Strigolactones (SLs) are phytohormones that regulate various developmental processes inside the plant body, including shoot branching and root growth, cause the germination of seeds from root-parasitic plants, and initiate symbiosis between plants and arbuscular mycorrhizal fungi. From an evolutionary standpoint, SLs are very old molecules whose roles in symbiosis might have helped plants colonize land approximately 450 million years ago. Bryophytes such as the moss Physcomitrium patens were among the first plants to appear on land and are excellent model systems to study the evolution of SL signaling. In angiosperms, SLs are perceived by the α/β hydrolase DWARF 14 (D14). D14 is structurally similar to KARRIKIN INSENSITIVE 2 (KAI2), which is mostly involved in seed germination and seedling development. KAI2’s endogenous signal is still unknown and is commonly referred to as KAI2 ligand (KL).
In a new study, Mauricio Lopez-Obando, Ambre Guillory, and colleagues (Lopez-Obando et al., 2021) used a combination of P. patens mutants, pharmacological treatments, in vitro binding assays, and cross-species complementation to dissect the proteins involved in SL versus KL signaling and perception in this moss (see Figure). Physcomitrium patens has undergone an expansion of the KAI2 receptor family to 13 members (PpKAI2L-A to M), classified in four clades: (ABCDE; also called eu-KAI2), (FK), (HIL), and (GJM). The authors created single and higher-order P. patens mutants to characterize these clades. When they abolished the (GJM) clade, the phenotype was reminiscent of the SL biosynthesis mutant Ppccd8 (Proust et al., 2011), and when they eliminated the eu-KAI2 clade, the phenotype was similar to the F-box signaling mutant Ppmax2, which remains SL sensitive (Lopez-Obando et al., 2018). Therefore, in P. patens, distinct clades of the PpKAI2L protein family perceive PpCCD8-derived SLs versus KL(s), with PpMAX2 taking part in KL signaling. Next, the authors applied different stereoisomers of the chemical analog GR24. The (+)-GR24 isomer is a well-established SL mimic. Interestingly, the (–)-GR24 isomer does not act like SL but instead binds to KAI2 and is considered to be a mimic of the unknown KL. Strikingly, the (gjm) mutant no longer responded to exogenous (+)-GR24, confirming that members of this clade indeed perceive SLs. Treatments with (–)-GR24 showed that the quintuple eu-KAI2 mutant still perceived (–)-GR24, suggesting that other proteins receive this signal as well.
Different pathways control protonema growth and gametophore development in P. patens. A, The weak KL mimic (–)-GR24 signals through the (eu-KAI2) clade of KAI2L proteins, whereas the SL mimic (+)-GR24 works via the (GJM) clade. Arrow width indicates effect strength. Modified from Lopez-Obando et al. (2021). B, Phylogenetic tree of PpKAI2L protein sequences. Created with BioRender.com.
Although (–)-GR24 appears to be a rather poor mimic for the actual P. patens KL(s), the authors elegantly complemented the mutant studies with biochemical and mass spectrometry analyses that demonstrated binding of (–)-GR24 to members of the eu-KAI2 clade and the diverging preferences of different clades for the different GR24 stereoisomers. By combining these approaches, they illustrate the existence of two pathways regulating the balance of protonema versus gametophore development in P. patens, triggered by KL(s) and SLs, respectively. Other exciting results reported in this paper should prompt future investigation, such as a possible gatekeeper residue in SL receptors causing their low turnover rates, and a potential interaction of eu-KAI2 clade proteins with the Arabidopsis thaliana KAI2 pathway.
The work by Lopez-Obando et al. is an important expansion of a previous study describing the structural mechanism for binding of non-natural SLs by members of the P. patens eu-KAI2 clade (Bürger et al., 2019). The identification of protein clades specific for SLs or KL(s) constitutes a significant advancement in our understanding of SL signaling evolution and the developmental biology of P. patens. How exactly the signal progresses downstream of the (GJM) clade remains to be established, since it would likely require a different F-box protein. Finally, the identity of the endogenous KL molecule(s) in P. patens and in fact any plant remains a mystery and will certainly be a focus of future research. The importance of KL signaling for plant developmental processes would make the identification of KL, which might be a new plant hormone, an important discovery for plant biology and agriculture. Desmethyl butenolides were recently identified as possible KAI2 ligands, and they are certainly among the candidates worth testing.
