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The intrinsically unstructured C9ORF78 protein was detected in spliceosomes but its role in splicing is presently unclear. We find that C9ORF78 tightly interacts with the spliceosome remodeling factor, BRR2, in vitro. Affinity purification/mass spectrometry and RNA UV-crosslinking analyses identify additional C9ORF78 interactors in spliceosomes. Cryogenic electron microscopy structures reveal how C9ORF78 and the spliceosomal B complex protein, FBP21, wrap around the C-terminal helicase cassette of BRR2 in a mutually exclusive manner. Knock-down of C9ORF78 leads to alternative NAGNAG 3′-splice site usage and exon skipping, the latter dependent on BRR2. Inspection of spliceosome structures shows that C9ORF78 could contact several detected spliceosome interactors when bound to BRR2, including the suggested 3′-splice site regulating helicase, PRPF22. Together, our data establish C9ORF78 as a late-stage splicing regulatory protein that takes advantage of a multi-factor trafficking site on BRR2, providing one explanation for suggested roles of BRR2 during splicing catalysis and alternative splicing.

Cationic antimicrobial peptides (AMPs) are active immune effectors of multicellular organisms and also considered as new antimicrobial drug candidates. One of the problems encountered when developing AMPs as drugs is the difficulty to reach sufficient killing concentrations under physiological conditions. Here, using pexiganan, a cationic peptide derived from a host defence peptide of the African clawed frog and the first AMP developed into an antibacterial drug, we studied if sub-lethal effects of AMPs can be harnessed to devise treatment combinations. We studied the pexiganan stress response ofStaphylococcus aureusat sub-lethal concentrations using quantitative proteomics. Several proteins involved in nucleotide metabolism were elevated, suggesting a metabolic demand. We then show thatS. aureusis highly susceptible to antimetabolite nucleoside analogues when exposed to pexiganan, even at sub-inhibitory concentrations. These findings could be used to enhance pexiganan potency while decreasing the risk of resistance emergence, and our findings can likely be extended to other antimicrobial peptides.