to be a ‘late’ auxin response gene (Terasaka et al., 2005). In general, protein abundances of ABCB1, 4, and 19 appear to reflect gene expression levels (Geisler et al., 2005; Terasaka et al., 2005; Blakeslee et al., 2007; Wu et al., 2007). Overexpression of ABCB4 does not result in the presence of ectopic ABCB4 protein in the shoot (Terasaka et al., 2005). This appears to be a result of regulation of gene expression or RNA stability, as steady-state ABCB4 RNA levels in the shoot were found to be very low in overexpressing lines (Terasaka et al., 2005). A similar disparity is found with expression of ABCB19, as visualized with a MDR1pro:GFP-MDR1 fusion. No signal is seen in root and anther filament epidermal cells where transcriptional reporters, RT-PCR, and microarray expression reports indicate gene expression (Blakeslee et al., 2007; Wu et al., 2007). However, immunolocalizations, epitope-tags, and other protein–reporter fusions indicate that the protein is present in these cells (Blakeslee et al., 2007). Further, ABCB19 appears to be highly stable and protein abundance responds slowly to decreases in gene expression (Titapiwatanakun et al., 2008).

Regulation of auxin transporters by cellular trafficking mechanisms

The trafficking pathways of the PIN and AUX1 auxin transport proteins have been extensively investigated. Less is known about the trafficking of the ABCBs, primarily due to a strong community bias against the acceptance of these proteins as auxin transporters until recently. In Fig. 2, a summary of the known subcellular trafficking pathways that mediate PIN1, PIN2, AUX1, ABCB1, and ABCB19 to and from the plasma membrane is presented.

Fig. 2.

Model of cellular trafficking pathways utilized by the auxin transporter proteins. Movement of auxin depends on the pH difference between the apoplast and the cytoplasm. The cellular uptake of auxin occurs via the diffusion of IAAH and the import of IAA through the proton-symporter protein AUX1 (purple). The passage out of the cell of IAA is characterized by PIN-formed proteins, shown here for PIN1 (blue) and PIN2 (green). An ATP-dependent transporter, ABCB proteins (pink), are also mediated by cellular auxin efflux. Although the trafficking of both PIN1 and PIN2 is sensitive to BFA, only PIN1 has been shown to utilize the GNOM-dependent pathway. The trafficking of PIN2 depends on the SNX1- and VPS29-containing retromer complex, which is inhibited by wortmannin. The intracellular pool of PIN2 is targeted to proteolysis though ubiquitination. AXR4, an ER-localized protein, is required for the plasma membrane localization of AUX1. The trafficking pathways of AUX1 and ABCB1 are also sensitive to BFA. The trafficking of ABCB19 is not regulated by the same dynamic mechanisms used by PIN1 and PIN2. Gravacin inhibits the targeting of ABCB19 to the plasma membrane.

Fig. 2.

Model of cellular trafficking pathways utilized by the auxin transporter proteins. Movement of auxin depends on the pH difference between the apoplast and the cytoplasm. The cellular uptake of auxin occurs via the diffusion of IAAH and the import of IAA through the proton-symporter protein AUX1 (purple). The passage out of the cell of IAA is characterized by PIN-formed proteins, shown here for PIN1 (blue) and PIN2 (green). An ATP-dependent transporter, ABCB proteins (pink), are also mediated by cellular auxin efflux. Although the trafficking of both PIN1 and PIN2 is sensitive to BFA, only PIN1 has been shown to utilize the GNOM-dependent pathway. The trafficking of PIN2 depends on the SNX1- and VPS29-containing retromer complex, which is inhibited by wortmannin. The intracellular pool of PIN2 is targeted to proteolysis though ubiquitination. AXR4, an ER-localized protein, is required for the plasma membrane localization of AUX1. The trafficking pathways of AUX1 and ABCB1 are also sensitive to BFA. The trafficking of ABCB19 is not regulated by the same dynamic mechanisms used by PIN1 and PIN2. Gravacin inhibits the targeting of ABCB19 to the plasma membrane.

Trafficking mechanism of PINs

The dynamic trafficking of PIN1 is the one of the best studied trafficking mechanisms in plants. PIN1 trafficking is mediated by ADP-ribosylation factors (ARFs), the GNOM ARF guanine nucleotide exchange factor (ARF-GEF) (Steinmann et al., 1999), and the ARF GTPase activating protein (ARF-GAP) SCARFACE (SFC) (Sieburth et al., 2006). Embryos of gnom mutants exhibit severe polarity defects and altered polar localizations of PIN1 (Steinmann et al., 1999). The Sec7 domain of GNOM has been shown to be a target of the fungal toxin brefeldin A (Geldner et al., 2003), and short-term treatments of brefeldin A (BFA) have been shown to reversibly cause intracellular aggregations of PIN1 in cells at the root apex (Geldner et al., 2001). Mutation of the Sec7 domain of GNOM to a BFA-insensitive form (GNOMM696L) prevents the formation of PIN1 aggregations after BFA treatment (Geldner et al., 2003), indicating that PIN1 trafficking is mediated by GNOM. PIN3 has also been shown to associate with the BFA compartment following BFA treatment (Friml et al., 2002b). PIN1 accumulation after BFA treatment in the sfc mutant resulted in multiple smaller organelles, and implicated a role of the SCARFACE ARF-GAP protein in PIN1 cycling as well (Sieburth et al., 2006). In addition, trafficking of PIN1 also requires an intact actin network, but appears not to require microtubules (Geldner et al., 2001).

The time- and dosage-dependent effects of BFA on PIN trafficking have recently been examined (Kleine-Vehn et al., 2008). Long-term incubation with BFA has been shown to promote transcytosis of PIN1 and PIN2, in which the ARF-GEF-mediated endocytotic vesicles was found to be translocated from one side of the cell to the other.

The subcellular trafficking of PIN2 appears to be regulated by different mechanisms than those mobilizing PIN1. Trafficking of PIN2 is mediated by endosomes containing SORTING NEXIN1 (SNX1), which are distinct from GNOM-mediated endosomes and are sensitive to the phosphatidylinositol-3-OH kinase (PI-3K) inhibitor wortmannin (Jaillais et al., 2006). SNX1 is a subunit of the retromer complex, which functions in recycling transmembrane proteins from endosomal multivesicular bodies (MVBs) to the trans-Golgi network in yeast and mammals (Bonifacino and Rojas, 2006). Another important subunit of the retromer complex is the Vacuolar Protein Sorting 29 (VPS29), which is also localized to MVBs. Arabidopsis mutants lacking VPS29 showed the alterations in the morphology of SNX1-containing endosomes and also altered PIN1 localization (Jaillais et al., 2007), suggesting that the retromer complex functions in the endosomal recycling of PIN1 as well.

The endocytosis of PIN proteins has been shown to be inhibited by auxin itself (Paciorek et al., 2005; Zazimalova et al., 2007). Treatment with high auxin concentrations stabilized PIN proteins (PIN1, 2, 3, and 4) on the plasma membrane. This phenomenon may reflect a need for increased PIN-mediated auxin efflux transport activity in order to prevent accumulation of auxin in the cytoplasm. However, there appears to be a difference between treatment with high concentrations of exogenous auxin and manipulation of auxin transport itself. The application of picomole amounts of auxin at the shoot apex to enhance auxin flux artificially altered the expression of PIN genes, and particularly increased PIN2 expression in the root (Peer et al., 2004). Consistent with this, elevated auxin transport in mutants lacking flavonoids, endogenous auxin transport inhibitors, resulted in increased IAA levels in the root which, in turn, enhanced PIN expression and altered PIN protein localization (Peer et al., 2004).

