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Genome-Scale Mapping of Escherichia coli σ54 Reveals Widespread, Conserved Intragenic Binding.

Bonocora RP, Smith C, Lapierre P, Wade JT - PLoS Genet. (2015)

Bottom Line: Strikingly, the majority of σ54 binding sites are located inside genes.We conclude that many intragenic σ54 binding sites are likely to be functional.Consistent with this assertion, we identify three conserved, intragenic σ54 promoters that drive transcription of mRNAs with unusually long 5' UTRs.

View Article: PubMed Central - PubMed

Affiliation: Wadsworth Center, New York State Department of Health, Albany, New York, United States of America.

ABSTRACT
Bacterial RNA polymerases must associate with a σ factor to bind promoter DNA and initiate transcription. There are two families of σ factor: the σ70 family and the σ54 family. Members of the σ54 family are distinct in their ability to bind promoter DNA sequences, in the context of RNA polymerase holoenzyme, in a transcriptionally inactive state. Here, we map the genome-wide association of Escherichia coli σ54, the archetypal member of the σ54 family. Thus, we vastly expand the list of known σ54 binding sites to 135. Moreover, we estimate that there are more than 250 σ54 sites in total. Strikingly, the majority of σ54 binding sites are located inside genes. The location and orientation of intragenic σ54 binding sites is non-random, and many intragenic σ54 binding sites are conserved. We conclude that many intragenic σ54 binding sites are likely to be functional. Consistent with this assertion, we identify three conserved, intragenic σ54 promoters that drive transcription of mRNAs with unusually long 5' UTRs.

No MeSH data available.


Related in: MedlinePlus

σ54 binds conserved sites in Salmonella enterica.Targeted validation of predicted σ54 binding sites in S. enterica. Enrichment (occupancy units) at predicted σ54 binding sites in S. enterica was measured by ChIP-qPCR with anti-σ54 (black bars) or no antibody (white bars). The cognate promoter IDs (Tables 1 & 2) are indicated in parentheses. Error bars represent the standard deviation from three independent biological replicates. Significant differences between wild-type and ΔrpoN values are indicated (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001). Note that the y-axis scale differs for first five regions indicated. Underlined genes indicate σ54 binding sites previously identified by [14].
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pgen.1005552.g009: σ54 binds conserved sites in Salmonella enterica.Targeted validation of predicted σ54 binding sites in S. enterica. Enrichment (occupancy units) at predicted σ54 binding sites in S. enterica was measured by ChIP-qPCR with anti-σ54 (black bars) or no antibody (white bars). The cognate promoter IDs (Tables 1 & 2) are indicated in parentheses. Error bars represent the standard deviation from three independent biological replicates. Significant differences between wild-type and ΔrpoN values are indicated (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001). Note that the y-axis scale differs for first five regions indicated. Underlined genes indicate σ54 binding sites previously identified by [14].

Mentions: The sequence-based phylogenetic analysis described above predicted that many of the σ54 sites we detected in E. coli are functionally conserved in Salmonella enterica. To test this prediction, we used ChIP-qPCR to measure association of σ54 with 14 sites in Salmonella enterica serovar Typhimurium that were predicted on the basis of sequence conservation. Only five of these sites are in the classical “outside sense” orientation (intergenic, oriented towards a gene). As a control, we performed ChIP-qPCR without antibody. In all cases, we detected robust association of σ54, dependent upon the presence of antibody in the ChIP (Fig 9). We conclude that many intragenic σ54 binding sites are functionally conserved in S. enterica. Moreover, these data validate the sequence-based predictions of conservation (Fig 8).


Genome-Scale Mapping of Escherichia coli σ54 Reveals Widespread, Conserved Intragenic Binding.

Bonocora RP, Smith C, Lapierre P, Wade JT - PLoS Genet. (2015)

σ54 binds conserved sites in Salmonella enterica.Targeted validation of predicted σ54 binding sites in S. enterica. Enrichment (occupancy units) at predicted σ54 binding sites in S. enterica was measured by ChIP-qPCR with anti-σ54 (black bars) or no antibody (white bars). The cognate promoter IDs (Tables 1 & 2) are indicated in parentheses. Error bars represent the standard deviation from three independent biological replicates. Significant differences between wild-type and ΔrpoN values are indicated (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001). Note that the y-axis scale differs for first five regions indicated. Underlined genes indicate σ54 binding sites previously identified by [14].
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4591121&req=5

pgen.1005552.g009: σ54 binds conserved sites in Salmonella enterica.Targeted validation of predicted σ54 binding sites in S. enterica. Enrichment (occupancy units) at predicted σ54 binding sites in S. enterica was measured by ChIP-qPCR with anti-σ54 (black bars) or no antibody (white bars). The cognate promoter IDs (Tables 1 & 2) are indicated in parentheses. Error bars represent the standard deviation from three independent biological replicates. Significant differences between wild-type and ΔrpoN values are indicated (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001). Note that the y-axis scale differs for first five regions indicated. Underlined genes indicate σ54 binding sites previously identified by [14].
Mentions: The sequence-based phylogenetic analysis described above predicted that many of the σ54 sites we detected in E. coli are functionally conserved in Salmonella enterica. To test this prediction, we used ChIP-qPCR to measure association of σ54 with 14 sites in Salmonella enterica serovar Typhimurium that were predicted on the basis of sequence conservation. Only five of these sites are in the classical “outside sense” orientation (intergenic, oriented towards a gene). As a control, we performed ChIP-qPCR without antibody. In all cases, we detected robust association of σ54, dependent upon the presence of antibody in the ChIP (Fig 9). We conclude that many intragenic σ54 binding sites are functionally conserved in S. enterica. Moreover, these data validate the sequence-based predictions of conservation (Fig 8).

Bottom Line: Strikingly, the majority of σ54 binding sites are located inside genes.We conclude that many intragenic σ54 binding sites are likely to be functional.Consistent with this assertion, we identify three conserved, intragenic σ54 promoters that drive transcription of mRNAs with unusually long 5' UTRs.

View Article: PubMed Central - PubMed

Affiliation: Wadsworth Center, New York State Department of Health, Albany, New York, United States of America.

ABSTRACT
Bacterial RNA polymerases must associate with a σ factor to bind promoter DNA and initiate transcription. There are two families of σ factor: the σ70 family and the σ54 family. Members of the σ54 family are distinct in their ability to bind promoter DNA sequences, in the context of RNA polymerase holoenzyme, in a transcriptionally inactive state. Here, we map the genome-wide association of Escherichia coli σ54, the archetypal member of the σ54 family. Thus, we vastly expand the list of known σ54 binding sites to 135. Moreover, we estimate that there are more than 250 σ54 sites in total. Strikingly, the majority of σ54 binding sites are located inside genes. The location and orientation of intragenic σ54 binding sites is non-random, and many intragenic σ54 binding sites are conserved. We conclude that many intragenic σ54 binding sites are likely to be functional. Consistent with this assertion, we identify three conserved, intragenic σ54 promoters that drive transcription of mRNAs with unusually long 5' UTRs.

No MeSH data available.


Related in: MedlinePlus