DNAI4 is required for flagellum assembly and male fertility in mice

Multiple morphological abnormalities of the sperm flagella (MMAF) is a major cause of male infertility, but identified gene variants can only explain about 60% of clinical cases. Here, dynein axonemal intermediate chain 4 (DNAI4) is identified as an essential regulator of sperm flagellum morphogenesis. RT-PCR and western blot analyses indicate that expression of DNAI4 is enriched in murine testes. Dnai4 deletion in mice causes male-specific infertility due to oligoasthenoteratospermia with MMAF. Electron microscopy analyses revealed that DNAI4 deficiency resulted in an abnormal ultrastructure of sperm flagella, including disorganized mitochondrial sheaths, outer dense fibers and '9+2' axonemes, and missing inner dynein arms (IDA) and outer dynein arms. The IDA component DNAH10 was remarkably reduced in testes and sperm tails of Dnai4 knockout mice. Immunoprecipitation demonstrated an interaction between DNAI4 and the intraflagellar transport protein IFT144 within the testes. Other IFT-A members, including IFT140, IFT122 and IFT121, were downregulated in sperm tails following Dnai4 deletion. Taken together, these findings establish DNAI4 as an essential regulator of sperm flagellum assembly and mammalian spermiogenesis, operating through the regulation of IDA assembly and retrograde intraflagellar transport.