scholarly journals A cell cycle-regulated adenine DNA methyltransferase from Caulobacter crescentus processively methylates GANTC sites on hemimethylated DNA

1998 ◽  
Vol 95 (6) ◽  
pp. 2874-2879 ◽  
Author(s):  
A. J. Berdis ◽  
I. Lee ◽  
J. K. Coward ◽  
C. Stephens ◽  
R. Wright ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Methyltransferase ◽  
Caulobacter Crescentus ◽  
A Cell

scholarly journals Beta class amino methyltransferases from bacteria to humans: evolution and structural consequences

2020 ◽  
Vol 48 (18) ◽  
pp. 10034-10044 ◽  
Author(s):  
Clayton B Woodcock ◽  
John R Horton ◽  
Xing Zhang ◽  
Robert M Blumenthal ◽  
Xiaodong Cheng
Keyword(s):  
Escherichia Coli ◽  
Cell Cycle ◽  
Dna Methyltransferase ◽  
Caulobacter Crescentus ◽  
Substrate Recognition ◽  
Its Sequence ◽  
Amino Groups ◽  
Dna And Rna ◽  
Acid Type ◽  
Target Molecules

Abstract S-adenosyl-l-methionine dependent methyltransferases catalyze methyl transfers onto a wide variety of target molecules, including DNA and RNA. We discuss a family of methyltransferases, those that act on the amino groups of adenine or cytosine in DNA, have conserved motifs in a particular order in their amino acid sequence, and are referred to as class beta MTases. Members of this class include M.EcoGII and M.EcoP15I from Escherichia coli, Caulobacter crescentus cell cycle–regulated DNA methyltransferase (CcrM), the MTA1-MTA9 complex from the ciliate Oxytricha, and the mammalian MettL3-MettL14 complex. These methyltransferases all generate N6-methyladenine in DNA, with some members having activity on single-stranded DNA as well as RNA. The beta class of methyltransferases has a unique multimeric feature, forming either homo- or hetero-dimers, allowing the enzyme to use division of labor between two subunits in terms of substrate recognition and methylation. We suggest that M.EcoGII may represent an ancestral form of these enzymes, as its activity is independent of the nucleic acid type (RNA or DNA), its strandedness (single or double), and its sequence (aside from the target adenine).

scholarly journals Cell cycle regulated DNA methyltransferase: fluorescent tracking of a DNA strand-separation mechanism and identification of the responsible protein motif

2020 ◽  
Vol 48 (20) ◽  
pp. 11589-11601
Author(s):  
Olivia Konttinen ◽  
Jason Carmody ◽  
Sarath Pathuri ◽  
Kyle Anderson ◽  
Xiaofeng Zhou ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Methyltransferase ◽  
Caulobacter Crescentus ◽  
Dna Recognition ◽  
Dna Methyltransferases ◽  
Terminal Segment ◽  
Recognition Site ◽  
Separation Mechanism ◽  
Human Pathogens ◽  
Strand Separation

Abstract DNA adenine methylation by Caulobacter crescentus Cell Cycle Regulated Methyltransferase (CcrM) is an important epigenetic regulator of gene expression. The recent CcrM-DNA cocrystal structure shows the CcrM dimer disrupts four of the five base pairs of the (5′-GANTC-3′) recognition site. We developed a fluorescence-based assay by which Pyrrolo-dC tracks the strand separation event. Placement of Pyrrolo-dC within the DNA recognition site results in a fluorescence increase when CcrM binds. Non-cognate sequences display little to no fluorescence changes, showing that strand separation is a specificity determinant. Conserved residues in the C-terminal segment interact with the phospho-sugar backbone of the non-target strand. Replacement of these residues with alanine results in decreased methylation activity and changes in strand separation. The DNA recognition mechanism appears to occur with the Type II M.HinfI DNA methyltransferase and an ortholog of CcrM, BabI, but not with DNA methyltransferases that lack the conserved C-terminal segment. The C-terminal segment is found broadly in N4/N6-adenine DNA methyltransferases, some of which are human pathogens, across three Proteobacteria classes, three other phyla and in Thermoplasma acidophilum, an Archaea. This Pyrrolo-dC strand separation assay should be useful for the study of other enzymes which likely rely on a strand separation mechanism.

scholarly journals Genomic Adaptations to the Loss of a Conserved Bacterial DNA Methyltransferase

mBio ◽  
2015 ◽  
Vol 6 (4) ◽  
Author(s):  
Diego Gonzalez ◽  
Justine Collier
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Experimental Evolution ◽  
Dna Methyltransferase ◽  
Metabolic Regulation ◽  
Caulobacter Crescentus ◽  
Point Mutations ◽  
Rich Medium ◽  
Content Type ◽  
Evolution Approach

