Origin of a Species-Specific Rheostat Controlling Testicular Growth and Steroidogenesis.

The prolactin (PRL) family of hormones/cytokines participates in the regulation of reproduction. Mouse mutagenesis has been used to characterize the biology of a subset of PRL family ligands. To date, PRL family members have been implicated in female reproduction. In this study, we used mouse mutagenesis to investigate the biology of PRL family 3, subfamily c, member 1 (PRL3C1), a heparin binding decidual cell cytokine previously implicated in the establishment of pregnancy. Male and female mice possessing a Prl3c1 null mutant allele on a C57BL/6 genetic background exhibited severe subfertility. Moving the Prl3c1 mutation to a mixed genetic background (C57BL/6 X CD1) improved fertility. Reproductive abnormalities remained in females and males possessing the Prl3c1 null mutation. In the remainder of this study, we focused on the unexpected male reproductive phenotype. Relative testis and seminal vesicle weights were increased in Prl3c1 null versus wild type (WT) mice (P<0.005 for both tissues). Consistent with the hypertrophy of the androgen responsive seminal vesicles, serum testosterone and LH levels were also elevated in the Prl3c1 null versus WT mice (P<0.001). These observations prompted an assessment of Prl3c1 expression in the male reproductive tract. Prl3c1 transcripts were identified in the testis, localized to Leydig cells, and showed a dramatic developmental increase between postnatal days 15 and 21, correlating with an increase in the expression of the inducible 3-beta hydroxysteroid dehydrogenase isoform, Hsd3b6. In Prl3c1 null testis, Hsd3b6 transcript levels were decreased (P<0.001), whereas, transcript levels for the constitutive isoform, Hsd3b1, were increased (P<0.001). 5-alpha reductase transcripts (Srd5a2, Srd5a3) were also decreased in the Prl3c1 null versus WT mouse testes (P<0.01). These differences in the expression of genes encoding proteins involved in steroid metabolism may in part explain the hyperandrogenic phenotype of the Prl3c1 null males. Upon closer examination, the 5' portion of the Prl3c1 transcript expressed by decidua (V1) differed from the transcript expressed by Leydig cells (V2), resulting in the transformation of a transcript (V1) encoding a secreted protein to a transcript (V2) encoding an intracellular protein. A mouse-specific transposable element (RMER17D) is situated within the first intron of the mouse Prl3c1 gene, immediately upstream of the first exon contributing to the V2 transcript. The presence of a transposable element at this site was not conserved in the rat nor was Prl3c1 expression detected in the rat testis. The Prl3c1-associated transposable element exhibited differential DNA methylation status. In decidua, CpG islands within the intronic transposable element were hyper-methylated, whereas the converse was true in Leydig cells. Taken together we have discovered a novel species-specific rheostat controlling testicular growth and steroidogenesis. The experimental evidence suggests that the evolutionary origin of this regulatory mechanism can be traced to the insertion of a transposable element into the first intron of the mouse Prl3c1 gene. This event resulted in the expression of a unique Prl3c1 transcript at a unique site, encoding for a protein with unique biological properties controlling testicular function. (Supported by NIH HD20676)