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While such divergence has long been suspected to contribute to hybrid male sterility, leading to reproductive isolation and thus speciation, the underlying mechanisms remain elusive. Here, we identify a molecular basis linking Y chromosome divergence to reproductive isolation in Drosophila. We show that male hybrids between D. simulans and D. mauritiana fail to properly express key Y-linked fertility genes. These genes contain unusually large introns, exceeding megabases and show substantial sequence divergence between species. In the hybrids, these gigantic introns are misprocessed, resulting in widespread splicing defects, including aberrant \u201cback-splicing\u201d events that join later exons to earlier ones. Our findings suggest that sequence divergence within introns can disrupt essential gene expression through defective splicing, providing a mechanistic link between rapid Y chromosome evolution and hybrid sterility. This work highlights the underappreciated role of intronic divergence in speciation.<\/jats:p>","DOI":"10.1093\/molbev\/msag045","type":"journal-article","created":{"date-parts":[[2026,2,13]],"date-time":"2026-02-13T12:46:07Z","timestamp":1770986767000},"source":"Crossref","is-referenced-by-count":1,"title":["Defective splicing of Y-chromosome-linked gigantic genes contributes to hybrid male sterility in Drosophila"],"prefix":"10.1093","volume":"43","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-1918-1393","authenticated-orcid":false,"given":"Adrienne","family":"Fontan","sequence":"first","affiliation":[{"name":"Whitehead Institute for Biomedical Research, Cambridge ,","place":["USA"]},{"name":"Department of Biology, Massachusetts Institute of Technology, Cambridge 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