Additional Targets of theBacillus subtilisGlobal Regulator CodY Identified by Chromatin Immunoprecipitation and Genome-Wide Transcript Analysis

Abstract

Additional targets of CodY, a GTP-activated repressor of early stationary-phase genes in Bacillus subtilis, were identified by combining chromatin immunoprecipitation, DNA microarray hybridization, and gel mobility shift assays. The direct targets of CodY newly identified by this approach included regulatory genes for sporulation, genes that are likely to encode transporters for amino acids and sugars, and the genes for biosynthesis of branched-chain amino acids.Bacteria have evolved a variety of mechanisms to accommodate gene expression to changes in nutritional availability. Some of these mechanisms are specific to a particular gene or operon. In other cases, regulatory proteins control large groups of genes of related function, such as the nitrogen metabolism genes regulated by the Ntr system in enteric bacteria (43) and by TnrA in Bacillus subtilis (17) and the carbon metabolism genes regulated by CcpA in gram-positive bacteria (13) and catabolic gene activator protein-cyclic AMP complex in gram-negative bacteria (59). Even broader forms of regulation are mediated by the leucine-responsive protein (Lrp) of gram-negative bacteria and the sigma-B protein of B. subtilis. Lrp and sigma-B control the transcription of operons that have diverse functions but have a common need to be expressed under a particular set of environmental conditions (50, 54). Lrp regulates the biosynthesis of leucine, isoleucine, valine, serine, glycine, and glutamate; the degradation of serine and threonine; transport of peptides, amino acids, and sugars; and production of fimbriae in response to the availability of leucine and serine (50). Sigma-B activates transcription of a host of genes when cells are exposed to excessive heat, ethanol, salt, or acid (54). Sigma-B responds through a complex, multibranched signal transduction pathway.The B. subtilis CodY protein also has broad effects on gene expression. CodY is a GTP-binding repressor of several genes that are normally quiescent when cells are growing in a rich medium (57). A high concentration of GTP activates CodY as a repressor (57). When the growth rate of B. subtilis slows down because of limitation of the carbon or nitrogen or phosphorus source, the GTP level drops (39, 40), CodY loses repressing activity, and targets of CodY repression are transcribed. The known targets of CodY in B. subtilis include the genes that encode transport systems for dipeptides (dpp) (65) and ␥-aminobutyrate (gabP) (16); catabolic pathways for acetate (acsA) (S. H. Fisher, personal communication), urea (ureABC) (71), histidine (hut) (18), arginine (rocABC and roc-DEF) (B. Belitsky, personal communication), and branchedchain keto acids (the bkd operon) (12); an enzyme of surfactin synthesis (srfAA) (63); the transcription factor for DNA uptake genes (comK) (63); a ComA aspartyl phosphate phosphatase and its inhibitor (rapC-phrC) (37); motility and chemotaxis (hag, fla/che) (45; F. Bergara, C. Ibarra, J. Iwamasa, R. Aguilera, and L. M. Màrquez-Magaña, submitted for publication); and aconitase (citB) (3...

Transcriptome Analysis of Sorbic Acid-Stressed Bacillus subtilis Reveals a Nutrient Limitation Response and Indicates Plasma Membrane Remodeling

“…This gene is known to be regulated only by CodY (52). The induction of transition-state and early-stationary-phase genes suggests that sorbic acidstressed cells respond as if they experience nutrient limitation.…”

Carbon Catabolite Control of the Metabolic Network inBacillus subtilis

“…119) Besides the probable necessity that the CcpA-dependent CCA of ilv-leu in glycolysis proceed continuously for the draining of accumulated pyruvate, it is notable that this positive regulation links carbon metabolism to amino acid anabolism. Recent global gene expression studies on amino acid availability 120) and CodY regulation, 121) as well as on the metabolic links of ilv-leu expression to glucose and nitrogen metabolism, 38,39,98,122) indicate that the ilv-leu operon is under direct negative transcriptional control through two major global regulators of nitrogen metabolism (CodY and TnrA).…”

Genome-wide identification of Bacillus subtilis CodY-binding sites at single-nucleotide resolution

“…C odY is a dimeric 259-residue Bacillus subtilis protein that binds to DNA through a winged helix-turn-helix domain (1) and acts as a global transcriptional regulator to control expression of many metabolic genes (2)(3)(4)(5). The DNA-binding activity of CodY is enhanced by interaction with two types of effectors, branched-chain amino acids [isoleucine, leucine, and valine (ILV)] (6, 7) and GTP (3,(7)(8)(9).…”

“…The DNA-binding activity of CodY is enhanced by interaction with two types of effectors, branched-chain amino acids [isoleucine, leucine, and valine (ILV)] (6, 7) and GTP (3,(7)(8)(9). CodY binding requires the presence of a 15-nt canonical consensus motif, AATTTTCW-GAAAATT (10)(11)(12).…”