Dataset Information

The PolS-PolR Two-Component System Regulates Genes Involved in Poly-P Metabolism and Phosphate Transport in Microlunatus phosphovorus.

ABSTRACT: Microlunatus phosphovorus NM-1 is a polyphosphate (poly-P)-accumulating bacterium that accumulates poly-P under aerobic conditions and degrades poly-P under anaerobic conditions. In this study, the two-component system (TCS) PolS-PolR was identified in NM-1, and the response regulator PolR was found to directly bind to the promoters of genes related to phosphate transport (MLP_RS00235, MLP_RS23035, and MLP_RS24590); poly-P catabolism (MLP_RS12905) and poly-P synthesis (MLP_RS23025). RT-qPCR assays showed that ppgk (MLP_RS12905), ppk (MLP_RS23025), pstS (MLP_RS23035), and pit (MLP_RS24590) were down-regulated during the aerobic-anaerobic shift. The sequence GTTCACnnnnnGTTCaC was identified as a recognition sequence for PolR by MEME analysis and DNase I footprinting. EMSAs and ChIP-qPCR assays indicated that PolR binds to the promoters of pit (MLP_RS00235), ppgk (MLP_RS12905), ppk (MLP_RS23025), pstS (MLP_RS23035) and pit (MLP_RS24590), and ChIP-qPCR further suggested that the binding affinity of PolR was lower under anaerobic conditions than under aerobic conditions in vivo. These findings indicate that the PolS-PolR TCS in M. phosphovorus may be involved in the regulation of poly-P metabolism in response to levels of dissolved oxygen in the environment, and our results provide insights into new approaches for understanding the mechanisms of phosphorus accumulation and release.

SUBMITTER: Zhong C

PROVIDER: S-EPMC6754071 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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The PolS-PolR Two-Component System Regulates Genes Involved in Poly-P Metabolism and Phosphate Transport in <i>Microlunatus phosphovorus</i>.

Zhong Chuanqing C Zhang Peipei P Liu Cheng C Liu Meng M Chen Wenbing W Fu Jiafang J Qi Xiaoyu X Cao Guangxiang G

Frontiers in microbiology 20190913

<i>Microlunatus phosphovorus</i> NM-1 is a polyphosphate (poly-P)-accumulating bacterium that accumulates poly-P under aerobic conditions and degrades poly-P under anaerobic conditions. In this study, the two-component system (TCS) PolS-PolR was identified in NM-1, and the response regulator PolR was found to directly bind to the promoters of genes related to phosphate transport (MLP_RS00235, MLP_RS23035, and MLP_RS24590); poly-P catabolism (MLP_RS12905) and poly-P synthesis (MLP_RS23025). RT-qP ...[more]

