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Dynamic distribution of seqa protein across the chromosome of escherichia coli K-12.

Sánchez-Romero MA, Busby SJ, Dyer NP, Ott S, Millard AD, Grainger DC - MBio (2010)

Bottom Line: Less SeqA is found in highly transcribed regions, as well as in the ter macrodomain.Using synchronized cultures, we show that SeqA distribution differs with the cell cycle.SeqA remains bound to some targets after replication has ceased, and these targets locate to genes encoding factors involved in nucleotide metabolism, chromosome replication, and methyl transfer.

View Article: PubMed Central - PubMed

Affiliation: School of Biosciences, the University of Birmingham, Edgbaston, Birmingham, United Kingdom.

ABSTRACT
The bacterial SeqA protein binds to hemi-methylated GATC sequences that arise in newly synthesized DNA upon passage of the replication machinery. In Escherichia coli K-12, the single replication origin oriC is a well-characterized target for SeqA, which binds to multiple hemi-methylated GATC sequences immediately after replication has initiated. This sequesters oriC, thereby preventing reinitiation of replication. However, the genome-wide DNA binding properties of SeqA are unknown, and hence, here, we describe a study of the binding of SeqA across the entire Escherichia coli K-12 chromosome, using chromatin immunoprecipitation in combination with DNA microarrays. Our data show that SeqA binding correlates with the frequency and spacing of GATC sequences across the entire genome. Less SeqA is found in highly transcribed regions, as well as in the ter macrodomain. Using synchronized cultures, we show that SeqA distribution differs with the cell cycle. SeqA remains bound to some targets after replication has ceased, and these targets locate to genes encoding factors involved in nucleotide metabolism, chromosome replication, and methyl transfer.

No MeSH data available.


Related in: MedlinePlus

Genome-wide view of SeqA binding at different points in the cell cycle. The figure shows ChIP-chip data for SeqA binding plotted against features of the E. coli genome in the form of a genome atlas. (A) SeqA binding signal generated using unsynchronized cultures of E. coli (time point A); (B) SeqA binding after chromosome replication had been blocked for 1 h (time point B); (C) SeqA binding in cultures where chromosome replication had been reinitiated in synchronicity for a period of 6 min.
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f2: Genome-wide view of SeqA binding at different points in the cell cycle. The figure shows ChIP-chip data for SeqA binding plotted against features of the E. coli genome in the form of a genome atlas. (A) SeqA binding signal generated using unsynchronized cultures of E. coli (time point A); (B) SeqA binding after chromosome replication had been blocked for 1 h (time point B); (C) SeqA binding in cultures where chromosome replication had been reinitiated in synchronicity for a period of 6 min.

Mentions: Figure 2 shows an overview of the SeqA binding profile at time points A, B, and C, derived from ChIP-chip data and plotted against the basic features of the E. coli chromosome and the local density of GATC motifs. In unsynchronized cells, the largest SeqA binding signal corresponds exactly with the location of oriC (Fig. 2A). A clear signal for SeqA binding is observed at the nearby dnaA locus, with binding being spread across the entire gene (see Fig. S1 in the supplemental material). Further SeqA binding signals, comparable in intensity to the signal seen at dnaA, are scattered throughout the genome, and these correspond well to locations where the frequency of GATC sites is higher (some examples are shown in Fig. S2 in the supplemental material). One hundred thirty-seven genes have a SeqA binding signal >4-fold above background levels (see Table S7 in the supplemental material). Of these genes, 24 are directly involved in nucleotide metabolism, DNA repair/replication, or methyl group transfer. Interestingly, SeqA binding across an ~1.3-Mbp segment that includes the ter region is greatly reduced. This region has a relatively low GATC content, and this most likely accounts for the reduced SeqA binding signal. Additionally, in unsynchronized cultures, weak SeqA binding signals in the ter region may be difficult to distinguish from experimental noise.


Dynamic distribution of seqa protein across the chromosome of escherichia coli K-12.

Sánchez-Romero MA, Busby SJ, Dyer NP, Ott S, Millard AD, Grainger DC - MBio (2010)

Genome-wide view of SeqA binding at different points in the cell cycle. The figure shows ChIP-chip data for SeqA binding plotted against features of the E. coli genome in the form of a genome atlas. (A) SeqA binding signal generated using unsynchronized cultures of E. coli (time point A); (B) SeqA binding after chromosome replication had been blocked for 1 h (time point B); (C) SeqA binding in cultures where chromosome replication had been reinitiated in synchronicity for a period of 6 min.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2912659&req=5

f2: Genome-wide view of SeqA binding at different points in the cell cycle. The figure shows ChIP-chip data for SeqA binding plotted against features of the E. coli genome in the form of a genome atlas. (A) SeqA binding signal generated using unsynchronized cultures of E. coli (time point A); (B) SeqA binding after chromosome replication had been blocked for 1 h (time point B); (C) SeqA binding in cultures where chromosome replication had been reinitiated in synchronicity for a period of 6 min.
Mentions: Figure 2 shows an overview of the SeqA binding profile at time points A, B, and C, derived from ChIP-chip data and plotted against the basic features of the E. coli chromosome and the local density of GATC motifs. In unsynchronized cells, the largest SeqA binding signal corresponds exactly with the location of oriC (Fig. 2A). A clear signal for SeqA binding is observed at the nearby dnaA locus, with binding being spread across the entire gene (see Fig. S1 in the supplemental material). Further SeqA binding signals, comparable in intensity to the signal seen at dnaA, are scattered throughout the genome, and these correspond well to locations where the frequency of GATC sites is higher (some examples are shown in Fig. S2 in the supplemental material). One hundred thirty-seven genes have a SeqA binding signal >4-fold above background levels (see Table S7 in the supplemental material). Of these genes, 24 are directly involved in nucleotide metabolism, DNA repair/replication, or methyl group transfer. Interestingly, SeqA binding across an ~1.3-Mbp segment that includes the ter region is greatly reduced. This region has a relatively low GATC content, and this most likely accounts for the reduced SeqA binding signal. Additionally, in unsynchronized cultures, weak SeqA binding signals in the ter region may be difficult to distinguish from experimental noise.

Bottom Line: Less SeqA is found in highly transcribed regions, as well as in the ter macrodomain.Using synchronized cultures, we show that SeqA distribution differs with the cell cycle.SeqA remains bound to some targets after replication has ceased, and these targets locate to genes encoding factors involved in nucleotide metabolism, chromosome replication, and methyl transfer.

View Article: PubMed Central - PubMed

Affiliation: School of Biosciences, the University of Birmingham, Edgbaston, Birmingham, United Kingdom.

ABSTRACT
The bacterial SeqA protein binds to hemi-methylated GATC sequences that arise in newly synthesized DNA upon passage of the replication machinery. In Escherichia coli K-12, the single replication origin oriC is a well-characterized target for SeqA, which binds to multiple hemi-methylated GATC sequences immediately after replication has initiated. This sequesters oriC, thereby preventing reinitiation of replication. However, the genome-wide DNA binding properties of SeqA are unknown, and hence, here, we describe a study of the binding of SeqA across the entire Escherichia coli K-12 chromosome, using chromatin immunoprecipitation in combination with DNA microarrays. Our data show that SeqA binding correlates with the frequency and spacing of GATC sequences across the entire genome. Less SeqA is found in highly transcribed regions, as well as in the ter macrodomain. Using synchronized cultures, we show that SeqA distribution differs with the cell cycle. SeqA remains bound to some targets after replication has ceased, and these targets locate to genes encoding factors involved in nucleotide metabolism, chromosome replication, and methyl transfer.

No MeSH data available.


Related in: MedlinePlus