Abstract
BACKGROUND: Hybridization can trigger phenotypic and transcriptomic changes, leading to reproductive isolation and genome evolution. Drosophila is a key model for understanding the genetic basis of speciation, yet the complete process leading to hybrid sterility remains unknown. We investigated the regulation of genes and transposable elements (TEs) from the onset of sterility to fertility recovery through backcrossing. RESULTS: We studied the testicular transcriptomes of Drosophila buzzatii, D. koepferae, sterile F1 hybrids, from several unidirectional crosses, and subsequent backcross generations. F1 hybrids presented testicular atrophy, absence of sperm, increased body size, and gene misregulation. Successive backcrosses with D. buzzatii restored gene expression and fertility in the third generation, but a set of 13 genes remained deregulated, potentially contributing to persistent gene flow barriers. TE expression exhibited a tendency towards underexpression, particularly for D. buzzatii-derived elements, with partial derepression in subsequent backcrosses. Importantly, 32% of differentially expressed genes (DEG) between parental species harbored lineage-specific TE insertions in flanking or intragenic regions, suggesting a putative role of these elements in shaping regulatory differences between these species. Regulatory divergence analyses identified DEG with cis and trans regulatory changes as prevalent in parental species, whereas compensatory regulation was enriched in misexpressed genes in hybrids. CONCLUSIONS: Hybrid incompatibilities may result from extensive deregulation of genes related to development, metabolism, and reproduction, possibly combined with the activity of specific TE copies. These findings demonstrate the complexity of regulatory incompatibilities in hybrids and the contribution of persistent deregulated genes and TE dynamics to postzygotic isolation.