MEIG1/PACRG associated and non-associated functions of axonemal dynein light intermediate polypeptide 1 (DNALI1) in mammalian spermatogenesis

Abstract

Axonemal dynein light intermediate polypeptide 1 (DNALI1) was originally cloned from Chlamydomonas reinhardtii in an effort to find motor proteins essential for flagellar motility. Here we report that DNALI1 is a binding partner of parkin co-regulated gene 1 (PACRG), which forms a complex with meiosis expressed gene 1 (MEIG1) in the manchette, a transient and unique structure only present in the elongating spermatids and required for normal spermiogenesis of the male germ cell differentiation process. DNALI1 recruits the PACRG protein in transfected CHO cells, and also stabilizes PACRG in bacteria and transfected mammalian cells. The untagged DNALI1 could also be co-purified with His-tagged PACRG in the gel filtration assay. Immunofluorescence staining on isolated male germ cells revealed that DNALI1 was present in the manchette of elongating spermatids, and colocalized with PACRG in this structure. In Pacrg mutant mice, localization of DNALI1 in the manchette was not changed, suggesting that DNALI1 and PACRG form a complex in the manchette, with DNALI1 being an upstream molecule. Mice deficiency in DNALI1 specifically in male germ cells showed dramatically reduced sperm numbers and were infertile. In addition, majority of the sperm exhibited abnormal morphology including misshapen heads, bent tails and enlarged midpiece, discontinuous accessory structure, and loss of sperm individualization, emphasizing the importance of DNALI1 in sperm development. Examination of testis histology revealed impaired spermiogenesis in the conditional Dnali1 knockout mice. Electron microscopy revealed disrupted ultrastructure in sperm of the Dnali1 mutant mice. Testicular levels of MEIG1, PACRG and SPAG16L proteins were not changed in the Dnali1 mutant mice. However, MEIG1 and SPAG16L were no longer present in the manchette in the absence of DNALI1. These findings demonstrate that DNALI1 is involved in the connection of the MEIG1/PACRG complex to carry cargo proteins along the manchette microtubules for sperm flagella formation. Given that Dnali1 mutant mice showed impaired sperm individualization that was not observed in the MEIG1 nor PACRG-deficient mice, DNALI1 might fulfill other functions beyond its role associated with the MEIG1/PACRG complex. Thus, DNALI1 plays multiple roles in sperm cell differentiation and function. Summary statementAxonemal dynein light intermediate polypeptide 1 (DNALI1) is required for sperm formation and male fertility. It associates with the MEIG1/PACRG complex in the manchette and is involved in a cargo transport system. In addition, it might be related to IFT and sperm individualization.