A number of auxin transport inhibitors have been used as pharmacological agents to probe the subcellular trafficking of PIN proteins. The competitive inhibitor triiodobenzoic acid (TIBA) and the non-competitive inhibitor pyrenoyl benzoic acid (PBA) both disrupt PIN vesicle trafficking processes by interfering with actin stability (Dhonukshe et al., 2008), and TIBA can inhibit restoration of PIN1 to the plasma membrane when included in washout solutions following BFA treatment (Geldner et al., 2001). NPA can function in a similar fashion if used in even higher concentrations (Geldner et al., 2001; Petrasek et al., 2003). However, in all cases, the concentrations of auxin transport inhibitors required for inhibition of cellular trafficking mechanisms are 10–100 times higher than those required to inhibit auxin transport (Petrasek et al., 2003).

Subcellular trafficking of AUX1

The trafficking of AUX1 exhibits both differences and similarities when compared to that of PIN proteins. Although AUX1 exhibits polar localization on the apical plasma membrane in some cells, in others, AUX1 exhibits a non-polar membrane distribution and is also accumulated at the Golgi apparatus and endosomal compartments (Kleine-Vehn et al., 2006). The connections between these plasma membrane and intercellular localizations have been shown to be dependent on actin filaments and the membrane sterols (Kleine-Vehn et al., 2006). Moreover, the apical localization of AUX1 on the plasma membrane of protophloem and epidermal cells requires the presence of AUXIN RESISTANT4 (AXR4), which is found in the endoplasmic reticulum (ER) (Dharmasiri et al., 2006). In contrast to AUX1, the localizations of PIN1 and PIN2 did not appear to be affected by AXR4, thus indicating that the root agravitropic phenotype observed in the axr4 mutant is probably caused by an alteration in the AUX1 function. Although AUX1 trafficking is also BFA-sensitive, it utilizes a GNOM-independent mechanism (Kleine-Vehn et al., 2006; Boutte et al., 2007). However, this is not surprising considering the likelihood of BFA interaction with Sec7-like motifs in multiple proteins.

Subcellular trafficking of ABCBs

ABCB19 is localized on the plasma membrane where it partially overlaps with PIN1 (Blakeslee et al., 2007). However, the trafficking of ABCB19 does not coincide with that of PIN1, PIN2, and AUX1. Compared with the more dynamic processes regulating membrane localization of the AUX1 and PIN-family proteins, ABCB19 is more stably situated on the plasma membrane (Titapiwatanakun et al., 2008). Whereas the dynamic cycling of PIN1 is disrupted by short-term treatments with actin depolymerising agents like latrunculin B (Geldner et al., 2001), the localization of ABCB19 is unaffected by a short-term treatment with latrunculin B. ABCB19 is not recycled by microtubule- or SNX1-dependent processes, as ABCB19 subcellular localization is insensitive to short-term treatments with the microtubule depolymerizing compound oryzalin and is also insensitive to wortmannin (Titapiwatanakun et al., 2008). However, treatment with gravacin does interfere with the trafficking of ABCB19 to the plasma membrane, resulting in aggregation of some ABCB19 protein in an unidentified compartment that does not coincide with the endocytic marker FM4-64 (Rojas-Pierce et al., 2007). Identification of this compartment should help the elucidation of the ABCB19 trafficking pathway.

As the trafficking of ABCB19 to the plasma membrane is not BFA-sensitive, it is not mediated by GNOM-dependent mechanisms (Titapiwatanakun et al., 2008). However, not surpisingly for a P-glycoprotein, ABCB19 appears to be trafficked by GNOM-LIKE1 (GNL1), a BFA-insensitive ARF-GEF in the GNOM family that mediates vesicular ER-Golgi trafficking (Richter et al., 2007; Teh and Moore, 2007). In Arabidopsis, mutations in GNL1 exhibit a reduction in the abundance and plasma membrane localization of ABCB19, but not of PIN1 and PIN2. The gnl1 mutants also exhibited a decrease in NPA-sensitive auxin transport.

On the other hand, ABCB1 does aggregate in intracellular bodies with PIN2 after BFA treatment, suggesting that it is less stable and more readily endocytosed than ABCB19 (Blakeslee et al., 2007; Titapiwatanakun et al., 2008). This is consistent with the synergistic phenotypic effects seen with loss of ABCB1 function in abcb19 mutants and suggests that ABCB1 may be a more dynamic regulator of ABCB19 function. This is even more likely, as ABCB1 exhibits stronger interactions with both FKBP42/TWD1 and the auxin transport inhibitor NPA (Murphy et al., 2002; Geisler et al., 2003; Bouchard et al., 2006; Bailly et al., 2008).

Recently, the involvement of small GTPases, such as the Rab proteins, in the trafficking of human ABCBs has been reported in a HeLa cells study (Fu et al., 2007). The intracellular localization of human ABCB1 became conspicuous when a dominant negative form of Rab5 and GFP-tagged HsABCB1 were co-expressed in HeLa cells (Fu et al., 2007). It will be interesting to investigate further whether the small GTPase family of proteins plays a role in regulating the vesicular trafficking of the ABCBs in plants.

Regulation of auxin transporters by protein phosphorylation

Protein phosphorylation and dephosphorylation are post-translational modifications that are commonly used to regulate activity and/or function of a particular protein. The serine-threonine kinase PINOID has been shown to promote the polar localization of PIN1 and the pinoid mutant exhibits a similar phenotype to that of pin1 (Christensen et al., 2000). The apical and basal membrane localization of PIN1 were also shown to be manipulated by the overexpression and inactivation of PID (Friml et al., 2004). Therefore, PID appears to function as a binary switch that can reverse the direction of auxin movement. The activation of PID, in turn, requires phosphorylation of the protein by a 3-phosphoinositide-dependent protein kinase 1 (PDK1) (Zegzouti et al., 2006). PIN proteins also appear to be regulated by dephosphorylation catalysed by the trimeric serine-threonine protein phosphatase 2A (PP2A) (Michniewicz et al., 2007). A regulatory subunit of PP2A is encoded by the ROOT CURLING IN NPA1 (RCN1) gene. The rcn1 mutant exhibits a decrease in PP2A activity, defects in root and hypocotyl elongation, and altered apical hook formation (Garbers et al., 1996). This suggests that PP2A may play a role in NPA-sensitive root acropetal auxin transport (DeLong et al., 2002).

The linker region between the two repeated nucleotide binding domains of ABCB proteins is a prominent target for phosphorylation in yeast and humans (Ambudkar et al., 1999). In Arabidopsis, phosphoproteomics of plasma membrane proteins revealed three possible sites in ABCB proteins that can be phosphorylated by related protein kinases (Nuhse et al., 2004). One potential candidate is PHOTOTROPIN1 (PHOT1), a plasma membrane serine-threonine protein kinase that functions in multiple blue-light responses (Inoue et al., 2008). The PHOT1 protein exhibits an intracellular localization upon blue-light illumination (Briggs and Christie, 2002; Wan et al., 2008) that is concurrent with the delocalization of PIN1 in vascular tissues of photoresponding hypocotyls (Blakeslee et al., 2004). However, to date there is no evidence of direct interactions between PHOT1 and PIN1. However, ABCB19 may regulate this interaction, as ABCB19 stabilizes PIN1 on the plasma membrane (Titapiwatanakun et al., 2008), and abcb19 mutants exhibit more rapid phototropic responses in hypocotyls (Noh et al., 2003; Lin and Wang, 2005; Nagashima et al., 2008). Another potential target of PHOT1 is PIN3, which is thought to contribute to lateral translocation of auxin (Friml et al., 2002b). PIN3 localization in hypocotyl epidermal cells, whose elongation is differentially regulated in tropic bending, suggests that PHOT1 functions in inactivating localized efflux of auxin mediated by PIN3 in cells below the site of illumination. However, blue light responses mediated by PHOT1 do not appear to regulate membrane localization of ABCB19 or PIN3, as, after the blue-light treatment, no alterations in the subcellular localization of ABCB19 and PIN3 are observed, whether native, epitope-tagged, or YFP/GFP translational fusion proteins are monitored (not shown).

