Male Sterility in Drosophila hybrids: A Multi-Generational Transcriptomic Analysis of Genes and Transposable Elements in Testes

Hybridization can trigger profound phenotypic and transcriptomic changes, revealing mechanisms of reproductive isolation and genome evolution. We studied the testicular transcriptomes of Drosophila buzzatii, D. koepferae, their sterile F1 hybrid males, and subsequent backcross generations to assess the impact of hybridization on the regulation of genes and transposable elements (TEs). F1 hybrids exhibited testicular atrophy, absence of sperm, increased body size, and widespread gene misregulation (~41% of genes), with enriched metabolic and reproductive pathway targets. Successive backcrossing with D. buzzatii restored gene expression and fertility by the third generation, but a core of 13 genes remained consistently deregulated. TE expression exhibited a predominant trend toward underexpression, especially of D. buzzatii-derived elements, with some derepression in later backcrosses. Regulatory divergence analyses indicated that cis-regulatory changes predominated in parental species, and compensatory regulation was enriched in misexpressed genes in hybrids. These findings demonstrate the complexity of regulatory incompatibilities in hybrids and underscore the contribution of persistent deregulated genes and TE dynamics to postzygotic isolation.