Small Proteins Can No Longer Be Ignored

Storz, Gisela; Wolf, Yuri I.; Ramamurthi, Kumaran S.

doi:10.1146/annurev-biochem-070611-102400

Cited by 342 publications

(412 citation statements)

References 107 publications

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“…Rather, DevI appears to be a small 40-residue protein that acts in a cell-autonomous manner. Small proteins have been shown to affect sporulation of Bacillus subtilis and various processes (signal transduction, enzyme activity, transport of ions and macromolecules across membranes, and cell division) in many bacteria (39). Recently, the 37-residue CmpA protein of B. subtilis has been shown to function as an adapter that targets SpoIVFA for proteolysis by ClpXP (40).…”

Section: Discussionmentioning

confidence: 99%

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

Rajagopalan

Kroos

2017

J Bacteriol

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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 the dev operon impair development. The dev operon encompasses a clustered regularly interspaced short palindromic repeat-associated (CRISPR-Cas) system. Null mutations in devI, a small gene at the beginning of the dev operon, suppress the developmental defects caused by null mutations in the downstream devR and devS genes but failed to suppress defects caused by a small in-frame deletion in devT. We provide evidence that the original mutant has a second-site mutation. We show that devT null mutants exhibit developmental defects indistinguishable from devR and devS null mutants, and a null mutation in devI suppresses the defects of a devT null mutation. The similarity of DevTRS proteins to components of the CRISPR-associated complex for antiviral defense (Cascade), together with our molecular characterization of dev mutants, support a model in which DevTRS form a Cascade-like subcomplex that negatively autoregulates dev 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 a devI devS mutant indicates that very little exo 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. The dev CRISPR-Cas system of M. xanthus has been proposed to prevent bacteriophage infection during development, but how dev 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 lacking devI than in the wild type. Only a minority of natural isolates appear to have a functional dev promoter and devI, suggesting that a functional dev CRISPR-Cas system evolved recently in niches where delayed sporulation and/or protection from bacteriophage infection proved advantageous.KEYWORDS CRISPR-Cas, Myxococcus xanthus, bacterial development, dev operon, gene regulation, signaling, sporulation M yxococcus xanthus is a Gram-negative soil bacterium that exhibits social behaviors (1). In nature, cells of M. xanthus move in groups, feeding on prey bacteria and organic detritus. When starved, thousands of cells aggregate to form mounds that mature into fruiting bodies as rod-shaped cells differentiate into dormant ovoid spores.

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Section: Discussionmentioning

confidence: 99%

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

Rajagopalan

Kroos

2017

J Bacteriol

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“…If the predicted 40-residue DevI protein acts cell autonomously, how might it inhibit sporulation? Small proteins have been shown to affect diverse processes, including spore formation, cell division, enzyme activity, transport of ions and macromolecules across membranes, and signal transduction (76). In Bacillus subtilis, two small proteins affect endospore formation.…”

Section: Figmentioning

confidence: 99%

devIIs an Evolutionarily Young Negative Regulator of Myxococcus xanthus Development

et al. 2015

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During starvation-induced development of Myxococcus xanthus, thousands of rod-shaped cells form mounds in which they differentiate into spores. The dev locus includes eight genes followed by clustered regularly interspaced short palindromic repeats (CRISPRs), comprising a CRISPR-Cas system (Cas stands for CRISPR associated) typically involved in RNA interference. Mutations in devS or devR of a lab reference strain permit mound formation but impair sporulation. We report that natural isolates of M. xanthus capable of normal development are highly polymorphic in the promoter region of the dev operon. We show that the dev promoter is predicted to be nonfunctional in most natural isolates and is dispensable for development of a laboratory reference strain. Moreover, deletion of the dev promoter or the small gene immediately downstream of it, here designated devI (development inhibitor), suppressed the sporulation defect of devS or devR mutants in the lab strain. Complementation experiments and the result of introducing a premature stop codon in devI support a model in which DevRS proteins negatively autoregulate expression of devI, whose 40-residue protein product DevI inhibits sporulation if overexpressed. DevI appears to act in a cellautonomous manner since experiments with conditioned medium and with cell mixtures gave no indication of extracellular effects. Strikingly, we report that devI is entirely absent from most M. xanthus natural isolates and was only recently integrated into the developmental programs of some lineages. These results provide important new insights into both the evolutionary history of the dev operon and its mechanistic role in M. xanthus sporulation. IMPORTANCECertain mutations in the dev CRISPR-Cas (clustered regularly interspaced short palindromic repeat-associated) system of Myxococcus xanthus impair sporulation. The link between development and a CRISPR-Cas system has been a mystery. Surprisingly, DNA sequencing of natural isolates revealed that many appear to lack a functional dev promoter, yet these strains sporulate normally. Deletion of the dev promoter or the small gene downstream of it suppressed the sporulation defect of a lab strain with mutations in dev genes encoding Cas proteins. The results support a model in which the Cas proteins DevRS prevent overexpression of the small gene devI, which codes for an inhibitor of sporulation. Phylogenetic analysis of natural isolates suggests that devI and the dev promoter were only recently acquired in some lineages. Myxococcus xanthus is a Gram-negative bacterium of the deltaproteobacteria group. Typically found in the soil, M. xanthus exhibits social behaviors (1). The rod-shaped cells move in groups on solid surfaces, feeding on prey bacteria and organic matter. When starved, cells modify their movements and build mounds containing thousands of individuals that differentiate into ovoid spores. The resulting mound of dormant and insultresistant spores is called a fruiting body. Spores within the fruiting body germinate when con...