Regulation of auxin transporters by ubiquitin-mediated proteolysis

The steady-state levels of many proteins that are implicated in auxin transport are actively regulated by ubiquitin-mediated protein degradation. The post-transcriptional stability of PIN proteins has been shown to require the presence of MODULATOR OF PIN (MOP) proteins (Malenica et al., 2007). The Arabidopsis mop2 and mop3 mutants were identified in a genetic screen in the eir1-1 mutant background for mutations that affect auxin responses and/or distribution. In addition to a root agravitropic phenotype, the mop2 and mop3 mutants also exhibited auxin-related phenotypes similar to those seen in pin mutants. Further characterization of these mutants showed a decrease in the abundance of PIN proteins, such as PIN1, PIN2, and PIN3, suggesting that MOP played an essential role in regulating the steady-state level of PIN proteins.

Cumulative evidence suggests that PIN2 is selectively regulated by ubiquitination (Abas et al., 2006). Gravistimulation of roots results in a greater abundance of plasma membrane-localized PIN2 in epidermal cells on the underside of the root. On the upper side of the root, PIN2 was found in an intracellular compartment as well as on the plasma membrane. This internalization of PIN2 appears to be controlled by proteasomic activity, as the internalization and degradation of PIN2 was prevented by pre-treatment with the proteasome inhibitor MG132 (Abas et al., 2006).

Regulation of PIN1 by ubiquitin-mediated turnover is suggested by the presence of a lower molecular weight band (∼50 kDa) typically observed in immunoblot analyses of PIN1 that is prevented by treatment with inhibitor cocktails containing MG132 (Titapiwatanakun et al., 2008). The presence of PIN1 proteolytic products may also explain differences observed between immunolocalizations of PIN proteins, which all utilize antisera generated against the central ‘soluble’ loop of the proteins, and some PIN-fluorescent protein fusions. Proteolytic turnover may also play a role in the stabilization of PIN1 associated with ABCB19-containing membrane subdomains, as PIN1 is more resistant to degradation in those fractions.

Regulation of auxin transporters by flavonoids

The effect of NPA can be mimicked by the activity of endogenous flavonoids, which can inhibit polar auxin transport and modify local auxin concentrations (Murphy et al., 2000). In addition, flavonol compounds, such as quercetin and kaempferol, can be displaced by NPA from the microsomal extract of Arabidopsis plants. Endogenous flavonols have also been shown to regulate auxin transport negatively and alter gene expressions and subcellular localization of PIN1, PIN2, and PIN4 (Peer et al., 2004; Peer and Murphy, 2007). To date, no evidence of flavonoid regulation of AUX1 has been reported.

The relationship between flavonoids and ABCB proteins has been demonstrated, as the addition of flavonols inhibits ABCB-mediated auxin transport in heterologous expression systems (Peer and Murphy, 2007). In abcb4 and abcb19 mutants, aggregations of quercetin accumulates in the same region where ABCB4 and ABCB19 would be localized (Terasaka et al., 2005; Titapiwatanakun et al., 2008). ABCB proteins, particularly ABCB4, are likely to be the principal targets of flavonoid regulation of auxin transport at the plasma membrane. Although the flavonoid-deficient transparent testa4 tt4 mutant displayed a slower gravitropic response compared to the wild type, the gravitropic response of abcb4 tt4 double mutant resembled that of abcb4, indicating that abcb4 was epistatic to tt4 (Lewis et al., 2007).

Regulation of auxin transporters by membrane composition

The mutant lacking STEROL METHYTRANSFERASE1 (SMT1) exhibits aberrant cell polarity, auxin distribution, and embryo development (Willemsen et al., 2003). SMT1 is required for the first step of sterol biosynthesis and for the correct membrane localization of PIN1 and PIN3, but not for localization of AUX1. The loss-of-function cpi1-1 mutant, which is deficient in the cyclopropylsterol isomerase1 that catalyses a step following SMT1 in the sterol biosynthesis pathway, was also recently shown to exhibit defects in PIN2 polarity (Men et al., 2008). In normal cytokinetic epidermal cells, PIN2 is detected in both apical and basal membranes, whereas, in the post-cytokinetic epidermal cells, PIN2 is localized only at the apical membrane. However, in the cpi1-1 mutant, where PIN2 endocytosis is altered, the subcellular localization of PIN2 following cytokinesis is associated with the cell-plate-like structure. Sterols internalized from the plasma membrane colocalize with the PIN2 recycling endosome and are both BFA-sensitive and actin-dependent, suggesting a link between endocytic sterol trafficking and PIN polarity (Grebe et al., 2003). However, not all phenoytpes resulting from altered sterol trafficking are derived from altered auxin transport. A mutation in a gene encoding an Arabidopsis sterol carrier protein-2 was recently shown to regulate normal seed and seedling metabolism (Zheng et al., 2008).

A defect in myo-inositol-1-phosphate synthase (MIPS) results in an altered embryogenesis, venation patterning, and root growth. MIPS plays a crucial role in inositol biosynthesis. Thus, it has been suggested that the phosphoinositide levels of the plasma membrane can directly impact polar auxin transport. Mutants lacking MIPS showed a slower rate of endocytosis and a higher degree of wortmannin sensitivity in PIN2-positive endosomes, thereby providing evidence for a contribution of MIPS to the regulation of vesicular trafficking in plants.

Specialized microdomains that are characteristically enriched in sterol lipids and remain soluble in high concentrations of detergents have been reported (Mongrand et al., 2004; Borner et al., 2005). These microdomains have also been shown to function in the association of membrane proteins and in endocytosis (Morel et al., 2006). Sterols and sphingolipids constitute a principal factor in the formation of plasma membrane microdomains (Heese-Peck et al., 2002; Jaillais and Gaude, 2008). Although the evidence that supports the existence of plasma membrane microdomains in plants is relatively limited, the lipid composition of the detergent-resistant membrane (DRM) fraction from Arabidopsis membranes is enriched in sterols, as well as glucosylceramide. A link between sterol composition and the endocytotic pathway has also been reported (Sharma et al., 2002). More recently, functional sphingolipid synthesis and trafficking has been shown to be required for normal plant development and growth (Chen et al., 2008), suggesting that sphingolipid content may also be an important factor in regulating membrane domains.

