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

Noriyasu Shikura,Naima Nhiri,Eric Jacquet,Sebastiaan Werten,Laila Sago,Cécile Martel,Clara Lejeune,David Cornu,Catherine Esnault,Delin Xu,Marie-Joelle Virolle,Emmanuelle Darbon,Ariane Deniset-Besseau

doi:10.3390/antibiotics10030325

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.

Highlights

Streptomyces are Gram-positive, filamentous soil bacteria, of great medical and economic importance since they are able to produce a great variety of bio-active molecules useful to human health or agriculture [1,2]
SLI_4383/pptA (GenBank entry EFD67904.1) to putative orthologues from other StreptoInterestingly, a comparison of the predicted protein product of SLI_4383/pptA (GenBank myces species revealed the presence of N-terminal extensions of various length whose entry EFD67904.1) to putative orthologues from other Streptomyces species revealed the extremity is highly conserved in almost all orthologues but which is missing in the propresence of N-terminal extensions of various length whose extremity is highly conserved tein of S. lividans as it is annotated at the Sanger center (Figure 1)
In order to establish the impact of phosphate availability on the transcriptional regulation of the genes of this operon, RNA was prepared from wt S. lividans, its phoP mutant and wt S. coelicolor grown for 40 h on solid R2YE medium with no phosphate added

Summary

Introduction

Streptomyces are Gram-positive, filamentous soil bacteria, of great medical and economic importance since they are able to produce a great variety of bio-active molecules useful to human health or agriculture [1,2]. The biosynthesis of these molecules usually takes place when growth slows down or stops and is thought to be triggered by some nutritional limitation [3]. The accumulation of large quantities of polyP was noticed in phoU mutants of E. coli [10] and was correlated with an activation of Pi up-take by the PstSCAB transport system.

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Results

Conclusion