The dev Operon Regulates the Timing of Sporulation during Myxococcus xanthus Development

Myxococcus xanthus undergoes multicellular development when starved. Thousands of rod-shaped cells coordinate their movements and aggregate into mounds in which cells differentiate into spores. Mutations in thedev operon impair development. Thedev operon encompasses a clustered regularly interspaced short palindromic repeat-associated (CRISPR-Cas) system. Null mutations indevI , a small gene at the beginning of thedev operon, suppress the developmental defects caused by null mutations in the downstreamdevR anddevS genes but failed to suppress defects caused by a small in-frame deletion indevT . We provide evidence that the original mutant has a second-site mutation. We show thatdevT null mutants exhibit developmental defects indistinguishable fromdevR anddevS null mutants, and a null mutation indevI suppresses the defects of adevT null mutation. The similarity of DevTRS proteins to components of the CRISPR-associated complex for antiviral defense (Cascade), together with our molecular characterization ofdev mutants, support a model in which DevTRS form a Cascade-like subcomplex that negatively autoregulatesdev transcript accumulation and prevents DevI overproduction that would strongly inhibit sporulation. Our results also suggest that DevI transiently inhibits sporulation when regulated normally. The mechanism of transient inhibition may involve MrpC, a key transcription factor, whose translation appears to be weakly inhibited by DevI. Finally, our characterization of adevI devS mutant indicates that very littleexo transcript is required for sporulation, which is surprising since Exo proteins help form the polysaccharide spore coat.IMPORTANCE CRISPR-Cas systems typically function as adaptive immune systems in bacteria. Thedev CRISPR-Cas system ofM. xanthus has been proposed to prevent bacteriophage infection during development, but howdev controls sporulation has been elusive. Recent evidence supported a model in which DevR and DevS prevent overproduction of DevI, a predicted 40-residue inhibitor of sporulation. We provide genetic evidence that DevT functions together with DevR and DevS to prevent DevI overproduction. We also show that spores form about 6 h earlier in mutants lackingdevI than in the wild type. Only a minority of natural isolates appear to have a functionaldev promoter anddevI , suggesting that a functionaldev CRISPR-Cas system evolved recently in niches where delayed sporulation and/or protection from bacteriophage infection proved advantageous.