Review of: "The nucleotide messenger (p)ppGpp is an anti-inducer of the purine synthesis transcription regulator PurR in Bacillus"
Year: 2022
Type: peer-review
Abstract: Potential competing interests: The author(s) declared that no potential competing interests exist.This is an outstanding article which investigates the first-known direct interaction of the nucleotide messenger and alarmone guanosine pentaphosphate, (p)ppGpp, with a transcription factor in Bacillus.Specifically, (p)ppGpp was found to be able to bind the PurR repressor's effector binding domain to allosterically compete with PurR's previously known inducer, phosphoribosyl pyrophosphate (PRPP).A thorough set of experiments were performed to dissect the overlapping binding domains of (p)ppGpp and PRPP, the effect of (p)ppGpp competition with PRPP on PurR-DNA interaction, the promoter occupancy at PurR binding sites across the B. subtilis genome following (p)ppGpp induction, and the anti-induction of purine biosynthesis genes by (p)ppGpp through PurR repressor activity during entry of cells into stationary phase.Altogether, the findings show that (p)ppGpp can directly regulate nucleotide synthesis in Bacillus by binding to a classical transcription repressor.While the concept of (p)ppGpp-induced inhibition of purine biosynthesis is established, the discovery that (p)ppGpp can directly modulate repressor activity introduces a new mechanism whereby (p)ppGpp relays stress-response signaling in Bacillus and highlights the versatility of (p)ppGpp as a second messenger.This work will allow the pursuit of novel experimental designs for future investigations of (p)ppGpp interactions both with PurR as well as with other protein candidates.Major strengths of this study include the thorough structural and physiological profiling of an interactor of (p)ppGpp, PurR, identified from a forward screen of the entire Bacillus proteome.Experiments methodically dissected the nature of the interaction of PurR with (p)ppGpp and the functional consequences of this interaction on PurR repressor activity in the context of nutrient stress adaptation. 1. Differential radial capillary action of ligand assay (DRaCALA) 1 , a tried and true method to detect interactors of low-molecular-weight ligands, was used to detect (p)ppGpp interactors in a proteomewide screen.Non-radiolabeled GTP was added as a competitor to enrich for ppGpp-specific interactors.DRaCALA is generally highly specific.False positives may occur due to spotting errors but can easily be identified visually and removed from analysis 2 .Validation of hits using purified protein further corrects for false positives 3 .In this study, PurR binding signal was enriched after GTP competitor addition, and Qeios, CC-BY 4.
DOI: 10.32388/ylh8ef
Authors: Christophe Herman, Xueyin Wen
Institution: Baylor College of Medicine
Language: English
Subfield: Genetics
Domain: Life Sciences
Open Access status: gold
OpenAlex ID: W4214875908