A deficiency screen identifies genomic regions critical for sperm head-tail connection

Abstract

A stable connection between the sperm head and tail is critical for fertility in species with flagellated sperm. The head-tail coupling apparatus (HTCA) serves as the critical link between the nucleus (head) and the axoneme (tail) via the centriole. To identify regions of the Drosophila melanogaster genome that contain genetic elements that influence HTCA formation, we undertook a two part screen using the Drosophila deficiency (Df) kit. For this screen, we utilized a sensitized genetic background that overexpresses the pericentriolar material regulatory protein Pericentrin-Like Protein (PLP). We had previously shown that PLP overexpression (PLPOE) disrupts the head-tail connection in some spermatids, but not to a degree sufficient to reduce fertility. In the first step of the screen we tested for Dfs that in combination with PLPOE cause a reduction in fertility. We ultimately identified 11 regions of the genome that showed an enhanced fertility defect when combined with PLP overexpression. In the second step of the screen we tested these Dfs for their ability to enhance the HTCA defect caused by PLPOE, finding six. We then tested smaller Dfs to narrow the region of the genome that contained these enhancers. To further analyze the regions of the genome removed by these Dfs, we examined the expression patterns of the genes within these Dfs in publicly available datasets of RNAseq of Drosophila tissues and snRNAseq of Drosophila testes. In total, our analysis suggests that some of these Dfs may contain a single gene that might influence HTCA formation and / or fertility, while others appear to be regions of the genome especially rich in testis-expressed genes that might affect the HTCA because of complex, multi-gene interactions. Article SummaryWe perform a genetic enhancer deficiency screen to uncover genomic regions required for proper sperm head-tail connection. We identified 6 regions and provide insight into these regions using publicly available RNA sequence data. Our data reveal that these regions are exceptionally rich in testes specific genes. Further analysis using small deficiencies resulted in two classes of enhancers: one class likely enhances head-tail connection by disrupting multiple genes, while the second class might house a single gene responsible for the reduction in fertility.