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abstracts
Pseudomonas aeruginosa relies on cell motility and ability to form biofilms to establish infections, however, the mechanism of regulation remains obscure. Here we report that BswR, a XRE (Xenobiotic Response Element) type transcriptional regulator, plays a critical role in regulation of bacterial motility and biofilm formation in P. aeruginosa. Transcriptomic and biochemical analyses showed that BswR counteracts the repressor activity of MvaT, controls the transcription of small RNA rsmZ and regulates the biogenesis of bacterial flagella. The crystal structure of BswR was determined at 2.3 Å resolution; the monomer comprises a DNA-binding domain with a helix-turn-helix (HTH) motif in the N terminus and two helices (6 and 7) with a V-shaped arrangement in the C-terminus. In addition to the contacts between the parallel helices α5 of two monomers, the two helical extensions (6 and 7) intertwine together to form a homodimer, which is the biological function unit. Based on the result of DNase I protection assay together with structural analysis BswR homodimer, we proposed a BswR-DNA model, which suggests a molecular mechanism with which BswR could interact with DNA. Taken together, our results unveiled a novel regulatory mechanism, in which BswR controls the motility and biofilm formation of P. aeruginosa by modulating the transcription of small RNA rsmZ.
abstracts
5-hydroxymethylation is a mysterious modification of cytosine discovered decades before, with its functional roles awaiting elucidation in eukaryotes. Cumulative evidence demonstrates that 5-hydroxymethylcytosine is an epigenetic marker serves critical roles in multiple biological processes. Moreover, the profile of 5-hydroxymethylcytosine is changed under disease conditions such as cancer. Several methods including AbaSI-coupled sequencing have been developed to decipher hydroxymethylome at single-base resolution. However, the technical hurdles of AbaSI-coupled sequencing derived from the enzymatic property of AbaSI, a member belonging to PvuRts1I family endonuclease, may impede the application of this method. PvuRts1I is a modification dependent endonuclease with high selectivity to 5-hydroxymethylcytosine over 5-methylcytosine and cytosine. To improve the substrate selectivity of PvuRts1I family member, we solved the crystal structure of PvuRts1I. One nuclease domain and one SRA-like domain are located at the N- and C-terminal half of the structure, respectively. In comparison with other SRA domain structures, the SRA-like domain of PvuRts1I has been proposed to be the 5-hmC recognition module. Several mutants of PvuRts1I with enzymatic activity to 5-hydroxymethylcytosine only have been generated based on structural analysis, providing perfect candidates for dissection of hydroxymethylome from methylome.
