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DamID-seq: Genome-wide Mapping of Protein-DNA Interactions by High Throughput Sequencing of Adenine-methylated DNA Fragments
Published on: January 27, 2016
GATC flanking sequences regulate Dam activity: evidence for how Dam specificity may influence pap expression
Stacey N Peterson1, Norbert O Reich
1Program in Biomolecular Science and Engineering and the Department of Chemistry and Biochemistry, University of California, Santa Barbara, CA 93106, USA.
Journal of Molecular Biology
|December 3, 2005
Summary
Flanking sequences around GATC sites in Escherichia coli regulate DNA adenine methyltransferase (Dam) activity. This epigenetic mechanism controls Dam processivity, impacting gene expression and bacterial adhesion.
Area of Science:
- Molecular Biology
- Epigenetics
- Microbiology
Background:
- Escherichia coli DNA adenine methyltransferase (Dam) is crucial for DNA replication, repair, and gene regulation.
- Differential methylation of GATC sites in the pap promoter by Dam controls pili gene expression in uropathogenic E.coli.
- Dam typically methylates GATC sites processively, but pap promoter sites exhibit distributive methylation due to flanking sequences.
Purpose of the Study:
- To investigate the role of flanking sequences in regulating Dam methylation processivity at pap GATC sites.
- To determine if altering flanking sequences can enhance Dam processivity.
- To elucidate the mechanism by which flanking sequences influence Dam's catalytic efficiency and gene regulation.
Main Methods:
- Genetic modification of pap GATC site flanking sequences.
- Assays to measure Dam methylation processivity and kinetics.
- Competition experiments to assess the direct impact of flanking sequences on Dam activity.
- Bioinformatic analysis of GATC flanking sequences in E.coli and other bacterial genomes.
Main Results:
- Replacing pap GATC sites with sequences predicted for processive methylation increased Dam processivity.
- Increased processivity resulted from altered methyltransferase kinetics, not binding efficiency.
- Competition experiments confirmed that flanking sequences directly regulate Dam's catalytic efficiency.
- Similar flanking sequences are found in other Dam-regulated promoters, including those with unassigned functions.
Conclusions:
- GATC flanking sequences critically influence Dam activity and processivity at specific genomic sites.
- This epigenetic mechanism, mediated by flanking sequences, is vital for regulating pap and other Dam-controlled genes.
- The findings propose a model where flanking sequences contribute to the diverse biological roles of Dam through epigenetic gene expression control.
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