DRM fractions derived from plant cell cultures were shown to contain ABCB and PIN proteins (Mongrand et al., 2004; Borner et al., 2005; Morel et al., 2006). Interestingly, the localization of PIN1 in DRMs requires intact ABCB19, as PIN1 was undetectable in DRM fractions derived from abcb19 seedlings and mature plants (Fig. 3; Titapiwatanakun et al., 2008). The abcb19 mutant also exhibits an altered rate of endocytosis compared to the wild type. As the DRM-associated proteins in mammals are thought to be sorted and internalized via a caveolin-dependent pathway (Sharma et al., 2002), the presence of ABCB19 in DRMs suggests that it may not only stabilize membrane microdomains, but also, indirectly, regulate endocytotic processes. Consistent with this interpretation, some BFA-induced aggregates containing PIN1 remain intact in abcb19 after BFA is washed out, suggesting that ABCB19 also has an important function in reconstitution of the endosomes to the plasma membrane (Titapiwatanakun et al., 2008).

Fig. 3.

Model depicting the interaction of ABCB19 and PIN1 in a membrane microdomain. Experimental evidence indicates that ABCB19 and PIN1 function independently to mediate auxin efflux on the plasma membrane, but co-ocurrence of ABCB19 and PIN1 synergistically enhances auxin efflux. PIN1 undergoes constitutive endocytotic cycling whereas ABCB19 appears to be relatively fixed once it is correctly inserted into the plasma membrane. However, when ABCB19 and PIN1 co-localize in sterol- and glucosylceramide-enriched microdomains, endocytosis and ubiquitin-mediated degradation of PIN1 are reduced. Retention of PIN1 in these microdomains is dependent on the presence of ABCB19.

Fig. 3.

Model depicting the interaction of ABCB19 and PIN1 in a membrane microdomain. Experimental evidence indicates that ABCB19 and PIN1 function independently to mediate auxin efflux on the plasma membrane, but co-ocurrence of ABCB19 and PIN1 synergistically enhances auxin efflux. PIN1 undergoes constitutive endocytotic cycling whereas ABCB19 appears to be relatively fixed once it is correctly inserted into the plasma membrane. However, when ABCB19 and PIN1 co-localize in sterol- and glucosylceramide-enriched microdomains, endocytosis and ubiquitin-mediated degradation of PIN1 are reduced. Retention of PIN1 in these microdomains is dependent on the presence of ABCB19.

The importance of sterols in defining the proper membrane environment may explain the difficulty encountered in attempting biochemically to demonstrate PIN1 auxin efflux activity by heterologous expression in multiple host organisms (Petrasek et al., 2006; Blakeslee et al., 2007). However, Schizosaccharomyces pombe was recently found to be suitable for the expression of PIN1 as a fully active transporter (Titapiwatanakun et al., 2008). This appears to be due to the presence of plant-like sterol-enriched microdomains in S. pombe (Wachtler et al., 2003), not found in Saccharomyces cerevisiae or Xenopus oocytes.

Conclusion

Regulation of auxin transport mechanisms is an important aspect of plant hormone biology, as impairment of auxin transport activity results in severe aberrations in plant growth and development. Plants regulate the activity and localization of these auxin transport proteins and protein complexes through a repertoire of cellular processes, many of which have now been identified. The use of Arabidopsis mutants, transgenic markers, and inducible expression constructs as well as concentration-dependent pharmalogical agents that interfere with specific proteins or enzymes have proven instrumental in deciphering these regulatory processes. While much attention has focused on their cellular trafficking pathways, the post-translational modifications of the auxin transport proteins, particularly by phosphorylation, ubiquitination, and maturation processes, are likely to be an important factor in regulating polar auxin transport and clearly need to be explored further. The role of ubiqutination and dynamic changes in the plasma membrane environment that affect the activity, localization and interaction of the auxin transport proteins deserve particular attention. The precise nature of the structural and biochemical properties that govern the interaction between auxin transport proteins and their membrane environment will require the development of new molecular genetic tools and pharmacological inhibitors to be fully elucidated.

We thank Dr Wendy Ann Peer for a careful reading of the manuscript. This work is supported by the US Department of Energy to ASM.

