Synergistic enhancement of the mouse Pramex1 and Pramel1 in repressing retinoic acid (RA) signaling during gametogenesis

Abstract

Background

PRAME constitutes one of the largest multi-copy gene families in Eutherians, encoding cancer-testis antigens (CTAs) with leucine-rich repeats (LRR) domains, highly expressed in cancer cells and gametogenic germ cells. This study aims to elucidate genetic interactions between two members, Pramex1 and Pramel1, in the mouse Prame family during gametogenesis using a gene knockout approach.

Result

Single-gene knockout (sKO) of either Pramex1 or Pramel1 resulted in approximately 7% of abnormal seminiferous tubules, characterized by a Sertoli-cell only (SCO) phenotype, impacting sperm count and fecundity significantly. Remarkably, sKO female mice displayed normal reproductive functions. In contrast, Pramex1/Pramel1 double knockout (dKO) mice exhibited reduced fecundity in both sexes. In dKO females, ovarian primary follicle count decreased by 50% compared to sKO and WT mice, correlating with a 50% fecundity decrease. This suggested compensatory roles during oogenesis in Pramex1 or Pramel1 sKO females. Conversely, dKO males showed an 18% frequency of SCO tubules, increased apoptotic germ cells, and decreased undifferentiated spermatogonia compared to sKO and WT testes. Western blot analysis with PRAMEX1- or PRAMEL1-specific antibodies on sKO testes revealed compensatory upregulation of each protein (30–50%) in response to the other gene’s deletion. Double KO males exhibited more severe defects in sperm count and litter size, surpassing Pramex1 and Pramel1 sKO accumulative effects, indicating a synergistic enhancement interaction during spermatogenesis. Additional experiments administering trans-retinoic acid (RA) and its inhibitor (WIN18,446) in sKO, dKO, and WT mice suggested that PRAMEX1 and PRAMEL1 synergistically repress the RA signaling pathway during spermatogenesis.

Conclusion

Data from sKO and dKO mice unveil a synergistic interaction via the RA signaling pathway between Pramex1 and Pramel1 genes during gametogenesis. This discovery sets the stage for investigating interactions among other members within the Prame family, advancing our understanding of multi-copy gene families involved in germ cell formation and function.