PMID: 31572333

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Project description:In bacteria, limited phosphate availability promotes the synthesis of active uptake systems, such as the Pst phosphate transport system. To understand the mechanisms that facilitate phosphate accumulation in the cyanobacterium Nostoc punctiforme, phosphate transport systems were identified, revealing a redundancy of Pst phosphate uptake systems that exists across three distinct operons. Four separate PstB system components were identified. pstB1 was determined to be a suitable target for creating phenotypic mutations that could result in the accumulation of excessive levels of phosphate through its overexpression or in a reduction of the capacity to accumulate phosphate through its deletion. Using quantitative real-time PCR (qPCR), it was determined that pstB1 mRNA levels increased significantly over 64 h in cells cultured in 0 mM added phosphate and decreased significantly in cells exposed to high (12.8 mM) phosphate concentrations compared to the level in cells cultured under normal (0.8 mM) conditions. Possible compensation for the loss of PstB1 was observed when pstB2, pstB3, and pstB4 mRNA levels increased, particularly in cells starved of phosphate. The overexpression of pstB1 increased phosphate uptake by N. punctiforme and was shown to functionally complement the loss of PstB in E. coli PstB knockout (PstB-) mutants. The knockout of pstB1 in N. punctiforme did not have a significant effect on cellular phosphate accumulation or growth for the most part, which is attributed to the compensation for the loss of PstB1 by alterations in the pstB2, pstB3, and pstB4 mRNA levels. This study provides novel in vivo evidence that PstB1 plays a functional role in phosphate uptake in N. punctiforme IMPORTANCE: Cyanobacteria have been evolving over 3.5 billion years and have become highly adept at growing under limiting nutrient levels. Phosphate is crucial for the survival and prosperity of all organisms. In bacteria, limited phosphate availability promotes the synthesis of active uptake systems. The Pst phosphate transport system is one such system, responsible for the internalization of phosphate when cells are in phosphate-limited environments. Our investigations reveal the presence of multiple Pst phosphate uptake systems that exist across three distinct operons in Nostoc punctiforme and functionally characterize the role of the gene product PstB1 as being crucial for the maintenance of phosphate accumulation. We demonstrate that the genes pstB2, pstB3, and pstB4 show alterations in expression to compensate for the deletion of pstB1 The overall outcomes of this work provide insights as to the complex transport mechanisms that exist in cyanobacteria like N. punctiforme, allowing them to thrive in low-phosphate environments.

Project description:Mycobacterium smegmatis and several other mycobacteria are able to utilize methanol as the sole source of carbon and energy. We recently showed that N,N-dimethyl-p-nitrosoaniline (NDMA)-dependent methanol dehydrogenase (Mno) is essential for the growth of M. smegmatis on methanol. Although Mno from this bacterium shares high homology with other known methanol dehydrogenases, methanol metabolism in M. smegmatis differs significantly from that of other described methylotrophs. In this study, we dissect the regulatory mechanism involved in the methylotrophic metabolism in M. smegmatis We identify a two-component system (TCS), mnoSR, that is involved in the regulation of mno expression. We show that the MnoSR TCS is comprised of a sensor kinase (MnoS) and a response regulator (MnoR). Our results demonstrate that MnoS undergoes autophosphorylation and is able to transfer its phosphate to MnoR by means of phosphotransferase activity. Furthermore, MnoR shows specific binding to the putative mno promoter region in vitro, thus suggesting its role in the regulation of mno expression. Additionally, we find that the MnoSR system is involved in the regulation of MSMEG_6239, which codes for a putative 1,3-propanediol dehydrogenase. We further show that M. smegmatis lacking mnoSR is unable to utilize methanol and 1,3-propanediol as the sole carbon source, which confirms the role of MnoSR in the regulation of alcohol metabolism. Our data, thus, suggest that the regulation of mno expression in M. smegmatis provides new insight into the regulation of methanol metabolism, which furthers our understanding of methylotrophy in mycobacteria.IMPORTANCE Methylotrophic metabolism has gained huge attention considering its broad application in ecology, agriculture, industries, and human health. The genus Mycobacterium comprises both pathogenic and nonpathogenic species. Several members of this genus are known to utilize methanol as the sole carbon source for growth. Although various pathways underlying methanol utilization have been established, the regulation of methylotrophic metabolism is not well studied. In the present work, we explore the regulation of methanol metabolism in M. smegmatis and discover a dedicated two-component system (TCS), MnoSR, that is involved in its regulation. We show that the loss of MnoSR renders the bacterium incapable of utilizing methanol and 1,3-propanediol as the sole carbon sources. Additionally, we establish that MnoS acts as the common sensor for the alcohols in M. smegmatis.

Dataset Information

The PolS-PolR Two-Component System Regulates Genes Involved in Poly-P Metabolism and Phosphate Transport in Microlunatus phosphovorus.

Publications

The PolS-PolR Two-Component System Regulates Genes Involved in Poly-P Metabolism and Phosphate Transport in <i>Microlunatus phosphovorus</i>.

Similar Datasets

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