References

Abas
L
Benjamins
R
Malenica
N
Paciorek
T
Wisniewska
J
Moulinier-Anzola
JC
Sieberer
T
Friml
J
Luschnig
C
Intracellular trafficking and proteolysis of the Arabidopsis auxin-efflux facilitator PIN2 are involved in root gravitropism
Nature Cell Biology
 , 
2006
, vol. 
8
 (pg. 
249
-
256
)
Ambudkar
SV
Dey
S
Hrycyna
CA
Ramachandra
M
Pastan
I
Gottesman
MM
Biochemical, cellular, and pharmacological aspects of the multidrug transporter
Annual Review of Pharmacology and Toxicology
 , 
1999
, vol. 
39
 (pg. 
361
-
398
)
Ambudkar
SV
Kim
IW
Sauna
ZE
The power of the pump: mechanisms of action of P-glycoprotein (ABCB1)
European Journal of Pharmaceutical Sciences
 , 
2006
, vol. 
27
 (pg. 
392
-
400
)
Bailly
A
Sovero
V
Vincenzetti
V
Santelia
D
Bartnik
D
Koenig
BW
Mancuso
S
Martinoia
E
Geisler
M
Modulation of P-glycoproteins by auxin transport inhibitors is mediated by interaction with immunophilins
Journal of Biological Chemistry
 , 
2008
, vol. 
283
 (pg. 
21817
-
21826
)
Bainbridge
K
Guyomarc'h
S
Bayer
E
Swarup
R
Bennett
M
Mandel
T
Kuhlemeier
C
Auxin influx carriers stabilize phyllotactic patterning
Genes and Development
 , 
2008
, vol. 
22
 (pg. 
810
-
823
)
Bandyopadhyay
A
Blakeslee
JJ
Lee
OR
, et al.  . 
Interactions of PIN and PGP auxin transport mechanisms
Biochemical Society Transactions
 , 
2007
, vol. 
35
 (pg. 
137
-
141
)
Benkova
E
Michniewicz
M
Sauer
M
Teichmann
T
Seifertova
D
Jurgens
G
Friml
J
Local, efflux-dependent auxin gradients as a common module for plant organ formation
Cell
 , 
2003
, vol. 
115
 (pg. 
591
-
602
)
Bennett
MJ
Marchant
A
Green
HG
May
ST
Ward
SP
Millner
PA
Walker
AR
Schulz
B
Feldmann
KA
Arabidopsis AUX1 gene: a permease-like regulator of root gravitropism
Science
 , 
1996
, vol. 
273
 (pg. 
948
-
950
)
Bhalerao
RP
Eklof
J
Ljung
K
Marchant
A
Bennett
M
Sandberg
G
Shoot-derived auxin is essential for early lateral root emergence in Arabidopsis seedlings
The Plant Journal
 , 
2002
, vol. 
29
 (pg. 
325
-
332
)
Blakeslee
JJ
Bandyopadhyay
A
Lee
OR
, et al.  . 
Interactions among PIN-FORMED and P-glycoprotein auxin transporters in Arabidopsis
The Plant Cell
 , 
2007
, vol. 
19
 (pg. 
131
-
147
)
Blakeslee
JJ
Bandyopadhyay
A
Peer
WA
Makam
SN
Murphy
AS
Relocalization of the PIN1 auxin efflux facilitator plays a role in phototropic responses
Plant Physiology
 , 
2004
, vol. 
134
 (pg. 
28
-
31
)
Blakeslee
JJ
Peer
WA
Murphy
AS
Auxin transport
Current Opinion in Plant Biology
 , 
2005
, vol. 
8
 (pg. 
494
-
500
)
Blakeslee
JJ
Peer
WA
Murphy
AS
MDR/PGP Auxin transport proteins and endocytic cycling
Plant Cell Monographs: Plant Endocytosis
 , 
2005
(pg. 
159
-
176
)
Blilou
I
Xu
J
Wildwater
M
Willemsen
V
Paponov
I
Friml
J
Heidstra
R
Aida
M
Palme
K
Scheres
B
The PIN auxin efflux facilitator network controls growth and patterning in Arabidopsis roots
Nature
 , 
2005
, vol. 
433
 (pg. 
39
-
44
)
Bonifacino
JS
Rojas
R
Retrograde transport from endosomes to the trans-Golgi network
Nature Reviews Molecular and Cellular Biology
 , 
2006
, vol. 
7
 (pg. 
568
-
579
)
Borner
GH
Sherrier
DJ
Weimar
T
Michaelson
LV
Hawkins
ND
Macaskill
A
Napier
JA
Beale
MH
Lilley
KS
Dupree
P
Analysis of detergent-resistant membranes in Arabidopsis. Evidence for plasma membrane lipid rafts
Plant Physiology
 , 
2005
, vol. 
137
 (pg. 
104
-
116
)
Bouchard
R
Bailly
A
Blakeslee
JJ
, et al.  . 
Immunophilin-like TWISTED DWARF1 modulates auxin efflux activities of Arabidopsis P-glycoproteins
Journal of Biological Chemistry
 , 
2006
, vol. 
281
 (pg. 
30603
-
30612
)
Boutte
Y
Ikeda
Y
Grebe
M
Mechanisms of auxin-dependent cell and tissue polarity
Current Opinion in Plant Biology
 , 
2007
, vol. 
10
 (pg. 
616
-
623
)
Briggs
WR
Christie
JM
Phototropins 1 and 2: versatile plant blue-light receptors
Trends in Plant Science
 , 
2002
, vol. 
7
 (pg. 
204
-
210
)
Campanoni
P
Nick
P
Auxin-dependent cell division and cell elongation. 1-Naphthaleneacetic acid and 2,4-dichlorophenoxyacetic acid activate different pathways
Plant Physiology
 , 
2005
, vol. 
137
 (pg. 
939
-
948
)
Casimiro
I
Marchant
A
Bhalerao
RP
, et al.  . 
Auxin transport promotes Arabidopsis lateral root initiation
The Plant Cell
 , 
2001
, vol. 
13
 (pg. 
843
-
852
)
Chen
R
Hilson
P
Sedbrook
J
Rosen
E
Caspar
T
Masson
PH
The Arabidopsis thaliana AGRAVITROPIC 1 gene encodes a component of the polar-auxin-transport efflux carrier
Proceedings of the National Academy of Sciences, USA
 , 
1998
, vol. 
95
 (pg. 
15112
-
15117
)
Chen
M
Markham
JE
Dietrich
CR
Jaworski
JG
Cahoon
EB
Sphingolipid long-chain base hydroxylation is important for growth and regulation of sphingolipid content and composition in Arabidopsis
Plant Cell
 , 
2008
, vol. 
20
 (pg. 
1862
-
78
)
Cho
M
Lee
SH
Cho
HT
P-glycoprotein4 displays auxin efflux transporter-like action in Arabidopsis root hair cells and tobacco cells
The Plant Cell
 , 
2007
, vol. 
19
 (pg. 
3930
-
3943
)
Christensen
SK
Dagenais
N
Chory
J
Weigel
D
Regulation of auxin response by the protein kinase PINOID
Cell
 , 
2000
, vol. 
100
 (pg. 
469
-
478
)
de Billy
F
Grosjean
C
May
S
Bennett
M
Cullimore
JV
Expression studies on AUX1-like genes in Medicago truncatula suggest that auxin is required at two steps in early nodule development
Molecular Plant–Microbe Interactions
 , 
2001
, vol. 
14
 (pg. 
267
-
277
)
De Smet
I
Jurgens
G
Patterning the axis in plants: auxin in control
Current Opinion in Genetic Development
 , 
2007
, vol. 
17
 (pg. 
337
-
343
)
De Smet
I
Tetsumura
T
De Rybel
B
, et al.  . 
Auxin-dependent regulation of lateral root positioning in the basal meristem of Arabidopsis
Development
 , 
2007
, vol. 
134
 (pg. 
681
-
690
)
Delker
C
Raschke
A
Quint
M
Auxin dynamics: the dazzling complexity of a small molecule's message
Planta
 , 
2008
, vol. 
227
 (pg. 
929
-
941
)
DeLong
A
Mockaitis
K
Christensen
S
Protein phosphorylation in the delivery of and response to auxin signals
Plant Molecular Biology
 , 
2002
, vol. 
49
 (pg. 
285
-
303
)
Dharmasiri
S
Swarup
R
Mockaitis
K
, et al.  . 
AXR4 is required for localization of the auxin influx facilitator AUX1
Science
 , 
2006
, vol. 
312
 (pg. 
1218
-
1220
)
Dhonukshe
P
Grigoriev
I
Fischer
R
, et al.  . 
Auxin transport inhibitors impair vesicle motility and actin cytoskeleton dynamics in diverse eukaryotes
Proceedings of the National Academy of Sciences, USA
 , 
2008
, vol. 
105
 (pg. 
4489
-
4494
)
Dubrovsky
JG
Sauer
M
Napsucialy-Mendivil
S
Ivanchenko
MG
Friml
J
Shishkova
S
Celenza
J
Benkova
E
Auxin acts as a local morphogenetic trigger to specify lateral root founder cells
Proceedings of the National Academy of Sciences, USA
 , 
2008
, vol. 
105
 (pg. 
8790
-
8794
)
Eckhoff
A
Granzin
J
Kamphausen
T
Buldt
G
Schulz
B
Weiergraber
OH
Crystallization and preliminary X-ray analysis of immunophilin-like FKBP42 from Arabidopsis thaliana
Acta Crystallogrophy Section F Structural Biology and Crystallization Communications
 , 
2005
, vol. 
61
 (pg. 
363
-
365
)
Friml
J
Benkova
E
Blilou
I
, et al.  . 
AtPIN4 mediates sink-driven auxin gradients and root patterning in Arabidopsis
Cell
 , 
2002
, vol. 
108
 (pg. 
661
-
673
)
Friml
J
Vieten
A
Sauer
M
Weijers
D
Schwarz
H
Hamann
T
Offringa
R
Jurgens
G
Efflux-dependent auxin gradients establish the apical-basal axis of Arabidopsis
Nature
 , 
2003
, vol. 
426
 (pg. 
147
-
153
)
Friml
J
Wisniewska
J
Benkova
E
Mendgen
K
Palme
K
Lateral relocation of auxin efflux regulator PIN3 mediates tropism in Arabidopsis
Nature
 , 
2002
, vol. 
415
 (pg. 
806
-
809
)
Friml
J
Yang
X
Michniewicz
M
, et al.  . 
A PINOID-dependent binary switch in apical-basal PIN polar targeting directs auxin efflux
Science
 , 
2004
, vol. 
306
 (pg. 
862
-
865
)
Fu
D
van Dam
EM
Brymora
A
Duggin
IG
Robinson
PJ
Roufogalis
BD
The small GTPases Rab5 and RalA regulate intracellular traffic of P-glycoprotein
Biochimica et Biophysica Acta
 , 
2007
, vol. 
1773
 (pg. 
1062
-
1072
)
Galweiler
L
Guan
C
Muller
A
Wisman
E
Mendgen
K
Yephremov
A
Palme
K
Regulation of polar auxin transport by AtPIN1 in Arabidopsis vascular tissue
Science
 , 
1998
, vol. 
282
 (pg. 
2226
-
2230
)
Garbers
C
DeLong
A
Deruere
J
Bernasconi
P
Soll
D
A mutation in protein phosphatase 2A regulatory subunit A affects auxin transport in Arabidopsis
EMBO Journal
 , 
1996
, vol. 
15
 (pg. 
2115
-
2124
)
Geisler
M
Kolukisaoglu
HU
Bouchard
R
, et al.  . 
TWISTED DWARF1, a unique plasma membrane-anchored immunophilin-like protein, interacts with Arabidopsis multidrug resistance-like transporters AtPGP1 and AtPGP19
Molecular Biology of the Cell
 , 
2003
, vol. 
14
 (pg. 
4238
-
4249
)
Geisler
M
Blakeslee
JJ
Bouchard
R
, et al.  . 
Cellular efflux of auxin catalysed by the Arabidopsis MDR/PGP transporter AtPGP1
The Plant Journal
 , 
2005
, vol. 
44
 (pg. 
179
-
194
)
Geldner
N
Friml
J
Stierhof
YD
Jurgens
G
Palme
K
Auxin transport inhibitors block PIN1 cycling and vesicle trafficking
Nature
 , 
2001
, vol. 
413
 (pg. 
425
-
428
)
Geldner
N
Anders
N
Wolters
H
Keicher
J
Kornberger
W
Muller
P
Delbarre
A
Ueda
T
Nakano
A
Jurgens
G
The Arabidopsis GNOM ARF-GEF mediates endosomal recycling, auxin transport, and auxin-dependent plant growth
Cell
 , 
2003
, vol. 
112
 (pg. 
219
-
230
)
Granzin
J
Eckhoff
A
Weiergraber
OH
Crystal structure of a multi-domain immunophilin from Arabidopsis thaliana: a paradigm for regulation of plant ABC transporters
Journal of Molecular Biology
 , 
2006
, vol. 
364
 (pg. 
799
-
809
)
Grebe
M
Xu
J
Mobius
W
Ueda
T
Nakano
A
Geuze
HJ
Rook
MB
Scheres
B
Arabidopsis sterol endocytosis involves actin-mediated trafficking via ARA6-positive early endosomes
Current Biology
 , 
2003
, vol. 
13
 (pg. 
1378
-
1387
)
Heese-Peck
A
Pichler
H
Zanolari
B
Watanabe
R
Daum
G
Riezman
H
Multiple functions of sterols in yeast endocytosis
Molecular Biology of the Cell
 , 
2002
, vol. 
13
 (pg. 
2664
-
2680
)
Inoue
S
Kinoshita
T
Matsumoto
M
Nakayama
KI
Doi
M
Shimazaki
K
Blue light-induced autophosphorylation of phototropin is a primary step for signaling
Proceedings of the National Academy of Sciences, USA
 , 
2008
, vol. 
105
 (pg. 
5626
-
5631
)
Jaillais
Y
Fobis-Loisy
I
Miege
C
Rollin
C
Gaude
T
AtSNX1 defines an endosome for auxin-carrier trafficking in Arabidopsis
Nature
 , 
2006
, vol. 
443
 (pg. 
106
-
109
)
Jaillais
Y
Gaude
T
Plant cell polarity: sterols enter into action after cytokinesis
Developmental Cell
 , 
2008
, vol. 
14
 (pg. 
318
-
320
)
Jaillais
Y
Santambrogio
M
Rozier
F
Fobis-Loisy
I
Miege
C
Gaude
T
The retromer protein VPS29 links cell polarity and organ initiation in plants
Cell
 , 
2007
, vol. 
130
 (pg. 
1057
-
1070
)
Kepinski
S
The anatomy of auxin perception
Bioessays
 , 
2007
, vol. 
29
 (pg. 
953
-
956
)
Kerr
ID
Bennett
MJ
New insight into the biochemical mechanisms regulating auxin transport in plants
Biochemistry Journal
 , 
2007
, vol. 
401
 (pg. 
613
-
622
)
Kimura
M
Kagawa
T
Phototropin and light-signaling in phototropism
Current Opinion in Plant Biology
 , 
2006
, vol. 
9
 (pg. 
503
-
508
)
Kimura
Y
Morita
SY
Matsuo
M
Ueda
K
Mechanism of multidrug recognition by MDR1/ABCB1
Cancer Science
 , 
2007
, vol. 
98
 (pg. 
1303
-
1310
)
Kleine-Vehn
J
Dhonukshe
P
Sauer
M
Brewer
PB
Wisniewska
J
Paciorek
T
Benkova
E
Friml
J
ARF GEF-dependent transcytosis and polar delivery of PIN auxin carriers in Arabidopsis
Current Biology
 , 
2008
, vol. 
18
 (pg. 
526
-
531
)
Kleine-Vehn
J
Dhonukshe
P
Swarup
R
Bennett
M
Friml
J
Subcellular trafficking of the Arabidopsis auxin influx carrier AUX1 uses a novel pathway distinct from PIN1
The Plant Cell
 , 
2006
, vol. 
18
 (pg. 
3171
-
3181
)
Kramer
EM
Bennett
MJ
Auxin transport: a field in flux
Trends in Plant Science
 , 
2006
, vol. 
11
 (pg. 
382
-
386
)
Lee
M
Choi
Y
Burla
B
Kim
Y-Y
Jeon
B
Maeshima
M
Yoo
J-Y
Martinoia
E
Lee
Y
The ABC transporter AtABCB14 is a malate importer and modulates stomatal response to CO2
Nature Cell Biology
 , 
2008
 
