Transcriptome Analysis of Acetate Metabolism inCorynebacterium glutamicumUsing a Newly Developed Metabolic Array

Abstract

Following the determination of the whole-genome sequence of Corynebacterium glutamicum, we have developed a DNA array to extensively investigate gene expression and regulation relevant to carbon metabolism. For this purpose, a total of 120 C. glutamicum genes, including those in central metabolism and amino acid biosyntheses, were amplified by PCR and printed onto glass slides. The resulting array, designated a "metabolic array", was used for hybridization with fluorescently labeled cDNA probes generated by reverse transcription from total RNA samples. As the first demonstration of transcriptome analysis in this industrially important microorganism, we applied the metabolic array to study differential transcription profiles between cells grown on glucose and on acetate as the sole carbon source. The changes in gene expression observed for the known acetate-regulated genes (aceA, aceB, pta, and ack) were well consistent with the literature data of northern analyses and enzyme assays, indicating the utility of the metabolic array in transcriptome analysis of C. glutamicum. In addition to the known responses, many previously unrecognized co-regulated genes were identified. For example, several TCA cycle genes, such as gltA, sdhA, sdhB, fumH, and mdh, and the gluconeogenic gene pck were up-regulated in the acetate medium. On the other hand, a few genes involved in glycolysis and the pentose phosphate pathway, as well as many amino acid biosynthetic genes, were down-regulated in acetate. Furthermore, two gap genes, gapA and gapB, were found to be inversely regulated, suggesting the presence of a new regulatory step for carbon metabolism between glycolysis and gluconeogenesis.

RamB, a Novel Transcriptional Regulator of Genes Involved in Acetate Metabolism of Corynebacterium glutamicum

“…In addition, recent genome-wide expression analyses of C. glutamicum cells grown on glucose and on acetate substantiated the idea of this transcriptional regulation of the aceA, aceB, pta, and ack genes (14,21,35). In accordance with induction or repression of these genes by a common regulator protein(s), in front of the genes we observed highly conserved 13-bp motifs (AA/GAACTTTGCAAA) showing a dyad symmetry at one end (underlined) (14).…”

“…Recent genome-wide and comparative expression analyses with C. glutamicum grown on glucose and on acetate revealed that in addition to the pta-ack operon and the aceA and aceB genes, a variety of other genes are obviously under transcriptional control in response to the presence or absence of acetate in the growth medium (14,21,35). These genes (thus belonging to the acetate stimulon) include those for some of the enzymes of the tricarboxylic acid cycle (citrate synthase, aconitase, succinate dehydrogenase subunits A, B, and C/D, and fumarase) and for some of the enzymes involved in sugar metabolism (glucose-specific phosphotransferase system enzyme II, glucose-6-phosphate dehydrogenase, transaldolase, transketolase, pyruvate kinase, and pyruvate dehydrogenase complex E1 subunit).…”

Effect of carbon source availability and growth phase on expression of Corynebacterium glutamicum genes involved in the tricarboxylic acid cycle and glyoxylate bypass

“…The transcriptional regulator GlxR contains a domain with similarity to the cAMP-binding motifs of Crp from E. coli and, in fact, cAMP is essential for binding of GlxR to the aceA/aceB intergenic region in C. glutamicum (Kim et al, 2004;Letek et al, 2006). The regulation of C. glutamicum metabolism in the presence of various carbon sources clearly differs from that of E. coli or B. subtilis (Gerstmeir et al, 2003;Hayashi et al, 2002;Letek et al, 2006;Muffler et al, 2002). No direct evidence has been found of a CCR system in C. glutamicum (Bruckner & Titgemeyer, 2002;Gerstmeir et al, 2003;Han et al, 2007), although a CCR-like phenomenon has been reported from cells grown on medium containing glucose with either glutamate (Gerstmeir et al, 2003;Kramer et al, 1990;Kronemeyer et al, 1995) or ethanol (Arndt & Eikmanns, 2007;Kotrbova-Kozak et al, 2007).…”

Involvement of Regulatory Interactions among Global Regulators GlxR, SugR, and RamA in Expression of ramA in Corynebacterium glutamicum

“…Carbon source-dependent regulation of gene expression in C. glutamicum has been investigated by comparing the expression profile in glucose-grown cells to that in acetate-grown cells (28)(29)(30). These studies reveal that genes for sugar uptake and metabolism, including the phosphotransferase system and glycolysis, are upregulated in the presence of glucose.…”