The eukaryotic cell exhibits compartmentalization of functions to various membrane-bound organelles and to specific domains within each membrane. The spatial distribution of the membrane chemoreceptors and associated cytoplasmic chemotaxis proteins in Escherichia coli were examined as a prototypic functional aggregate in bacterial cells. Bacterial chemotaxis involves a phospho-relay system brought about by ligand association with a membrane receptor, culminating in a switch in the direction of flagellar rotation. The transduction of the chemotaxis signal is initiated by a chemoreceptor-CheW-CheA ternary complex at the inner membrane. These ternary complexes aggregate predominantly at the cell poles. Polar localization of the cytoplasmic CheA and CheW proteins is dependent on membrane-bound chemoreceptor. Chemoreceptors are not confined to the cell poles in strains lacking both CheA and CheW. The chemoreceptor-CheW binary complex is polarly localized in the absence of CheA, whereas the chemoreceptor-CheA binary complex is not confined to the cell poles in strains lacking CheW. The subcellular localization of the chemotaxis proteins may reflect a general mechanism by which the bacterial cell sequesters different regions of the cell for specialized functions.

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The CcrM DNA methyltransferase of the ␣-proteobacteria catalyzes the methylation of the adenine in the sequence GAnTC. Like Dam in the enterobacteria, CcrM plays a regulatory role in Caulobacter crescentus and Rhizobium meliloti. CcrM is essential for viability in both of these organisms, and we show here that it is also essential in Brucella abortus. Further, increased copy number of the ccrM gene results in striking changes in B. abortus morphology, DNA replication, and growth in murine macrophages. We generated strains that carry ccrM either on a low-copy-number plasmid (strain GR131) or on a moderate-copy-number plasmid (strain GR132). Strain GR131 has wild-type morphology and chromosome number, as assessed by flow cytometry. In contrast, strain GR132 has abnormal branched morphology, suggesting aberrant cell division, and increased chromosome number. Although these strains exhibit different morphologies and DNA content, the replication of both strains in macrophages is attenuated. These data imply that the reduction in survival in host cells is not due solely to a cell division defect but is due to additional functions of CcrM. Because CcrM is essential in B. abortus and increased ccrM copy number attenuates survival in host cells, we propose that CcrM is an appropriate target for new antibiotics.Changes in DNA methylation patterns signal changes in cellular physiology in both prokaryotes and eukaryotes. In bacteria, DNA methyltransferases not only participate in restriction-modification systems (5) but also play regulatory roles in the cell. For example, methylation of the origin of replication by the Dam methyltransferase governs the timing of the initiation of DNA replication in Escherichia coli (6,29). Dam methylation also contributes to strand discrimination in methyl-directed DNA mismatch repair (19) and plays a role in pathogenesis (10, 12). For instance, changes in Dam methylation patterns control pyelonephritis-associated pilus (pap) transcription in uropathogenic E. coli by altering the binding of Lrp (leucine-responsive protein) to the pap promoter (21, 38). In addition, Dam methylation either directly or indirectly regulates the transcription of a number of genes in Salmonella enterica serovar Typhimurium that are induced following infection of the host (12), suggesting that DNA methylation plays a role in the virulence of this organism.The Dam methyltransferases of E. coli and other ␥-proteobacteria have a counterpart in ␣-proteobacteria, a DNA adenine methyltransferase called CcrM (for cell cycle-regulated methyltransferase). This enzyme was originally described in Caulobacter crescentus, a bacterium that is easily synchronized and therefore amenable to cell cycle studies (42). Both Dam and CcrM catalyze the transfer of the methyl group from S-adenosylmethionine to the adenine of their target DNA sequences. Both enzymes apparently lack cognate restriction enzymes and instead regulate cell cycle events. However, unlike Dam, CcrM is required for cell viability, and its activity is tightly regula...

Only one of the two chromosomes in the asymmetric Caulobacter predivisional cell initiates replication in the progeny cells. Transcription from a strong promoter within the origin occurs uniquely from the replication-competent chromosome at the stalked pole of the predivisional cell. This regulated promoter has an unusual sequence organization, and transcription from this promoter is essential for regulated {cell type-specific) replication. Our analysis defines a new class of bacterial origins and suggests a coupling between transcription and replication that is consistent with the phylogenetic relationship of Caulobacter to the ancestral mitochondrion.

Transcription of flagellar genes in Caulobacter crecentus is programmed to occur during the predivisional stage of the cell cycle. The mechanism of activation of Class II flagellar genes, the highest identified genes in the Caulobacter flagellar hierarchy, is unknown. As a step toward understanding this process, we have defined cis-acting sequences necessary for expression of a Class II flagellar operon, fliLM. Deletion analysis indicated that a 55 bp DNA fragment was sufficient for normal, temporally regulated promoter activity. Transcription from this promoter-containing fragment was severely reduced when chromosomal DNA replication was inhibited. Extensive mutational analysis of the promoter region from -42 to -5 identified functionally important nucleotides at -36 and -35, between -29 and -22, and at -12, which correlates well with sequences conserved between fliLM and the analogous regions of two other Class II flagellar operons. The promoter sequence does not resemble that recognized by any known bacterial sigma factor. Models for regulation of Caulobacter early flagellar promoters are discussed in which RNA polymerase containing a novel sigma subunit interacts with an activation factor bound to the central region of the promoter.