The functions of DNA methylation by CcrM in Caulobacter crescentus: a global approach

Abstract

DNA methylation is involved in a diversity of processes in bacteria, including maintenance of genome integrity and regulation of gene expression. Here, using Caulobacter crescentus as a model, we exploit genome-wide experimental methods to uncover the functions of CcrM, a DNA methyltransferase conserved in most Alphaproteobacteria. Using single molecule sequencing, we provide evidence that most CcrM target motifs (GANTC) switch from a fully methylated to a hemi-methylated state when they are replicated, and back to a fully methylated state at the onset of cell division. We show that DNA methylation by CcrM is not required for the control of the initiation of chromosome replication or for DNA mismatch repair. By contrast, our transcriptome analysis shows that >10% of the genes are misexpressed in cells lacking or constitutively over-expressing CcrM. Strikingly, GANTC methylation is needed for the efficient transcription of dozens of genes that are essential for cell cycle progression, in particular for DNA metabolism and cell division. Many of them are controlled by promoters methylated by CcrM and co-regulated by other global cell cycle regulators, demonstrating an extensive cross talk between DNA methylation and the complex regulatory network that controls the cell cycle of C. crescentus and, presumably, of many other Alphaproteobacteria.

Short-Stalked Prosthecomicrobium hirschii Cells Have a Caulobacter-Like Cell Cycle

“…4B and D). This observation is consistent with the described role of CcrMmediated methylation in mediating cell cycle progression, DNA metabolism, and cell division in C. crescentus (57,58).…”

“…In C. crescentus, the methylation of DNA by CcrM plays an important role in the regulation of cell cycle progression (57)(58)(59), and methylated sites are subject to transcriptional regulation by GcrA (14). Methylation by CcrM is conserved in other Alphaproteobacteria, including B. abortus (18), A. tumefaciens (9), and S. meliloti (60).…”

Molecular Characterization of a Novel Temperate Sinorhizobium Bacteriophage, ФLM21, Encoding DNA Methyltransferase with CcrM-Like Specificity

“…The cell cycle-regulating methyltransferase CcrM can modify the adenine residue in the GANTC sequence and is widespread among members of Alphaproteobacteria. It has been demonstrated that the CcrM protein is essential for the viability of S. meliloti, A. tumefaciens, Brucella abortus, and Caulobacter crescentus in rich medium cultures (49,50) and that it participates in the regulation of the bacterial cell cycle (51,52).…”

“…None of the GANTC sequences were found within the predicted "lysogeny" control region (see above). Analysis of the frequency and distribution of GANTC motifs in the available genomes of Alphaproteobacteria showed a distribution bias of GANTC motifs between the protein-coding and intergenic sequences, being on average Ͼ1.5-fold overrepresented in the intergenic regions and slightly underrepresented in the coding regions (52). It was also shown that the expression levels of 546 genes of C. crescentus were significantly altered in the strain with constitutive overexpression of CcrM compared to the wild-type strain, with 214 of these genes being affected Ͼ2-fold (52).…”

Fifty Years after the Replicon Hypothesis: Cell-Specific Master Regulators as New Players in Chromosome Replication Control

“…It is worth mentioning that, during the swarmer-cell-to-stalked-cell transition, DnaA accumulation (accompanied by CtrA degradation) leads to activation of gcrA tran- scription and subsequent CtrA expression induced by the GcrA cell cycle regulator (98). Interestingly, though there are also CcrM methylation sites within the C. crescentus origin, it seems that methylation of the origin does not play a role in regulating the initiation of chromosomal replication as it does with Dam methylase (99,100).…”