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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

Conservation analysis of σ54 binding sites.Heat-maps depicting the match to the σ54 consensus binding site for each (A) canonical and (B) non-canonical σ54 binding site across a range of bacterial species. Genera are listed across the top, binding site ID numbers, and fold above threshold (FAT) scores in parentheses are listed to the left of the heat-map. For σ54 binding sites in panel A, the gene immediately downstream of each binding site is indicated to the right of the heat-map. (C) Comparison of the level of σ54 binding, as indicated by FAT score (Tables 1 & 2), versus Motif Score (S5 Table) for E. coli only. Different classes of binding site are indicated by color.
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pgen.1005552.g008: Conservation analysis of σ54 binding sites.Heat-maps depicting the match to the σ54 consensus binding site for each (A) canonical and (B) non-canonical σ54 binding site across a range of bacterial species. Genera are listed across the top, binding site ID numbers, and fold above threshold (FAT) scores in parentheses are listed to the left of the heat-map. For σ54 binding sites in panel A, the gene immediately downstream of each binding site is indicated to the right of the heat-map. (C) Comparison of the level of σ54 binding, as indicated by FAT score (Tables 1 & 2), versus Motif Score (S5 Table) for E. coli only. Different classes of binding site are indicated by color.

Mentions: We speculated that other intragenic σ54 binding sites represent genuine promoters. We therefore compared each site across a range of bacterial species, mostly from the family Enterobacteriaceae. σ54 is well conserved across these species (e.g. 62% identical, 79% similar amino acid sequence between E. coli and V. cholerae), suggesting that it binds with similar DNA sequence specificity. Furthermore, ChIP-seq of σ54 in V. cholerae identified a very similar motif to the E. coli σ54 motif identified here (Fig 1B) [19], despite the two species having diverged >600 million years ago [40]. Therefore, we used a position weight matrix derived from our MEME analysis (Fig 1B) to score sequences, from other species, that correspond to homologous regions to those surrounding the 135 high stringency σ54 binding sites in E. coli. In some cases, no homologous region was identified. A summary of the conservation analysis is shown in Fig 8, and a complete list of conservation scores is shown in S5 Table. Our analysis indicated that, as a group, intergenic binding sites (Fig 8A) are better conserved than intragenic sites (Fig 8B). However, many intragenic σ54 binding sites are conserved, suggesting that these sites are functional. Importantly, the computationally determined motif score correlates well with in vivo binding of σ54 (Fig 8C): the Spearman’s Rank Correlation Coefficient for the “OS” class of binding site (intergenic, oriented towards a gene) is 0.67, and for the rest of the binding sites is 0.38.


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

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

Conservation analysis of σ54 binding sites.Heat-maps depicting the match to the σ54 consensus binding site for each (A) canonical and (B) non-canonical σ54 binding site across a range of bacterial species. Genera are listed across the top, binding site ID numbers, and fold above threshold (FAT) scores in parentheses are listed to the left of the heat-map. For σ54 binding sites in panel A, the gene immediately downstream of each binding site is indicated to the right of the heat-map. (C) Comparison of the level of σ54 binding, as indicated by FAT score (Tables 1 & 2), versus Motif Score (S5 Table) for E. coli only. Different classes of binding site are indicated by color.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4591121&req=5

pgen.1005552.g008: Conservation analysis of σ54 binding sites.Heat-maps depicting the match to the σ54 consensus binding site for each (A) canonical and (B) non-canonical σ54 binding site across a range of bacterial species. Genera are listed across the top, binding site ID numbers, and fold above threshold (FAT) scores in parentheses are listed to the left of the heat-map. For σ54 binding sites in panel A, the gene immediately downstream of each binding site is indicated to the right of the heat-map. (C) Comparison of the level of σ54 binding, as indicated by FAT score (Tables 1 & 2), versus Motif Score (S5 Table) for E. coli only. Different classes of binding site are indicated by color.
Mentions: We speculated that other intragenic σ54 binding sites represent genuine promoters. We therefore compared each site across a range of bacterial species, mostly from the family Enterobacteriaceae. σ54 is well conserved across these species (e.g. 62% identical, 79% similar amino acid sequence between E. coli and V. cholerae), suggesting that it binds with similar DNA sequence specificity. Furthermore, ChIP-seq of σ54 in V. cholerae identified a very similar motif to the E. coli σ54 motif identified here (Fig 1B) [19], despite the two species having diverged >600 million years ago [40]. Therefore, we used a position weight matrix derived from our MEME analysis (Fig 1B) to score sequences, from other species, that correspond to homologous regions to those surrounding the 135 high stringency σ54 binding sites in E. coli. In some cases, no homologous region was identified. A summary of the conservation analysis is shown in Fig 8, and a complete list of conservation scores is shown in S5 Table. Our analysis indicated that, as a group, intergenic binding sites (Fig 8A) are better conserved than intragenic sites (Fig 8B). However, many intragenic σ54 binding sites are conserved, suggesting that these sites are functional. Importantly, the computationally determined motif score correlates well with in vivo binding of σ54 (Fig 8C): the Spearman’s Rank Correlation Coefficient for the “OS” class of binding site (intergenic, oriented towards a gene) is 0.67, and for the rest of the binding sites is 0.38.

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