The heterodimeric hypoxia-inducible factor (HIF)-1 is a transcriptional master regulator of several genes involved in mammalian oxygen homeostasis. We described previously testis-specific mouse (mHIF-1aI.1) and human isoforms of HIF-1a(hHIF-1aTe), which are the result of two distinct promoter/first exon combinations. Performing a yeast two-hybrid screen using mHIF-1aI.1 as bait, we identified a protein-protein interaction between HIF-1a and the fibrous sheath protein TSGA10. The interaction was verified by pull-down assays. TSGA10 is a structural protein processed during spermiogenesis. Immunofluorescence analysis indicated that the C-terminal part of processed TSGA10 accumulates in the midpiece of the spermatozoa tail, where it co-localizes with HIF-1a. HIF-1a was not affecting TSGA10-mediated fibre formation after overexpression in vitro. However, HIF-1a nuclear accumulation and HIF-1 transcriptional activity was significantly affected by overexpressed TSGA10. These data indicate that TSGA10 might localize the usually nuclear HIF-1a protein to this unusual localization in the sperm tail.