Loss of RPL27a expression promotes p53 activation by modulating the RPL5-MDM2 interaction in mouse spermatogonia

Ribosomal proteins (RPs) regulate p53 activation, which is mediated by mouse double minute 2 homolog (MDM2), ribosomal protein L5 (RPL5) and/or ribosomal protein L11. Thus, this study aimed to explore whether ribosomal protein L27a (RPL27a) interacts with RPL5 and regulates p53 activation in mouse spermatogonia. Immunohistochemistry and immunofluorescence were used to analyze protein expression in vivo . Protein and mRNA expression was analyzed by immunoblotting and real-time polymerase chain reaction, respectively. The bindings of RPL27a with RPL5, and MDM2 were verified by molecular docking and glutathione S-transferase (GST)-fusion protein assays. Co-immunoprecipitation and in vitro ubiquitination assays were used to confirm protein interactions and p53 accumulation. An adeno-associated virus was used to knock down RPL27a in vivo . The results showed that knockdown of RPL27a weakened the binding of RPL27a to RPL5 and MDM2, but promoted the interaction between RPL5 and MDM2. Meanwhile, knockdown of RPL27a induced p53-dependent cell cycle arrest and RPL5-dependent p53 activation in mouse spermatogonial GC-1 spg cells. Moreover, RPL27a knockdown induced apoptosis of spermatogonia via the accumulation of p53, as verified in vivo . This study indicates that RPL27a negatively regulates p53 activation via enhancing the RPL5-MDM2 interaction in spermatogonia, and provides a better understanding of male infertility induced apoptosis of spermatogonia.