doi:10.1038/ncb1782
Lewis
DR
Miller
ND
Splitt
BL
Wu
G
Spalding
EP
Separating the roles of acropetal and basipetal auxin transport on gravitropism with mutations in two Arabidopsis multidrug resistance-like ABC transporter genes
The Plant Cell
 , 
2007
, vol. 
19
 (pg. 
1838
-
1850
)
Lin
R
Wang
H
Two homologous ATP-binding cassette transporter proteins, AtMDR1 and AtPGP1, regulate Arabidopsis photomorphogenesis and root development by mediating polar auxin transport
Plant Physiology
 , 
2005
, vol. 
138
 (pg. 
949
-
964
)
Ljung
K
Hull
AK
Celenza
J
Yamada
M
Estelle
M
Normanly
J
Sandberg
G
Sites and regulation of auxin biosynthesis in Arabidopsis roots
The Plant Cell
 , 
2005
, vol. 
17
 (pg. 
1090
-
1104
)
Luckie
DB
Wilterding
JH
Krha
M
Krouse
ME
CFTR and MDR: ABC transporters with homologous structure but divergent function
Current Genomics
 , 
2003
, vol. 
4
 (pg. 
225
-
235
)
Luschnig
C
Gaxiola
RA
Grisafi
P
Fink
GR
EIR1, a root-specific protein involved in auxin transport, is required for gravitropism in Arabidopsis thaliana
Genes and Development
 , 
1998
, vol. 
12
 (pg. 
2175
-
2187
)
Malenica
N
Abas
L
Benjamins
R
Kitakura
S
Sigmund
HF
Jun
KS
Hauser
MT
Friml
J
Luschnig
C
MODULATOR OF PIN genes control steady-state levels of Arabidopsis PIN proteins
The Plant Journal
 , 
2007
, vol. 
51
 (pg. 
537
-
550
)
Marchant
A
Kargul
J
May
ST
Muller
P
Delbarre
A
Perrot-Rechenmann
C
Bennett
MJ
AUX1 regulates root gravitropism in Arabidopsis by facilitating auxin uptake within root apical tissues
EMBO Journal
 , 
1999
, vol. 
18
 (pg. 
2066
-
2073
)
Mattsson
J
Ckurshumova
W
Berleth
T
Auxin signaling in Arabidopsis leaf vascular development
Plant Physiology
 , 
2003
, vol. 
131
 (pg. 
1327
-
1339
)
Men
S
Boutte
Y
Ikeda
Y
Li
X
Palme
K
Stierhof
YD
Hartmann
MA
Moritz
T
Grebe
M
Sterol-dependent endocytosis mediates post-cytokinetic acquisition of PIN2 auxin efflux carrier polarity
Nature Cell Biology
 , 
2008
, vol. 
10
 (pg. 
237
-
244
)
Michniewicz
M
Zago
MK
Abas
L
, et al.  . 
Antagonistic regulation of PIN phosphorylation by PP2A and PINOID directs auxin flux
Cell
 , 
2007
, vol. 
130
 (pg. 
1044
-
1056
)
Mongrand
S
Morel
J
Laroche
J
Claverol
S
Carde
JP
Hartmann
MA
Bonneu
M
Simon-Plas
F
Lessire
R
Bessoule
JJ
Lipid rafts in higher plant cells: purification and characterization of Triton X-100-insoluble microdomains from tobacco plasma membrane
Journal of Biological Chemistry
 , 
2004
, vol. 
279
 (pg. 
36277
-
36286
)
Morel
J
Claverol
S
Mongrand
S
Furt
F
Fromentin
J
Bessoule
JJ
Blein
JP
Simon-Plas
F
Proteomics of plant detergent-resistant membranes
Molecular Cell Proteomics
 , 
2006
, vol. 
5
 (pg. 
1396
-
1411
)
Muller
A
Guan
C
Galweiler
L
Tanzler
P
Huijser
P
Marchant
A
Parry
G
Bennett
M
Wisman
E
Palme
K
AtPIN2 defines a locus of Arabidopsis for root gravitropism control
EMBO Journal
 , 
1998
, vol. 
17
 (pg. 
6903
-
6911
)
Multani
DS
Briggs
SP
Chamberlin
MA
Blakeslee
JJ
Murphy
AS
Johal
GS
Loss of an MDR transporter in compact stalks of maize br2 and sorghum dw3 mutants
Science
 , 
2003
, vol. 
302
 (pg. 
81
-
84
)
Murphy
A
Peer
WA
Taiz
L
Regulation of auxin transport by aminopeptidases and endogenous flavonoids
Planta
 , 
2000
, vol. 
211
 (pg. 
315
-
324
)
Murphy
AS
Hoogner
KR
Peer
WA
Taiz
L
Identification, purification, and molecular cloning of N-1-naphthylphthalmic acid-binding plasma membrane-associated aminopeptidases from Arabidopsis
Plant Physiology
 , 
2002
, vol. 
128
 (pg. 
935
-
950
)
Nagashima
A
Suzuki
G
Uehara
Y
, et al.  . 
Phytochromes and cryptochromes regulate the differential growth of Arabidopsis hypocotyls in both a PGP19-dependent and a PGP19-independent manner
The Plant Journal
 , 
2008
, vol. 
53
 (pg. 
516
-
529
)
Noh
B
Bandyopadhyay
A
Peer
WA
Spalding
EP
Murphy
AS
Enhanced gravi- and phototropism in plant mdr mutants mislocalizing the auxin efflux protein PIN1
Nature
 , 
2003
, vol. 
423
 (pg. 
999
-
1002
)
Noh
B
Murphy
AS
Spalding
EP
Multidrug resistance-like genes of Arabidopsis required for auxin transport and auxin-mediated development
The Plant Cell
 , 
2001
, vol. 
13
 (pg. 
2441
-
2454
)
Nuhse
TS
Stensballe
A
Jensen
ON
Peck
SC
Phosphoproteomics of the Arabidopsis plasma membrane and a new phosphorylation site database
The Plant Cell
 , 
2004
, vol. 
16
 (pg. 
2394
-
2405
)
Paciorek
T
Zazimalova
E
Ruthardt
N
, et al.  . 
Auxin inhibits endocytosis and promotes its own efflux from cells
Nature
 , 
2005
, vol. 
435
 (pg. 
1251
-
1256
)
Palme
K
Dovzhenko
A
Ditengou
FA
Auxin transport and gravitational research: perspectives
Protoplasma
 , 
2006
, vol. 
229
 (pg. 
175
-
181
)
Peer
WA
Bandyopadhyay
A
Blakeslee
JJ
Makam
SN
Chen
RJ
Masson
PH
Murphy
AS
Variation in expression and protein localization of the PIN family of auxin efflux facilitator proteins in flavonoid mutants with altered auxin transport in Arabidopsis thaliana
The Plant Cell
 , 
2004
, vol. 
16
 (pg. 
1898
-
1911
)
Peer
WA
Murphy
AS
Flavonoids and auxin transport: modulators or regulators?
Trends in Plant Science
 , 
2007
, vol. 
12
 (pg. 
556
-
563
)
Petrasek
J
Cerna
A
Schwarzerova
K
Elckner
M
Morris
DA
Zazimalova
E
Do phytotropins inhibit auxin efflux by impairing vesicle traffic?
Plant Physiology
 , 
2003
, vol. 
131
 (pg. 
254
-
263
)
Petrasek
J
Mravec
J
Bouchard
R
, et al.  . 
PIN proteins perform a rate-limiting function in cellular auxin efflux
Science
 , 
2006
, vol. 
312
 (pg. 
914
-
918
)
Quint
M
Gray
WM
Auxin signaling
Current Opinion in Plant Biology
 , 
2006
, vol. 
9
 (pg. 
448
-
453
)
Reinhardt
D
Phyllotaxis: a new chapter in an old tale about beauty and magic numbers
Current Opinion in Plant Biology
 , 
2005
, vol. 
8
 (pg. 
487
-
493
)
Richter
S
Geldner
N
Schrader
J
Wolters
H
Stierhof
YD
Rios
G
Koncz
C
Robinson
DG
Jurgens
G
Functional diversification of closely related ARF-GEFs in protein secretion and recycling
Nature
 , 
2007
, vol. 
448
 (pg. 
488
-
492
)
Rojas-Pierce
M
Titapiwatanakun
B
Sohn
EJ
, et al.  . 
Arabidopsis P-glycoprotein19 participates in the inhibition of gravitropism by gravacin
Chemistry and Biology
 , 
2007
, vol. 
14
 (pg. 
1366
-
1376
)
Sachs
T
The control of the patterned differentiation of vascular tissues
Advances in Botanical Research Incorporating Advances in Plant Pathology
 , 
1981
, vol. 
9
 (pg. 
151
-
262
)
Santelia
D
Vincenzetti
V
Azzarello
E
Bovet
L
Fukao
Y
Duchtig
P
Mancuso
S
Martinoia
E
Geisler
M
MDR-like ABC transporter AtPGP4 is involved in auxin-mediated lateral root and root hair development
FEBS Letters
 , 
2005
, vol. 
579
 (pg. 
5399
-
5406
)
Sauer
M
Balla
J
Luschnig
C
Wisniewska
J
Reinohl
V
Friml
J
Benkova
E
Canalization of auxin flow by Aux/IAA-ARF-dependent feedback regulation of PIN polarity
Genes and Development
 , 
2006
, vol. 
20
 (pg. 
2902
-
2911
)
Sharma
P
Sabharanjak
S
Mayor
S
Endocytosis of lipid rafts: an identity crisis
Seminars in Cell Development Biology
 , 
2002
, vol. 
13
 (pg. 
205
-
214
)
Shitan
N
Bazin
I
Dan
K
Obata
K
Kigawa
K
Ueda
K
Sato
F
Forestier
C
Yazaki
K
Involvement of CjMDR1, a plant multidrug-resistance-type ATP-binding cassette protein, in alkaloid transport in Coptis japonica
Proceedings of the National Academy of Sciences, USA
 , 
2003
, vol. 
100
 (pg. 
751
-
756
)
Sidler
M
Hassa
P
Hasan
S
Ringli
C
Dudler
R
Involvement of an ABC transporter in a developmental pathway regulating hypocotyl cell elongation in the light
The Plant Cell
 , 
1998
, vol. 
10
 (pg. 
1623
-
1636
)
Sieburth
LE
Muday
GK
King
EJ
Benton
G
Kim
S
Metcalf
KE
Meyers
L
Seamen
E
Van Norman
JM
SCARFACE encodes an ARF-GAP that is required for normal auxin efflux and vein patterning in Arabidopsis
The Plant Cell
 , 
2006
, vol. 
18
 (pg. 
1396
-
1411
)
Steinmann
T
Geldner
N
Grebe
M
Mangold
S
Jackson
CL
Paris
S
Galweiler
L
Palme
K
Jurgens
G
Co-ordinated polar localization of auxin efflux carrier PIN1 by GNOM ARF GEF
Science
 , 
1999
, vol. 
286
 (pg. 
316
-
318
)
Surpin
M
Rojas-Pierce
M
Carter
C
Hicks
GR
Vasquez
J
Raikhel
NV
The power of chemical genomics to study the link between endomembrane system components and the gravitropic response
Proceedings of the National Academy of Sciences, USA
 , 
2005
, vol. 
102
 (pg. 
4902
-
4907
)
Swarup
K
Benkova
E
Swarup
R
, et al.  . 
The auxin influx carrier LAX3 promotes lateral root emergence
Nature Cell Biology
 , 
2008
, vol. 
10
 (pg. 
946
-
954
)
Swarup
R
Parry
G
Graham
N
Allen
T
Bennett
M
Auxin cross-talk: integration of signalling pathways to control plant development
Plant Molecular Biology
 , 
2002
, vol. 
49
 (pg. 
411
-
426
)
Swarup
R
Kargul
J
Marchant
A
, et al.  . 
Structure–function analysis of the presumptive Arabidopsis auxin permease AUX1
The Plant Cell
 , 
2004
, vol. 
16
 (pg. 
3069
-
3083
)
Tanaka
H
Dhonukshe
P
Brewer
PB
Friml
J
Spatiotemporal asymmetric auxin distribution: a means to co-ordinate plant development
Cellular and Molecular Life Sciences
 , 
2006
, vol. 
63
 (pg. 
2738
-
2754
)
Teh
OK
Moore
I
An ARF-GEF acting at the Golgi and in selective endocytosis in polarized plant cells
Nature
 , 
2007
, vol. 
448
 (pg. 
493
-
496
)
Terasaka
K
Blakeslee
JJ
Titapiwatanakun
B
, et al.  . 
PGP4, an ATP binding cassette P-glycoprotein, catalyses auxin transport in Arabidopsis thaliana roots
The Plant Cell
 , 
2005
, vol. 
17
 (pg. 
2922
-
2939
)
Timpte
C
Auxin binding protein: curiouser and curiouser
Trends in Plant Science
 , 
2001
, vol. 
6
 (pg. 
586
-
590
)
Titapiwatanakun
B
Blakeslee
J
Bandyopadhyay
A
, et al.  . 
ABCB19/PGP19 stabilizes PIN1 on membrane microdomains in Arabidopsis
The Plant Journal
 , 
2008
 
