An epididymis-specific secretory protein Clpsl2 critically regulates sperm motility, acrosomal integrity, and male fertility.

The epididymis performs an important role in the maturation of spermatozoa including their acquisition of progressive motility and fertilizing ability. However, the molecular mechanisms that govern these maturational events are still poorly defined. Here we report that Clpsl2, a novel colipase homology, is exclusively expressed in the caupt epididymis and conserved in mammalian. Clpsl2 was secreted into the lumen and covered the acrosome region and principal piece of spermatozoa tail. And during epididymal transit, the binding rate between Clpsl2 protein and the spermatozoa gradually decreased. Though Clpsl2 had the highest identifies with pancreatic colipase (Clps), Clpsl2 lacked those conserved amino acids in pancreatic Clps that interacting with lipase, correspondingly, the recombinant Clpsl2 protein did not possess the Clps function such as promoting the hydrolysis of lipase to its substrate glycerine trioleate. However, sequence analysis showed that Clpsl2 has the potency to bind with lipid. Knockdown expression of Clpsl2 by lentivirus-mediated RNAi in vivo caused an attenuation of spermatozoa motility, a suppressed acrosomal reaction, a decrease of cauda spermatozoa number, and subfertility. This study identified a novel and conserved molecule, Clpsl2, was specifically expressed in epididymis and involved in the regulation of spermatozoa motility, acrosomal integrity, and male fertility.

[Lentiviral vector-mediated RNA interference of mouse epididymis-specific meClps gene lowers mouse sperm mobility].

OBJECTIVE: To analyze the effect of small interfering RNA (siRNA) targeting mouse epididymis-specific colipase-like (meClps) gene on mouse sperm mobility. METHODS: The eukaryotic expression vector pDsRed2.0-C1-meClps was constructed and transfected into NIH-3T3 cells, and the protein expression was detected with anti-meClps serum. Three interfering sequences targeting meClps (RNAi-251, 224 and 286) were inserted into lentiviral vectors pRNAT-U6.2/lenti, which were co-transfected with pDsRed2.0-C1-meClps into NIH-3T3 cells. The RNA interfering efficiency was confirmed by semi-quantitative PCR and Western blotting. The lentivirus, packed with the lentiviral vector with the highest interfering efficiency, was injected into the caput tissues of mouse epididymis, and its effect on sperm mobility of the cauda epididymis was evaluated. RESULTS: All the 3 lentiviral RNAi vectors targeting meClps could inhibit the mRNA and protein expressions of meClps, among which pRNAT-U6.2/lenti-RNAi-251 had the highest interfering efficiency. The lentivirus packed with pRNAT-U6.2/lenti-RNAi-251 significantly reduced the path velocity of cauda sperm after injection into the caput epididymis of the mice (P<0.05). CONCLUSION: Knock-down meClps expression by lentiviral-mediated RNA interference can lower sperm mobility of mice.