DNAH3

DNAH3 is an axonemal dynein heavy chain essential for inner dynein arm (IDA) assembly and motility of sperm flagella. It localizes to the sperm flagellum, physically interacts with DNALI1, and is required for stable expression of other IDA components including DNAH1, DNAH6, and DNALI1; loss of DNAH3 causes IDA deficiency, central pair microtubule loss, mitochondrial and fibrous sheath dislocation, and severely impaired sperm motility (PMID:38312775, PMID:39503742, PMID:41731497). Biallelic loss-of-function variants in DNAH3 cause asthenoteratozoospermia and male infertility in humans, a phenotype faithfully recapitulated by Dnah3-knockout mice (PMID:38312775, PMID:39503742, PMID:39588341). DNAH3 deficiency also disturbs sperm energy metabolism, reducing levels of L-palmitoylcarnitine and glycocholic acid, suggesting an additional role in metabolic support of flagellar function (PMID:39774634).

1. 2008 Medium

   Before any connection to flagellar biology, retroviral insertional mutagenesis identified Dnah3 as a gene whose overexpression or truncation impairs microtubule function in colon epithelial cells, providing the first hint that DNAH3 influences cytoskeletal dynamics.

   Evidence Retroviral integration mapping and Dnah3 overexpression in mouse IMCE colon epithelial cells with microtubule function assays

   PMID:18294281

   Open questions at the time

   • Overexpression artefact in a non-ciliated cell type; relevance to endogenous DNAH3 function unclear
   • No loss-of-function experiment performed
   • Connection to dynein arm biology not established
2. 2016 Medium

   Detection of DNAH3 protein in human spermatozoa by targeted proteomics confirmed that this dynein heavy chain is expressed in the relevant cell type for motility function.

   Evidence Mining of PRIDE mass spectrometry data with SRM/MRM validation in human sperm

   PMID:27581094

   Open questions at the time

   • Sub-flagellar localization not determined
   • No functional data linking DNAH3 protein to sperm motility
3. 2024 High

   Genetic and ultrastructural studies in patients and knockout mice established that biallelic DNAH3 loss causes asthenoteratozoospermia by disrupting IDA assembly, central pair microtubules, and flagellar accessory structures, resolving the question of whether DNAH3 is required for sperm motility.

   Evidence WES in infertile men, two independent Dnah3-KO mouse lines, TEM, immunostaining, CASA, and Western blot across multiple studies

   PMID:38312775 PMID:39503742 PMID:39588341

   Open questions at the time

   • Whether DNAH3 is a structural subunit versus an assembly factor for IDA was not resolved
   • Mechanism by which DNAH3 loss disrupts the central pair of microtubules remains unknown
   • No structural or biochemical reconstitution of the DNAH3-containing IDA complex
4. 2025 Medium

   Metabolomic profiling of Dnah3-KO mouse sperm revealed that DNAH3 deficiency alters energy metabolism — reducing L-palmitoylcarnitine and glycocholic acid — suggesting that impaired motility is not solely due to structural defects but also involves metabolic dysregulation.

   Evidence Sperm metabolomics in Dnah3-KO mice with CASA and histology

   PMID:39774634

   Open questions at the time

   • Causal relationship between metabolite changes and motility loss not established
   • Whether metabolic effects are direct or secondary to structural disorganization is unknown
   • Single-lab finding not independently replicated
5. 2026 High

   Co-immunoprecipitation and mass spectrometry identified DNALI1 as a direct physical interactor of DNAH3, establishing that DNAH3 forms a complex with a known IDA light intermediate chain and that this interaction is required for IDA integrity and sperm motility gene expression programs.

   Evidence Co-IP/MS in testis, comparative proteomics, scRNA-seq of elongating spermatids, and Dnah3-KO mice with EM and immunostaining

   PMID:41731497

   Open questions at the time

   • Full stoichiometry and additional subunits of the DNAH3-containing IDA complex not determined
   • Whether DNAH3 interacts directly with DNAH1 or DNAH6 was not tested by reciprocal co-IP
   • scRNA-seq changes in motility gene expression lack causal mechanistic follow-up

• Key unresolved questions include the precise molecular architecture of the DNAH3-containing IDA complex, whether DNAH3 has ATPase/motor activity distinct from other dynein heavy chains, how its loss leads to central pair defects, and whether its metabolic role is direct or secondary.
• No in vitro reconstitution or cryo-EM structure of DNAH3-containing IDA
• ATPase activity of DNAH3 not directly measured
• Mechanism linking IDA deficiency to central pair and accessory structure loss is unknown