The Phosin PptA Plays a Negative Role in the Regulation of Antibiotic Production in Streptomyces lividans

Abstract

In Streptomyces, antibiotic biosynthesis is triggered in phosphate limitation that is usually correlated with energetic stress. Polyphosphates constitute an important reservoir of phosphate and energy and a better understanding of their role in the regulation of antibiotic biosynthesis is of crucial importance. We previously characterized a gene, SLI_4384/ppk, encoding a polyphosphate kinase, whose disruption greatly enhanced the weak antibiotic production of Streptomyces lividans. In the condition of energetic stress, Ppk utilizes polyP as phosphate and energy donor, to generate ATP from ADP. In this paper, we established that ppk is co-transcribed with its two downstream genes, SLI_4383, encoding a phosin called PptA possessing a CHAD domain constituting a polyphosphate binding module and SLI_4382 encoding a nudix hydrolase. The expression of the ppk/pptA/SLI_4382 operon was shown to be under the positive control of the two-component system PhoR/PhoP and thus mainly expressed in condition of phosphate limitation. However, pptA and SLI_4382 can also be transcribed alone from their own promoter. The deletion of pptA resulted into earlier and stronger actinorhodin production and lower lipid content than the disruption of ppk, whereas the deletion of SLI_4382 had no obvious phenotypical consequences. The disruption of ppk was shown to have a polar effect on the expression of pptA, suggesting that the phenotype of the ppk mutant might be linked, at least in part, to the weak expression of pptA in this strain. Interestingly, the expression of phoR/phoP and that of the genes of the pho regulon involved in phosphate supply or saving were strongly up-regulated in pptA and ppk mutants, revealing that both mutants suffer from phosphate stress. Considering the presence of a polyphosphate binding module in PptA, but absence of similarities between PptA and known exo-polyphosphatases, we proposed that PptA constitutes an accessory factor for exopolyphosphatases or general phosphatases involved in the degradation of polyphosphates into phosphate.

A Proteomic Analysis Indicates That Oxidative Stress Is the Common Feature Triggering Antibiotic Production in Streptomyces coelicolor and in the pptA Mutant of Streptomyces lividans

“…Our analysis revealed that proteins of most ontological classes of the two SL strains had a similar abundance and responded in a similar, but not identical, way to phosphate availability and thus belonged to the same groups. The slight differences in the response to Pi availability between the two SL strains are thought to be due to the previously reported upregulation of proteins of the Pho regulon involved in Pi scavenging and transport, in the pptA mutant in a condition of Pi limitation ( Shikura et al, 2021 ). However, interestingly, the main difference between the wt strain of SL and its pptA mutant concerned enzymes involved in nicotinamide adenine dinucleotide phosphate (reduced form) [NADP(H)] metabolism.…”

“…However, the extensively studied model strain, Streptomyces coelicolor ( SC ), escapes this general rule since it produces the blue pigmented polyketide antibiotic, actinorhoddin (ACT), in both phosphate conditions and even more abundantly in phosphate proficiency than in phosphate limitation ( Esnault et al, 2017 ). In contrast, Streptomyces lividans ( SL ), a strain phylogenetically closely related to SC , which has the genetic capability to produce ACT, does not produce it in any Pi condition, whereas the pptA mutant derived from it produces ACT in phosphate limitation but not in phosphate proficiency ( Shikura et al, 2021 ), as most Streptomyces species. The pptA mutant is an in-frame deletion of sco4144 that encodes a phosin-like protein possessing a polyphosphate (polyP) binding module (CHAD domain) ( Werten et al, 2019 ).…”

“…The pptA mutant is an in-frame deletion of sco4144 that encodes a phosin-like protein possessing a polyphosphate (polyP) binding module (CHAD domain) ( Werten et al, 2019 ). PptA, which is mainly expressed in conditions of phosphate limitation and belongs to the Pho regulon ( Shikura et al, 2021 ), was proposed to act as an accessory factor forcing polyP into a conformation allowing their efficient use by various enzymes using polyP as donor of phosphate and/or energy ( Werten et al, 2019 ). These potentially include exo-polyphosphatases or general phosphatases involved in the degradation of polyP into phosphate ( Shikura et al, 2021 ) or the polyphosphate kinase Ppk/SCO4145 that uses polyP to regenerate ATP from ADP ( Chouayekh and Virolle, 2002 ; Ghorbel et al, 2006b ).…”

“…PptA, which is mainly expressed in conditions of phosphate limitation and belongs to the Pho regulon ( Shikura et al, 2021 ), was proposed to act as an accessory factor forcing polyP into a conformation allowing their efficient use by various enzymes using polyP as donor of phosphate and/or energy ( Werten et al, 2019 ). These potentially include exo-polyphosphatases or general phosphatases involved in the degradation of polyP into phosphate ( Shikura et al, 2021 ) or the polyphosphate kinase Ppk/SCO4145 that uses polyP to regenerate ATP from ADP ( Chouayekh and Virolle, 2002 ; Ghorbel et al, 2006b ). The pptA mutant thus suffers phosphate/energetic stress, and a strong induction of the expression of the two-component system (TCS) PhoR/PhoP that controls positively the expression of genes of the Pho regulon involved in phosphate scavenging and uptake was observed in this strain ( Shikura et al, 2021 ).…”

“…These potentially include exo-polyphosphatases or general phosphatases involved in the degradation of polyP into phosphate ( Shikura et al, 2021 ) or the polyphosphate kinase Ppk/SCO4145 that uses polyP to regenerate ATP from ADP ( Chouayekh and Virolle, 2002 ; Ghorbel et al, 2006b ). The pptA mutant thus suffers phosphate/energetic stress, and a strong induction of the expression of the two-component system (TCS) PhoR/PhoP that controls positively the expression of genes of the Pho regulon involved in phosphate scavenging and uptake was observed in this strain ( Shikura et al, 2021 ). In contrast, the expression of PhoR/PhoP and proteins under its positive control was shown to be lower in SC than in SL in Pi limitation ( Millan-Oropeza et al, 2020 ), and SC has a 2–3-fold higher intracellular ATP content than SL ( Esnault et al, 2017 ).…”