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“…In this regulatory network, the role of small proteins has largely been ignored, possibly due to the assigned size limits of open reading frames (ORFs) during annotation and the lack of bioinformatics approaches to annotate these small proteins (5). However, in today's genomic age, several small proteins that were previously unknown have been identified and their functions have been determined (see reference 6 and references therein).…”

mentioning

confidence: 99%

“…In B. subtilis, the Fur-regulated FbpA, FbpB, and FbpC proteins, which are small and basic in nature, are also known to act as metal chaperones in the iron-sparing response of B. subtilis (16). Understandably, the diverse functions of small proteins have prompted refinement in the annotation of genomes, leading to the identification of more small proteins in other bacteria by exploration of their small proteomes (5). Our group previously explored the regulatory function of HLP, which is an immunogenic nucleoid-associated small protein that contributes to the virulence of S. mutans (17).…”

mentioning

confidence: 99%

Pleiotropic Regulation of Virulence Genes in Streptococcus mutans by the Conserved Small Protein SprV

2017

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Streptococcus mutans, an oral pathogen associated with dental caries, colonizes tooth surfaces as polymicrobial biofilms known as dental plaque. S. mutans expresses several virulence factors that allow the organism to tolerate environmental fluctuations and compete with other microorganisms. We recently identified a small hypothetical protein (90 amino acids) essential for the normal growth of the bacterium. Inactivation of the gene, SMU.2137, encoding this protein caused a significant growth defect and loss of various virulence-associated functions. An S. mutans strain lacking this gene was more sensitive to acid, temperature, osmotic, oxidative, and DNA damage-inducing stresses. In addition, we observed an altered protein profile and defects in biofilm formation, bacteriocin production, and natural competence development, possibly due to the fitness defect associated with SMU.2137 deletion. Transcriptome sequencing revealed that nearly 20% of the S. mutans genes were differentially expressed upon SMU.2137 deletion, thereby suggesting a pleiotropic effect. Therefore, we have renamed this hitherto uncharacterized gene as sprV (streptococcal pleiotropic regulator of virulence). The transcript levels of several relevant genes in the sprV mutant corroborated the phenotypes observed upon sprV deletion. Owing to its highly conserved nature, inactivation of the sprV ortholog in Streptococcus gordonii also resulted in poor growth and defective UV tolerance and competence development as in the case of S. mutans. Our experiments suggest that SprV is functionally distinct from its homologs identified by structure and sequence homology. Nonetheless, our current work is aimed at understanding the importance of SprV in the S. mutans biology.IMPORTANCE Streptococcus mutans employs several virulence factors and stress resistance mechanisms to colonize tooth surfaces and cause dental caries. Bacterial pathogenesis is generally controlled by regulators of fitness that are critical for successful disease establishment. Sometimes these regulators, which are potential targets for antimicrobials, are lost in the genomic context due to the lack of annotated homologs. This work outlines the regulatory impact of a small, highly conserved hypothetical protein, SprV, encoded by S. mutans. We show that SprV affects the transcript levels of various virulence factors required for normal growth, biofilm formation, stress tolerance, genetic competence, and bacteriocin production.KEYWORDS pleiotropic regulator, Streptococcus mutans, bacteriocins, biofilms, global virulence, stress response S treptococcus mutans expresses various stress tolerance mechanisms and virulence factors, which allow the organism to endure the detrimental oral environment, colonize tooth surfaces as dental plaque, and eventually cause caries by demineralization of the enamel (1). The coordinated expression of stress responsive and virulencerelated genes is critical for the persistence of S. mutans in the oral environment that

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Small Proteins Can No Longer Be Ignored

Cited by 342 publications

References 107 publications

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

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

devIIs an Evolutionarily Young Negative Regulator of Myxococcus xanthus Development

Pleiotropic Regulation of Virulence Genes in Streptococcus mutans by the Conserved Small Protein SprV

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