(in press)
Verrier
PJ
Bird
D
Burla
B
, et al.  . 
Plant ABC proteins: a unified nomenclature and updated inventory
Trends in Plant Science
 , 
2008
, vol. 
13
 (pg. 
151
-
159
)
Vieten
A
Vanneste
S
Wisniewska
J
Benkova
E
Benjamins
R
Beeckman
T
Luschnig
C
Friml
J
Functional redundancy of PIN proteins is accompanied by auxin-dependent cross-regulation of PIN expression
Development
 , 
2005
, vol. 
132
 (pg. 
4521
-
4531
)
Wachtler
V
Rajagopalan
S
Balasubramanian
MK
Sterol-rich plasma membrane domains in the fission yeast Schizosaccharomyces pombe
Journal of Cell Science
 , 
2003
, vol. 
116
 (pg. 
867
-
874
)
Wan
YL
Eisinger
W
Ehrhardt
D
Kubitscheck
U
Baluska
F
Briggs
W
The subcellular localization and blue-light-induced movement of phototropin 1-GFP in etiolated seedlings of Arabidopsis thaliana
Molecular Plant
 , 
2008
, vol. 
1
 (pg. 
103
-
117
)
Went
FW
On growth-accelerating substances in the coleoptile of Avena sativa
Proceedings of the Koninklijke Akademie Van Wetenschappen Te Amsterdam
 , 
1927
, vol. 
30
 (pg. 
10
-
19
)
Willemsen
V
Friml
J
Grebe
M
van den Toorn
A
Palme
K
Scheres
B
Cell polarity and PIN protein positioning in Arabidopsis require STEROL METHYLTRANSFERASE1 function
The Plant Cell
 , 
2003
, vol. 
15
 (pg. 
612
-
625
)
Wisniewska
J
Xu
J
Seifertova
D
Brewer
PB
Ruzicka
K
Blilou
I
Rouquie
D
Benkova
E
Scheres
B
Friml
J
Polar PIN localization directs auxin flow in plants
Science
 , 
2006
, vol. 
312
 pg. 
883
 
