Uncovering roles for Caspar/dFaf1 in Drosophila development

Abstract

Fas associating Factors (Faf1/Casp and Faf2) are members of the UBX domain containing proteasomal adaptors family of proteins. The primary sequence analysis suggests conservation of structure and function between flies and humans. Casp has been previously reported to be involved in Drosophila innate immunity and also has been characterized as a maternal effect gene in development. The physiological function of Faf2 is not well characterized in Drosophila. In this study, we attempt to characterize the role of the different functional domains of Casp by generating Casp transgenic animals with targeted domain deletions. The deletions of interest were the ΔUBX, and a combined deletion ΔUASΔUBX. Our first attempt used CRISPR/Cas9 based genome editing, which did not succeed. As an alternative, we were successful in generating a number of overexpression transgenic animals, pUASP-ΔDomain flies, which we plan to express in a Caspnull animal using the UAS Gal4 system. We are also redoing the CRISPR screen using a modified design. Additionally, a loss of function analysis using reverse genetics on the Casp homolog Faf2 suggest that Faf2 is a paternal effect gene. Zygotic copy of Faf2 is not sufficient to rescue the paternal phenotype. Faf2 knockdown does not cause sterility of the male germline as the embryos fathered by Faf2 knockdown males proceed through nuclear divisions. Paternal germline specific Faf2 knockdown results in embryonic developmental arrest at early stages of gastrulation. Given that Faf2 is not present in the Drosophila sperm and that Faf2 influences protein dynamics in the cell, we hypothesize that Faf2 functions in the observed paternal effect phenotype by altering the chromatin architecture of paternal chromosomes.