Two outer membrane proteins contribute to Caulobacter crescentus cellular fitness by preventing intracellular S-layer protein accumulation

Caulobacter crescentus is unique in that it has two homologous, seemingly redundant outer membrane proteins, RsaFa and RsaFb, that, together with other components, form a type I protein translocation pathway for S-layer export. RsaFa and RsaFb are required to prevent intracellular accumulation and aggregation of the S-layer protein RsaA; deletion of RsaFa and RsaFb led to a general growth defect and lowered cellular fitness. We show that loss of both RsaFa and RsaFb led to accumulation of insoluble RsaA in the cytoplasm, which in turn caused upregulation of a number of genes involved in protein mis-folding and degradation pathways. These findings provide new insight into the requirement for RsaFa and RsaFb in cellular fitness and tolerance to antimicrobial agents and further our understanding of the S-layer export mechanism on both the transcriptional and translational levels in C. crescentus. RNA-seq of biological triplicate samples of Caulobacter crescentus NA1000 wild type, single RsaFa mutant, single RsaFb mutant, double RsaFa/RsaFb mutant, and triple RsaFa/RsaFb/RsaA mutant