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Direct repeat sequences are implicated in the regulation of two Streptomyces chitinase promoters that are subject to carbon catabolite control.

ABSTRACT: We report the identification and partial characterization of the promoters for two chitinase genes from Streptomyces plicatus. Chitinases are a family of enzymes made by Streptomyces and other soil microbes to digest chitin, an abundant source of carbon and nitrogen in the soil. The promoter regions of two chitinases were defined by using transcriptional fusions to the xylE reporter gene. Transcription was shown to be glucose-sensitive and chitin-dependent. Each promoter contains a putative RNA polymerase binding site with a recognition sequence very similar to that observed in many eubacterial vegetatively expressed genes. In both promoters, a pair of 12-base-pair direct repeat sequences overlap the putative RNA polymerase binding sites. Further analysis of one of the promoters revealed that a single-base change within the direct repeat sequences resulted in glucose-resistant, chitin-independent expression in vivo. In addition, the promoter region that includes the direct repeat sequences was shown to interact with a sequence-specific DNA binding factor in vitro. Similar direct repeat sequences are present in other chitinase genes recently characterized, and we suggest that these repeats may be involved in repression and induction for this entire class of catabolite-controlled genes.

SUBMITTER: Delic I

PROVIDER: S-EPMC48558 | biostudies-other | 1992 Mar

REPOSITORIES: biostudies-other

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Direct repeat sequences are implicated in the regulation of two Streptomyces chitinase promoters that are subject to carbon catabolite control.

Delic I I Robbins P P Westpheling J J

Proceedings of the National Academy of Sciences of the United States of America 19920301 5

We report the identification and partial characterization of the promoters for two chitinase genes from Streptomyces plicatus. Chitinases are a family of enzymes made by Streptomyces and other soil microbes to digest chitin, an abundant source of carbon and nitrogen in the soil. The promoter regions of two chitinases were defined by using transcriptional fusions to the xylE reporter gene. Transcription was shown to be glucose-sensitive and chitin-dependent. Each promoter contains a putative RNA ...[more]

PMID: 1542688

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Project description:Ethanolamine (EA) is a compound prevalent in the gastrointestinal (GI) tract that can be used as a carbon, nitrogen, and/or energy source. Enterococcus faecalis, a GI commensal and opportunistic pathogen, contains approximately 20 ethanolamine utilization (eut) genes encoding the necessary regulatory, enzymatic, and structural proteins for this process. Here, using a chemically defined medium, two regulatory factors that affect EA utilization were examined. First, the functional consequences of loss of the small RNA (sRNA) EutX on the efficacy of EA utilization were investigated. One effect observed, as loss of this negative regulator causes an increase in eut gene expression, was a concomitant increase in the number of catabolic bacterial microcompartments (BMCs) formed. However, despite this increase, the growth of the strain was repressed, suggesting that the overall efficacy of EA utilization was negatively affected. Second, utilizing a deletion mutant and a complement, carbon catabolite control protein A (CcpA) was shown to be responsible for the repression of EA utilization in the presence of glucose. A predicted cre site in one of the three EA-inducible promoters, PeutS, was identified as the target of CcpA. However, CcpA was shown to affect the activation of all the promoters indirectly through the two-component system EutV and EutW, whose genes are under the control of the PeutS promoter. Moreover, a bioinformatics analysis of bacteria predicted to contain CcpA and cre sites revealed that a preponderance of BMC-containing operons are likely regulated by carbon catabolite repression (CCR).IMPORTANCE Ethanolamine (EA) is a compound commonly found in the gastrointestinal (GI) tract that can affect the behavior of human pathogens that can sense and utilize it, such as Enterococcus faecalis and Salmonella Therefore, it is important to understand how the genes that govern EA utilization are regulated. In this work, we investigated two regulatory factors that control this process. One factor, a small RNA (sRNA), is shown to be important for generating the right levels of gene expression for maximum efficiency. The second factor, a transcriptional repressor, is important for preventing expression when other preferred sources of energy are available. Furthermore, a global bioinformatics analysis revealed that this second mechanism of transcriptional regulation likely operates on similar genes in related bacteria.

Project description:BackgroundIn the genus Streptomyces, one of the most remarkable control mechanisms of physiological processes is carbon catabolite repression (CCR). This mechanism regulates the expression of genes involved in the uptake and utilization of alternative carbon sources. CCR also affects the synthesis of secondary metabolites and morphological differentiation. Even when the outcome effect of CCR in different bacteria is the same, their essential mechanisms can be quite different. In several streptomycetes glucose kinase (Glk) represents the main glucose phosphorylating enzyme and has been regarded as a regulatory protein in CCR. To evaluate the paradigmatic model proposed for CCR in Streptomyces, a high-density microarray approach was applied to Streptomyces coelicolor M145, under repressed and non-repressed conditions. The transcriptomic study was extended to assess the ScGlk role in this model by comparing the transcriptomic profile of S. coelicolor M145 with that of a ∆glk mutant derived from the wild-type strain, complemented with a heterologous glk gene from Zymomonas mobilis (Zmglk), insensitive to CCR but able to grow in glucose (ScoZm strain).ResultsMicroarray experiments revealed that glucose influenced the expression of 651 genes. Interestingly, even when the ScGlk protein does not have DNA binding domains and the glycolytic flux was restored by a heterologous glucokinase, the ScGlk replacement modified the expression of 134 genes. From these, 91 were also affected by glucose while 43 appeared to be under the control of ScGlk. This work identified the expression of S. coelicolor genes involved in primary metabolism that were influenced by glucose and/or ScGlk. Aside from describing the metabolic pathways influenced by glucose and/or ScGlk, several unexplored transcriptional regulators involved in the CCR mechanism were disclosed.ConclusionsThe transcriptome of a classical model of CCR was studied in S. coelicolor to differentiate between the effects due to glucose or ScGlk in this regulatory mechanism. Glucose elicited important metabolic and transcriptional changes in this microorganism. While its entry and flow through glycolysis and pentose phosphate pathway were stimulated, the gluconeogenesis was inhibited. Glucose also triggered the CCR by repressing transporter systems and the transcription of enzymes required for secondary carbon sources utilization. Our results confirm and update the agar model of the CCR in Streptomyces and its dependence on the ScGlk per se. Surprisingly, the expected regulatory function of ScGlk was not found to be as global as thought before (only 43 out of 779 genes were affected), although may be accompanied or coordinated by other transcriptional regulators. Aside from describing the metabolic pathways influenced by glucose and/or ScGlk, several unexplored transcriptional regulators involved in the CCR mechanism were disclosed. These findings offer new opportunities to study and understand the CCR in S. coelicolor by increasing the number of known glucose and ScGlk -regulated pathways and a new set of putative regulatory proteins possibly involved or controlling the CCR.

Dataset Information

Direct repeat sequences are implicated in the regulation of two Streptomyces chitinase promoters that are subject to carbon catabolite control.

Publications

Direct repeat sequences are implicated in the regulation of two Streptomyces chitinase promoters that are subject to carbon catabolite control.

Similar Datasets

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