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RNA-seq of zinc-replete wild type and zinc-limited wild type, altRP operon deletion mutant and its complement strain in Mycolicibacterium (Mycobacterium) smegmatis

ABSTRACT: RNA-seq of zinc-replete wild type and zinc-limited wild type, altRP operon deletion mutant and its complement strain in Mycolicibacterium (Mycobacterium) smegmatis

PROVIDER: PRJNA777953 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR16775601_1.fastq.gz	Fastqsanger.gz
	SRR16775601_2.fastq.gz	Fastqsanger.gz
	SRR16775602_1.fastq.gz	Fastqsanger.gz
	SRR16775602_2.fastq.gz	Fastqsanger.gz
	SRR16775603_1.fastq.gz	Fastqsanger.gz

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Project description:We assessed changes in gene expression in response to zinc availability for the human pathogen Corynebacterium diphtheriae. Expression profiles of wild-type C. diphtheriae strain 1737 grown in semi-defined metal-poor media (mPGT) without and with 5 µM zinc chloride supplementation were compared; the expression profile of wild-type C. diphtheriae strain 1737 in zinc-replete conditions was also compared against an isogenic Δzur mutant grown in zinc-replete conditions. Three biological replicates were prepared by isolating total RNA from mid-logarithmic growth cultures and eleven genes (dip0013, dip0093, dip0173, dip0438, dip1087, dip1486, dip1724, dip2114, dip2128, dip2162, and dip2325) were assessed by real-time PCR to validate the array results. (Abstract) Corynebacterium diphtheriae is a Gram-positive bacterial pathogen and the causative agent of diphtheria, a severe disease of the upper respiratory tract of humans. Factors required for C. diphtheriae to survive in the human host are not well defined, but likely include the acquisition of essential metals such as zinc. In C. diphtheriae, zinc-responsive global gene regulation is controlled by the Zinc Uptake Regulator (Zur), a member of the Fur-family of transcriptional regulators. In this study, we use transcriptomics to identify zinc-regulated genes in C. diphtheriae by comparing gene expression of a wild-type strain grown without and with zinc supplementation. Zur-regulated genes were identified by comparing wild-type gene expression with that of an isogenic zur mutant. We observed zinc repression of several putative surface proteins, the heme efflux system hrtBA, various ABC transporters, and the non-ribosomal peptide synthetase/polyketide synthase cluster sidAB. Furthermore, increased gene expression in response to zinc was observed for the alcohol dehydrogenase, adhA. Zinc and Zur regulation were confirmed for several genes by complementing the zur deletion and subsequent qPCR analysis. We used MEME to predict Zur binding sites within the promoter regions of zinc- and Zur-regulated genes, and verified Zur binding by electrophoretic mobility shift assays. Additionally, we characterized cztA (dip1101), which encodes a putative cobalt/zinc/cadmium efflux family protein. Deletion of cztA results in increased sensitivity to zinc, but not to cobalt or cadmium. This study advances our knowledge of changes to Zur-dependent global gene expression in response to zinc in C. diphtheriae. The identification of zinc-regulated ABC transporters herein will facilitate future studies to characterize zinc transport in C. diphtheriae.

Dataset Information

RNA-seq of zinc-replete wild type and zinc-limited wild type, altRP operon deletion mutant and its complement strain in Mycolicibacterium (Mycobacterium) smegmatis

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