A document by Gabriele Pradel. Click on the document to view its contents.

RNA-binding proteins (RBPs) govern the lifespan of nearly all transcripts and play key roles in adaptive responses in microbes. A robust approach to examine protein-RNA interactions involves irradiating cells with UV light to form covalent adducts between RBPs and their cognate RNAs. Combined with RNA or protein purification, these procedures can provide global RBP censuses or transcriptomic maps for all target sequences of a single protein in living cells. Recent development of novel methods has quickly populated the RBP landscape in microorganisms. Here, we provide an overview of prominent UV cross-linking techniques which have been applied to investigate RNA interactomes in microbes. By assessing their advantages and caveats, this technical evaluation intends to guide the selection of appropriate methods and experimental design as well as to encourage the use of complementary UV-dependent techniques to inspect RNA-binding activity.

DNA glycosylases protect genetic fidelity during DNA replication by removing potentially mutagenic chemically damaged DNA bases. Bacterial Lhr proteins are well-characterized DNA repair helicases that are fused to additional 600-700 amino acids of unknown function, but with structural homology to SecB chaperones and AlkZ DNA glycosylases. Here we identify that E. coli Lhr is a uracil-DNA glycosylase that depends on an active site aspartic acid residue. We show that the Lhr DNA helicase activity is functionally independent of the uracil-DNA glycosylase activity, but that the helicase domains are required for fully active uracil DNA glycosylase activity. Consistent with uracil DNA glycosylase activity, deletion of lhr from the E. coli chromosome sensitized cells to oxidative stress that triggers cytosine deamination to uracil. The ability of Lhr to translocate single-stranded DNA and remove uracil bases suggests a surveillance role to seek and remove potentially mutagenic base changes during replication stress.

In the last two decades, an increasing number of bacterial species have been recognized that are able to generate a phenotypically diverse population that shares an identical genotype. This ability is dependent on a complex genetic regulatory network that includes cellular and environmental signals, as well as stochastic elements. Among Bacilli, a broadly distributed family of Rap (Response-regulator aspartyl phosphate) phosphatases is known to modulate the function of the main phenotypic heterogeneity regulators by controlling their phosphorylation. Even more, their related extracellular Phr (Phosphatase regulator) peptides function as quorum sensing signals, creating a cell-cell communication network that regulates the phenotypic development of the entire population. In this review, we examine the role that the Rap phosphatases and their Phr peptides play in the regulation of B. subtilis phenotypic differentiation, and in other members of the Bacillus genus. We also highlight the contribution of these regulatory elements to the fitness of bacterial cells and mobile genetic elements, e.g. prophages and conjugative vectors.