Wu
G
Lewis
DR
Spalding
EP
Mutations in Arabidopsis multidrug resistance-like ABC transporters separate the roles of acropetal and basipetal auxin transport in lateral root development
The Plant Cell
 , 
2007
, vol. 
19
 (pg. 
1826
-
1837
)
Yang
Y
Hammes
UZ
Taylor
CG
Schachtman
DP
Nielsen
E
High-affinity auxin transport by the AUX1 influx carrier protein
Current Biology
 , 
2006
, vol. 
16
 (pg. 
1123
-
1127
)
Zazimalova
E
Krecek
P
Skupa
P
Hoyerova
K
Petrasek
J
Polar transport of the plant hormone auxin: the role of PIN-FORMED (PIN) proteins
Cellular and Molecular Life Sciences
 , 
2007
, vol. 
64
 (pg. 
1621
-
1637
)
Zegzouti
H
Anthony
RG
Jahchan
N
Bogre
L
Christensen
SK
Phosphorylation and activation of PINOID by the phospholipid signaling kinase 3-phosphoinositide-dependent protein kinase 1 (PDK1) in Arabidopsis
Proceedings of the National Academy of Sciences, USA
 , 
2006
, vol. 
103
 (pg. 
6404
-
6409
)
Zheng
BS
Ronnberg
E
Viltanen
L
Salminen
TA
Lundgren
K
Mortiz
T
Edqvist
J
Arabidopsis sterol carrier protein-2 is required for normal development of seeds and seedlings
Journal of Experimental Botany
 , 
2008
 
doi:10.1093/jxb/ern201

Comments

0 Comments