Complex Regulation of the Phosphoenolpyruvate Carboxykinase Gene pck and Characterization of Its GntR-Type Regulator IolR as a Repressor of myo-Inositol Utilization Genes in Corynebacterium glutamicum

Abstract

c DNA affinity chromatography with the promoter region of the Corynebacterium glutamicum pck gene, encoding phosphoenolpyruvate carboxykinase, led to the isolation of four transcriptional regulators, i.e., RamA, GntR1, GntR2, and IolR. Determination of the phosphoenolpyruvate carboxykinase activity of the ⌬ramA, ⌬gntR1 ⌬gntR2, and ⌬iolR deletion mutants indicated that RamA represses pck during growth on glucose about 2-fold, whereas GntR1, GntR2, and IolR activate pck expression about 2-fold irrespective of whether glucose or acetate served as the carbon source. The DNA binding sites of the four regulators in the pck promoter region were identified and their positions correlated with the predicted functions as repressor or activators. The iolR gene is located upstream and in a divergent orientation with respect to a iol gene cluster, encoding proteins involved in myoinositol uptake and degradation. Comparative DNA microarray analysis of the ⌬iolR mutant and the parental wild-type strain revealed strongly (>100-fold) elevated mRNA levels of the iol genes in the mutant, indicating that the primary function of IolR is the repression of the iol genes. IolR binding sites were identified in the promoter regions of iolC, iolT1, and iolR. IolR therefore is presumably subject to negative autoregulation. A consensus DNA binding motif (5=-KGWCHTRACA-3=) which corresponds well to those of other GntR-type regulators of the HutC family was identified. Taken together, our results disclose a complex regulation of the pck gene in C. glutamicum and identify IolR as an efficient repressor of genes involved in myo-inositol catabolism of this organism.

A unique gene cluster for the utilization of the mucosal and human milk‐associated glycans galacto‐N‐biose and lacto‐N‐biose in Lactobacillus casei

“…Unlike Bifidobacterium species, in L. casei the gnb cluster containing the genes involved in the metabolism of those disaccharides does not encode sugar kinases. We showed that the genes gnbF, encoding GalNAc-6P deacetylase, and nagA, encoding GlcNAc-6P deacetylase, were involved in the metabolism of GNB and LNB, respectively, which indicates that the GalNAc and GlcNAc resulting from (Hu et al, 1989); FarR (Quail et al, 1994); TreRo1 and TreRo2 (Schöck and Dahl, 1996); KorSA (Sezonov et al, 2000); NagRdre nagP and NagRdre nagA (Bertram et al, 2011); PckR (Hyeon et al, 2012); IolR (Klaffl et al, 2013). Bases conserved in all sequences are shown against a dark background.…”

Genome-Wide Analysis of the Role of Global Transcriptional Regulator GntR1 in Corynebacterium glutamicum

“…Seventy genes were upregulated more than 1.5-fold, whereas 229 genes were downregulated more than 1.5-fold (see Table S2 in the supplemental material). Decreased expression of ptsG and pckA and increased expression of tkt, gnd, gntK, gntP, and gntV were observed as described previously (13)(14)(15). In addition to these genes, we found that the expression of many of the genes involved in carbon metabolism was changed; these genes are gapB, rbsK2, rbsK1, dctP2, ugpE, dhaS, maeB, dctA, dctM1, dctP1, bglF2, and bglF.…”

“…GntR1/2 is known to control the expression of genes of gluconate utilization (gntP and gntK), the pentose phosphate pathway (tkt, tal, zwf, opcA, devB, and gnd), the phosphotransferase system (ptsG and ptsS), and gluconeogenesis (pckA) in C. glutamicum ATCC 13032, indicating that GntR1/2 is one of the important transcriptional regulators for carbon metabolism (13)(14)(15). In C. glutamicum R, the gntR2 gene is missing and gntR1 is solely responsible for the induction of the gnd gene by gluconate (14).…”

“…Of these regions, 51 sites are located upstream of coding sequences, suggesting that GntR1 regulates transcription of these genes. The identified 56 sites include the upstream regions of genes gntP, gntK, gnd, tkt, ptsG, ptsS, and pckA, the regions that GntR1 was shown to bind to in vitro (13)(14)(15).…”

“…The dual functions of GntR1/2, repression of gluconate utilization and activation of the PTS-dependent sugar uptake, probably contribute to the simultaneous utilization of gluconate and glucose in C. glutamicum. Recently, it was reported that the pck gene, encoding phosphoenolpyruvate carboxykinase, is also transcriptionally activated by GntR1/2 (15). Microarray analysis revealed that disruption of gntR1/2 resulted in the upregulation of 19 genes and downregulation of 26 genes (13).…”