Activation of chloramphenicol biosynthesis in Streptomyces venezuelae ATCC 10712 by ethanol shock: insights from the promoter fusion studies.

Olga N Sekurova,Jianhai Zhang,Sergey B Zotchev,Kåre A Kristiansen

doi:10.1186/s12934-016-0484-9

Olga N Sekurova, Jianhai Zhang + Show 2 more

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https://doi.org/10.1186/s12934-016-0484-9

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Abstract

BackgroundStreptomyces venezuelae ATCC 10712 produces antibiotics chloramphenicol (Cml) and jadomycin (Jad) in response to nutrient limitation and ethanol shock (ES), respectively. Biosynthesis of Cml and Jad was shown to be reciprocally regulated via the action of regulatory proteins JadR1 and JadR2 encoded by the jad cluster, and mechanism of such regulation has been characterized. However, detailed analysis of the regulatory mechanism controlling Cml biosynthesis is still lacking.ResultsIn the present study, several promoters from the cml cluster were fused to the reporter gene gusA. Reporter protein activity and Cml production were assayed in the wild-type strain with and without ES, followed by similar experiments with the jadR1 deletion mutant. The latter gene was earlier reported to negatively control Cml biosynthesis, while serving as a positive regulator for the jad cluster. A double deletion mutant deficient in both jadR1 and the cml cluster was also constructed and used in promoter fusion studies. Analyses of the results revealed that ES activates Cml biosynthesis in both wild-type and jadR1 deletion mutant, while Cml production by the latter was ca 80 % lower.ConclusionsThese results contradict earlier reports regarding the function of JadR1, but correlate well with the reporter activity data for some promoters, while reaction of others to the ES is genotype-dependent. Remarkably, the absence of Cml production in the double mutant has a profound effect on the way certain cml promoters react to ES. The latter suggests direct involvement of Cml in this complex regulatory mechanism.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-016-0484-9) contains supplementary material, which is available to authorized users.

Highlights

Streptomyces venezuelae ATCC 10712 produces antibiotics chloramphenicol (Cml) and jadomycin (Jad) in response to nutrient limitation and ethanol shock (ES), respectively
Cml production could not be detected in S. venezuelae, and cmlR transcription level was very low in the conditions tested, suggesting the latter being the main reason for Cml non-producing phenotype
According to the currently accepted model for reciprocal regulation of Cml and Jad biosynthesis, no Cml production is possible upon ES due to the repression of cml promoters by the JadR1 regulatory protein [4, 5]

Summary

Introduction

Streptomyces venezuelae ATCC 10712 produces antibiotics chloramphenicol (Cml) and jadomycin (Jad) in response to nutrient limitation and ethanol shock (ES), respectively. Biosynthesis of secondary metabolites in Gram-positive bacteria of the genus Streptomyces is controlled at several levels, involving global, pleiotropic, and pathwayspecific regulators These regulators represent an intricate network that switches on biosynthesis of secondary metabolites, which is energy- and resourcedemanding process, in response to particular environmental stimuli. The latter can be nutrient deprivation, The intriguing mechanism behind regulation of Jad biosynthesis has recently been revealed, and involves a complex interaction between four regulatory proteins, JadR1, JadR2, JadR3 and JadR* encoded within the jad biosynthetic gene cluster [4,5,6,7,8]. Heterologous expression of the cml cluster in Streptomyces coelicolor was demonstrated, and deletion of cmlR led to complete abrogation of Cml biosynthesis, confirming the role of CmlR as a positive regulator and implying that in S. venezuelae cmlR is repressed

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