Nuclear progestin receptor (Pgr) knockouts resulted in subfertility in male tilapia (Oreochromis niloticus)

It was documented that 17 alpha, 20 beta-dihydroxy-4-pregnen-3-one (DHP), a fish specific progestin, might play critical roles in spermatogenesis, sperm maturation and spermiation partially through activating nuclear receptor (Pgr). However, no direct evidence is available to demonstrate the functions of DHP in fish spermatogenesis. To further elucidate the roles of DHP in teleosts, we generated a pgr homozygous mutant line in XY Nile tilapia (Oreochromis niloticus). Pgr gene mutation resulted in the development of a smaller, thinner testis and a lower GSI compared with normal testis. Pgr gene knockout led to irregular arrangement of spermatogenic cysts, decline of sperm count and sperm motility. Significant decrease of spermatocytes and spermatozoa was observed, which was further proved by the PCNA and Ph3 staining. Real-time PCR analysis demonstrated that mutation of pgr gene resulted in a significant up-regulation of steroidogenesis-related genes of cyp17 alpha, cyp11b2, StAR, scc, 20 beta-HSD, and sf1, and down-regulation of fshb, fshr, oct4, sycp3, cdk1, prm, cyclinB1, cyclinB2 and cdc25 genes Furthermore, both Immunohistochemistry and Western blotting experiments revealed a remarkable increase of Cyp17a1, Cyp17a2 and Cyp11b2 expressions in the pgr -/- testis. EIA measurement showed that an evident increase of 11-KT level was found in the pgr(-/-) XY fish. There was a significant increase in the mortality of offspring when crossing pgr(-/-) XY fish with wild type XX fish. Increased TUNEL staining and enhanced apoptosis maker gene (bax) expressions were also observed. Taken together, our data suggested that DHP-activated physiology via pgr is crucial for the fertility in the XY tilapia.