DNAI4 is Required for Flagellum Assembly and Male Fertility in Mice

Knockout mice lacking DNAI4 are completely infertile with shattered sperm flagella, pointing to a candidate gene for the ~40% of human MMAF cases still unexplained.

Journal: Development (Cambridge, England) | Published: 2026-05-19 | Type: Journal Article | PMID: 42083518 Authors: Yang Fan et al., Yangzhou University; one co-author from University of Pennsylvania Funding/COI: National Natural Science Foundation of China, Jiangsu Province grants, China Postdoctoral Science Foundation. No competing interests declared.

Summary

Multiple morphological abnormalities of the sperm flagella (MMAF) is a recognized cause of male infertility, but known gene variants account for only ~60% of clinical cases. This study from Yangzhou University used a knockout mouse model to show that deleting Dnai4 causes complete male infertility, with sperm exhibiting structural collapse across every major flagellar compartment. The proposed mechanism runs through retrograde intraflagellar transport (IFT-A): DNAI4 interacts with IFT144, and without it, downstream IFT-A members and inner dynein arms fail to assemble properly.

Claims

• Dnai4 knockout male mice are entirely infertile; female knockouts are fertile, confirming the phenotype is male-specific
• Sperm from knockout mice display oligoasthenoteratospermia with MMAF: shattered mitochondrial sheaths, disorganized outer dense fibers, disrupted '9+2' axonemes, and missing both inner and outer dynein arms
• DNAH10 (an inner dynein arm component) is markedly reduced in testes and sperm tails of knockout mice
• Co-immunoprecipitation confirmed a physical interaction between DNAI4 and the IFT-A protein IFT144 in testicular tissue
• IFT-A subunits IFT140, IFT122, and IFT121 are all downregulated in sperm tails following Dnai4 deletion
• Dnai4 expression is testis-enriched in mice (RT-PCR and western blot)

Study Quality

This is a well-executed mouse knockout study. The authors used a standard panel of techniques — RT-PCR, western blot, co-immunoprecipitation, and transmission electron microscopy — and the EM data in particular is the backbone of the structural claims. The knockout model is clean: male infertility is fully penetrant, female fertility is unaffected, which rules out a general developmental catastrophe and points to a flagellum-specific defect.

The mechanistic story is coherent. IFT-A retrograde transport is a plausible node for MMAF pathology, and the finding that multiple IFT-A subunits are destabilized downstream of DNAI4 loss gives the model some depth. That said, the paper doesn't establish causality between IFT-A disruption and dynein arm loss — the connection is correlative within the mouse.

Red Flags

• Mice, not men. No human genetic data is presented. The case that DNAI4 variants cause human infertility is entirely inferential.
• No patient cohort. The obvious next step — screening infertile men with unexplained MMAF for DNAI4 variants — is not done here.
• Mechanism is correlative. The paper shows that IFT-A members decrease when DNAI4 is absent, but doesn't rescue or confirm the causal chain with targeted experiments.
• No quantitative sperm parameters reported in a structured way (sperm count, motility percentage, morphology rates) — the abstract uses qualitative descriptors rather than numbers.
• All authors from a single institution (with one Penn collaborator), limiting independent validation.

Strengths

• Robust multi-technique approach: EM, immunoprecipitation, western blot, and gene expression converge on the same conclusion
• Clean mouse phenotype with male-specific infertility is compelling
• No competing interests declared; funding is government academic, not industry
• Adds a candidate gene to an underdetermined disease (40% of MMAF cases still lack a genetic explanation)
• Published in Development, a peer-reviewed journal with appropriate scope for this work

Verdict

This is solid basic science that earns its place in the MMAF gene discovery literature. The mouse knockout data is convincing, the structural phenotype is thorough, and the IFT-A mechanistic link is a legitimate contribution. What it is not is clinical evidence: until someone screens infertile men for DNAI4 variants and finds them, this remains a well-characterized mouse gene with human implications yet to be demonstrated. File it as a strong candidate for the unsolved 40% — not a solved case.