ABSTRACTCcrM is an orphan DNA methyltransferase nearly universally conserved in a vast group ofAlphaproteobacteria.InCaulobacter crescentus, it controls the expression of key genes involved in the regulation of the cell cycle and cell division. Here, we demonstrate, using an experimental evolution approach, thatC. crescentuscan significantly compensate, through easily accessible genetic changes like point mutations, the severe loss in fitness due to the absence of CcrM, quickly improving its growth rate and cell morphology in rich medium. By analyzing the compensatory mutations genome-wide in 12 clones sampled from independent ΔccrMpopulations evolved for ~300 generations, we demonstrated that each of the twelve clones carried at least one mutation that potentially stimulatedftsZexpression, suggesting that the low intracellular levels of FtsZ are the major burden of ΔccrMmutants. In addition, we demonstrate that the phosphoenolpyruvate-carbohydrate phosphotransfer system (PTS) actually modulatesftsZandmipZtranscription, uncovering a previously unsuspected link between metabolic regulation and cell division inAlphaproteobacteria. We present evidence that point mutations found in genes encoding proteins of the PTS provide the strongest fitness advantage to ΔccrMcells cultivated in rich medium despite being disadvantageous in minimal medium. This environmental sign epistasis might prevent such mutations from getting fixed under changing natural conditions, adding a plausible explanation for the broad conservation of CcrM.IMPORTANCEIn bacteria, DNA methylation has a variety of functions, including the control of DNA replication and/or gene expression. The cell cycle-regulated DNA methyltransferase CcrM modulates the transcription of many genes and is critical for fitness inCaulobacter crescentus. Here, we used an original experimental evolution approach to determine which of its many targets make CcrM so important physiologically. We show that populations lacking CcrM evolve quickly, accumulating an excess of mutations affecting, directly or indirectly, the expression of theftsZcell division gene. This finding suggests that the most critical function of CcrM inC. crescentusis to promote cell division by enhancing FtsZ intracellular levels. During this work, we also discovered an unexpected link between metabolic regulation and cell division that might extend to otherAlphaproteobacteria.

scholarly journals Conserved Promoter Motif Is Required for Cell Cycle Timing of dnaX Transcription inCaulobacter

2001 ◽  
Vol 183 (16) ◽  
pp. 4860-4865 ◽  
Author(s):  
Kenneth C. Keiler ◽  
Lucy Shapiro
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Caulobacter Crescentus ◽  
Transcriptional Networks ◽  
Cellular Processes ◽  
Genes Encoding ◽  
Constitutive Gene Expression ◽  
A Cell ◽  
Replication Factors

ABSTRACT Cells use highly regulated transcriptional networks to control temporally regulated events. In the bacterium Caulobacter crescentus, many cellular processes are temporally regulated with respect to the cell cycle, and the genes required for these processes are expressed immediately before the products are needed. Genes encoding factors required for DNA replication, includingdnaX, dnaA, dnaN,gyrB, and dnaK, are induced at the G1/S-phase transition. By analyzing mutations in thednaX promoter, we identified a motif between the −10 and −35 regions that is required for proper timing of gene expression. This motif, named RRF (for repression of replication factors), is conserved in the promoters of other coordinately induced replication factors. Because mutations in the RRF motif result in constitutive gene expression throughout the cell cycle, this sequence is likely to be the binding site for a cell cycle-regulated transcriptional repressor. Consistent with this hypothesis, Caulobacter extracts contain an activity that binds specifically to the RRF in vitro.

scholarly journals The CtrA Response Regulator Mediates Temporal Control of Gene Expression during the Caulobacter Cell Cycle

1999 ◽  
Vol 181 (8) ◽  
pp. 2430-2439 ◽  
Author(s):  
Ann Reisenauer ◽  
Kim Quon ◽  
Lucy Shapiro
Keyword(s):  
Cell Cycle ◽  
Inverted Repeat ◽  
Transcription Initiation ◽  
Dna Methyltransferase ◽  
Caulobacter Crescentus ◽  
Response Regulator ◽  
Temporal Control ◽  
Temporal Expression ◽  
Recognition Sequence ◽  
Control Of Gene Expression

ABSTRACT In its role as a global response regulator, CtrA controls the transcription of a diverse group of genes at different times in theCaulobacter crescentus cell cycle. To understand the differential regulation of CtrA-controlled genes, we compared the expression of two of these genes, the fliQ flagellar gene and the ccrM DNA methyltransferase gene. Despite their similar promoter architecture, these genes are transcribed at different times in the cell cycle. PfliQ is activated earlier than PccrM. Phosphorylated CtrA (CtrA∼P) bound to the CtrA recognition sequence in both promoters but had a 10- to 20-fold greater affinity for PfliQ. This difference in affinity correlates with temporal changes in the cellular levels of CtrA. Disrupting a unique inverted repeat element in PccrMsignificantly reduced promoter activity but not the timing of transcription initiation, suggesting that the inverted repeat does not play a major role in the temporal control of ccrMexpression. Our data indicate that differences in the affinity of CtrA∼P for PfliQ and PccrM regulate, in part, the temporal expression of these genes. However, the timing offliQ transcription but not of ccrMtranscription was altered in cells expressing a stable CtrA derivative, indicating that changes in CtrA∼P levels alone cannot govern the cell cycle transcription of these genes. We propose that changes in the cellular concentration of CtrA∼P and its interaction with accessory proteins influence the temporal expression offliQ, ccrM, and other key cell cycle genes and ultimately the regulation of the cell cycle.

scholarly journals A cell cycle-regulated bacterial DNA methyltransferase is essential for viability.

