69 Oral - Immunity and Microbiome
Friday April 08, 9:15 AM - 9:30 AM

The cytoplasmic incompatibility factor proteins CifA and CifB are both nucleases in Drosophila melanogaster


Authors:
Rupinder Kaur 1,2; J. Dylan Shropshire 3; Brittany Leigh 1,2; Seth Bordenstein 1,2,4,5

Affiliations:
1) Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA; 2) Vanderbilt Microbiome Initiative, Vanderbilt University, Nashville, TN, USA; 3) Division of Biological Sciences, University of Montana, Missoula, MT, USA ; 4) Department of Pathology, Microbiology & Immunology, Vanderbilt University Medical Center, Nashville, TN, USA; 5) Vanderbilt Institute for Infection, Immunology and Inflammation, Vanderbilt University Medical Center, Nashville, TN, USA

Keywords:
l. Wolbachia; a. spermatogenesis

The widespread symbiosis between invertebrates and Wolbachia bacteria is due in part to cytoplasmic incompatibility (CI) that is significant to arthropod vector control and evolution. While CI selectively favors the fitness of Wolbachia-transmitting females and thus the bacterial spread, the CI mechanism is not broadly understood. In Drosophila melanogaster, dual expression of the genes cifA and cifB from wMel causes CI, resulting in embryonic lethality between symbiotic male and aposymbiotic female cross. These two co-diverging genes span five phylogenetic Types, and previous work suggested different CifB Types are distinguishable by the presence/absence of nuclease activity. Here, we re-evaluate this claim using transgenic, mutant, enzymatic, and cytochemical assays in vitro and in situ. We demonstrate that contrary to prior conclusions, T1 CifB is an in vitro DNase that cleaves single- and double-stranded DNA. Moreover, we provide the first characterization of T1 CifA as a nuclease that cleaves both DNA and RNA. We further show that in vitro DNase activity translates to in situ spermatid DNA fragmentation in Cif-expressing and Wolbachia-bearing testes, and CifB-mediated spermatid DNA damage alone is insufficient to cause CI. Cif enzymatic activity is ablated by deletions and amino acid substitutions. In sum, these molecular insights (i) highlight previously unrecognized nuclease functions of T1 CifA and CifB proteins (ii) reinforce a conservative gene nomenclature and mechanistic model that is agnostic to presumptions about distinct Cif enzymatic activities and (iii) support a postulate of the Host Modification model of CI that spermatid DNA is altered by the Cif proteins.