Novel Roles of cAMP Receptor Protein (CRP) in Regulation of Transport and Metabolism of Carbon Sources

Abstract

CRP (cAMP receptor protein), the global regulator of genes for carbon source utilization in the absence of glucose, is the best-studied prokaryotic transcription factor. A total of 195 target promoters on the Escherichia coli genome have been proposed to be under the control of cAMP-bound CRP. Using the newly developed Genomic SELEX screening system of transcription factor-binding sequences, however, we have identified a total of at least 254 CRP-binding sites. Based on their location on the E. coli genome, we predict a total of at least 183 novel regulation target operons, altogether with the 195 hitherto known targets, reaching to the minimum of 378 promoters as the regulation targets of cAMP-CRP. All the promoters selected from the newly identified targets and examined by using the lacZ reporter assay were found to be under the control of CRP, indicating that the Genomic SELEX screening allowed to identify the CRP targets with high accuracy. Based on the functions of novel target genes, we conclude that CRP plays a key regulatory role in the whole processes from the selective transport of carbon sources, the glycolysis-gluconeogenesis switching to the metabolisms downstream of glycolysis, including tricarboxylic acid (TCA) cycle, pyruvate dehydrogenase (PDH) pathway and aerobic respiration. One unique regulation mode is that a single and the same CRP molecule bound within intergenic regions often regulates both of divergently transcribed operons.

Circuitry Linking the Catabolite Repression and Csr Global Regulatory Systems of Escherichia coli

“…Integration host factor IHF is the only factor known to differentially activate csrB transcription without affecting csrC (3,41). However, the ihfA and ihfB genes, which encode the IHF subunits, were not among the E. coli genes found to contain cAMP-CRP binding sites by genomic SELEX analyses (47). Thus, IHF seems unlikely to mediate the effects of cAMP-CRP on csrB.…”

“…For example, (i) crp and cya mRNAs copurified with the CsrA protein (25), (ii) potential cAMP-CRP binding sites were identified upstream of the csrB gene by bioinformatics analysis (47), and (iii) a number of genes and processes have been reported to be regulated by both CsrA and cAMP-CRP (14,18,21,66) (Fig. 10).…”

“…The cAMP-CRP complex regulates transcription in response to the availability of a preferred carbon source, e.g., glucose (45)(46)(47). Under conditions of carbon limitation, the PTS proteins, including the glucose-specific protein EIIA Glc , are predominantly phosphorylated.…”

“…The cAMP-CRP complex mediates hierarchical utilization of nonpreferred carbon sources, referred to as carbon catabolite repression (CCR), by activating the expression of genes required for the transport and utilization of alternative carbon sources (49). cAMP-CRP also influences the expression of genes not directly involved in carbon metabolism such as those encoding ribosomal proteins, tRNAs, amino acid biosynthesis enzymes, heat shock proteins, sRNAs, and perhaps as many as 70 transcription factors (45)(46)(47)(50)(51)(52)(53)(54)(55). cAMP levels and cAMP-CRP regulatory functions have been suggested to respond to both the carbon status and the nitrogen status of the cell, leading to reorganization of the proteome (56).…”

“…Bioinformatics analysis for potential cAMP-CRP binding sites in the E. coli genome identified the coding region of syd, the gene immediately upstream from csrB, as a possible target (47). In addition, an online tool (Virtual Footprint [www.prodoric.de/vfp]) for predicting the binding sequences of regulatory proteins identified potential CRP binding sites in csrB, csrC, csrA, csrD, and uvrY.…”