Chromatin state contributions to cytoplasmic incompatibility

About This Grant

Project Summary Among all endosymbionts, maternally transmitted Wolbachia bacteria are the most common, due in large part to their ability to influence host reproduction to spread. Many Wolbachia cause cytoplasmic incompatibility (CI), a sperm modification that kills embryos without Wolbachia. Two prophage-associated genes (cifA and cifB) cause and one gene (cifA) rescues CI. Cytologically, CI produces cycle-1 defects in embryogenesis when paternal chromosomes fail to properly condense. However, CI-derived embryos regularly escape cycle-1 defects and display chromosome segregation defects later in embryogenesis during the cortical divisions (10- 14). Further, some individuals fully escape CI, proceeding to larval and even adult life stages. This has resulted in a large gap in knowledge about whether a common acute CI mechanism acts at different time points, or if independent and temporally distinct mechanisms underlie these patterns. The proposed research will fill this gap in knowledge by integrating genetic, cytogenetic, and genomic analyses to determine the primary cellular defect underlying embryonic lethality in the context of two mechanistic models. The host-modification (HM) model predicts that Cifs modify male gametes prior to fertilization, while the toxin-antidote (TA) model predicts that Wolbachia produce a Cif toxin that is transferred directly to the embryo. The presence of independent defects beyond cycle-1 lends support to the HM model since a putative toxin is unlikely to persist throughout embryogenesis. Elegant experiments have tracked cycle-1 mitotic defects to errors at the histone-protamine transition in spermatogenesis that cause abnormal levels of paternal histones in sperm nuclei. Very recent data also indicate a reduction of H3K9me2,3—a marker of inactive heterochromatin—in CI crosses. These data support the hypothesis that CI-causing Wolbachia modify host chromatin and interfere with the histone post- translational-modifications (PTM) required for normal histone removal. The proposed work will determine if histone PTMs are maintained through development in embryos and larvae which ‘escape’ embryonic death. Chromosome structure will then be evaluated in larvae to determine the relationship between histone PTMs and decondensed chromosomes. To determine the genome-wide effects of CI-induced chromatin changes, chromatin accessibility in embryos will be mapped using single-cell ATAC-seq. This technique was chosen to specifically identify differentially condensed genomic regions in the presence of Wolbachia. Finally, a Position Effect Variegation screen will be used to assay effects on gene expression as a functional output of the chromatin states. In addition to preparing the applicant for a long-term career as a principal investigator, this work will fill critical gaps in knowledge by tracking the chromatin state changes that occur with CI and determining if a single mechanism underlies CI death. This work will contribute to our broader knowledge of CI, which is critical for explaining global Wolbachia prevalence in natural populations and to improving the efficacy of Wolbachia biocontrol of human disease transmission in transinfected vector populations.