1996 ◽  
Vol 93 (3) ◽  
pp. 1210-1214 ◽  
Author(s):  
C. Stephens ◽  
A. Reisenauer ◽  
R. Wright ◽  
L. Shapiro
Keyword(s):  
Cell Cycle ◽  
Dna Methyltransferase ◽  
Bacterial Dna ◽  
A Cell

scholarly journals The cell cycle-regulated DNA adenine methyltransferase CcrM opens a bubble at its DNA recognition site

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
John R. Horton ◽  
Clayton B. Woodcock ◽  
Sifa B. Opot ◽  
Norbert O. Reich ◽  
Xing Zhang ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Methyltransferase ◽  
Caulobacter Crescentus ◽  
Recognition Site ◽  
Van Der Waals Interactions ◽  
Target Sequence ◽  
Recognition Sequence ◽  
Methyltransferase Domain ◽  
Target Strand ◽  
Dna Strands

Abstract The Caulobacter crescentus cell cycle-regulated DNA methyltransferase (CcrM) methylates the adenine of hemimethylated GANTC after replication. Here we present the structure of CcrM in complex with double-stranded DNA containing the recognition sequence. CcrM contains an N-terminal methyltransferase domain and a C-terminal nonspecific DNA-binding domain. CcrM is a dimer, with each monomer contacting primarily one DNA strand: the methyltransferase domain of one molecule binds the target strand, recognizes the target sequence, and catalyzes methyl transfer, while the C-terminal domain of the second molecule binds the non-target strand. The DNA contacts at the 5-base pair recognition site results in dramatic DNA distortions including bending, unwinding and base flipping. The two DNA strands are pulled apart, creating a bubble comprising four recognized base pairs. The five bases of the target strand are recognized meticulously by stacking contacts, van der Waals interactions and specific Watson–Crick polar hydrogen bonds to ensure high enzymatic specificity.

scholarly journals Functional dichotomy and distinct nanoscale assemblies of a cell cycle-controlled bipolar zinc-finger regulator

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Johann Mignolet ◽  
Seamus Holden ◽  
Matthieu Bergé ◽  
Gaël Panis ◽  
Ezgi Eroglu ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Zinc Finger ◽  
Cell Cycle Progression ◽  
Zinc Finger Protein ◽  
Caulobacter Crescentus ◽  
Super Resolution ◽  
Swarming Motility ◽  
Cycle Progression ◽  
Pilus Biogenesis ◽  
A Cell

Protein polarization underlies differentiation in metazoans and in bacteria. How symmetric polarization can instate functional asymmetry remains elusive. Here, we show by super-resolution photo-activated localization microscopy and edgetic mutations that the bitopic zinc-finger protein ZitP implements specialized developmental functions – pilus biogenesis and multifactorial swarming motility – while shaping distinct nanoscale (bi)polar architectures in the asymmetric model bacterium Caulobacter crescentus. Polar assemblage and accumulation of ZitP and its effector protein CpaM are orchestrated in time and space by conserved components of the cell cycle circuitry that coordinate polar morphogenesis with cell cycle progression, and also act on the master cell cycle regulator CtrA. Thus, this novel class of potentially widespread multifunctional polarity regulators is deeply embedded in the cell cycle circuitry.

scholarly journals Spatiotemporal control of PopZ localization through cell cycle–coupled multimerization

2013 ◽  
Vol 201 (6) ◽  
pp. 827-841 ◽  
Author(s):  
Géraldine Laloux ◽  
Christine Jacobs-Wagner
Keyword(s):  
Cell Cycle ◽  
Self Assembly ◽  
Cell Cycle Progression ◽  
Caulobacter Crescentus ◽  
Protein Localization ◽  
Order Structure ◽  
Asymmetric Distribution ◽  
Physiological Processes ◽  
Anchoring Protein ◽  
A Cell

Bacterial cell poles constitute defined subcellular domains where numerous proteins localize, often at specific times, to affect various physiological processes. How pole recognition occurs and what governs the timing of protein localization are often unknown. In this paper, we investigate the mechanisms governing the localization of PopZ, a chromosome-anchoring protein whose unipolar to bipolar localization pattern is critical for cell cycle progression in Caulobacter crescentus. We provide evidence that polar localization of PopZ relied on its self-assembly into a higher-order structure (matrix) and that the unipolar to bipolar transition was coupled to the asymmetric distribution of ParA during the translocation of the origin-proximal ParB–parS partition complex. Collectively, our data suggest a model in which a local increase of ParA concentration promotes the assembly of a PopZ matrix precisely when and where this matrix is needed. Such coupling of protein assembly with a cell cycle–associated molecular asymmetry may represent a principle of cellular organization for controlling protein localization in both time and space.

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