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Pseudomonas aeruginosa biofilm matrix polysaccharide Psl is regulated transcriptionally by RpoS and post-transcriptionally by RsmA.

Irie Y, Starkey M, Edwards AN, Wozniak DJ, Romeo T, Parsek MR - Mol. Microbiol. (2010)

Bottom Line: In this study, we demonstrate that the alternative σ-factor RpoS is a positive transcriptional regulator of psl gene expression.Furthermore, we show that psl mRNA has an extensive 5' untranslated region, to which the post-transcriptional regulator RsmA binds and represses psl translation.This constitutes a novel mechanism for translational repression by this family of regulators.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of Washington, Seattle, WA 98195, USA.

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Transcriptional fusion studies confirm the 5′ RACE-derived transcriptional start site of psl.A. Transcriptional lacZ fusion constructs of psl promoter region. Four representative transcriptional fusion constructs span the corresponding regions as indicated by the black bars, with the locations of the two putative transcriptional start sites. The approximate positions of the TR1, TR2, TR5, DN1 and DN3 primers that were used to construct the transcriptional fusions are shown as horizontal arrows.B. Relative expression of the four representative transcriptional fusion constructs. The results indicate that the promoter associated with the +1 transcriptional start site has the highest activity instead of the previously published +108 transcriptional start position. The y-axis unit is described as β-galactosidase activity determined by the Galacto-Light Plus kit divided by total mg of protein from the cell lysates as determined by Bradford assay.
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fig02: Transcriptional fusion studies confirm the 5′ RACE-derived transcriptional start site of psl.A. Transcriptional lacZ fusion constructs of psl promoter region. Four representative transcriptional fusion constructs span the corresponding regions as indicated by the black bars, with the locations of the two putative transcriptional start sites. The approximate positions of the TR1, TR2, TR5, DN1 and DN3 primers that were used to construct the transcriptional fusions are shown as horizontal arrows.B. Relative expression of the four representative transcriptional fusion constructs. The results indicate that the promoter associated with the +1 transcriptional start site has the highest activity instead of the previously published +108 transcriptional start position. The y-axis unit is described as β-galactosidase activity determined by the Galacto-Light Plus kit divided by total mg of protein from the cell lysates as determined by Bradford assay.

Mentions: To investigate the discrepancies in the results, we designed a series of nested transcriptional fusion reporter constructs that spanned various regions upstream of pslA. The representative constructs are displayed in Fig. 2A. As shown in Fig. 2B, the reporters containing the promoter region from our study (TR1→DN1 and TR1→DN3) had significantly higher transcriptional activity compared with the reporter containing only the promoter reported in the Overhage study (TR2→DN1 and TR5→DN1). While it is possible that psl has two distinct promoters, we conclude that the majority of the psl transcripts initiate at 148 bp upstream of pslA ORF (designated as +1 in this paper).


Pseudomonas aeruginosa biofilm matrix polysaccharide Psl is regulated transcriptionally by RpoS and post-transcriptionally by RsmA.

Irie Y, Starkey M, Edwards AN, Wozniak DJ, Romeo T, Parsek MR - Mol. Microbiol. (2010)

Transcriptional fusion studies confirm the 5′ RACE-derived transcriptional start site of psl.A. Transcriptional lacZ fusion constructs of psl promoter region. Four representative transcriptional fusion constructs span the corresponding regions as indicated by the black bars, with the locations of the two putative transcriptional start sites. The approximate positions of the TR1, TR2, TR5, DN1 and DN3 primers that were used to construct the transcriptional fusions are shown as horizontal arrows.B. Relative expression of the four representative transcriptional fusion constructs. The results indicate that the promoter associated with the +1 transcriptional start site has the highest activity instead of the previously published +108 transcriptional start position. The y-axis unit is described as β-galactosidase activity determined by the Galacto-Light Plus kit divided by total mg of protein from the cell lysates as determined by Bradford assay.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2984543&req=5

fig02: Transcriptional fusion studies confirm the 5′ RACE-derived transcriptional start site of psl.A. Transcriptional lacZ fusion constructs of psl promoter region. Four representative transcriptional fusion constructs span the corresponding regions as indicated by the black bars, with the locations of the two putative transcriptional start sites. The approximate positions of the TR1, TR2, TR5, DN1 and DN3 primers that were used to construct the transcriptional fusions are shown as horizontal arrows.B. Relative expression of the four representative transcriptional fusion constructs. The results indicate that the promoter associated with the +1 transcriptional start site has the highest activity instead of the previously published +108 transcriptional start position. The y-axis unit is described as β-galactosidase activity determined by the Galacto-Light Plus kit divided by total mg of protein from the cell lysates as determined by Bradford assay.
Mentions: To investigate the discrepancies in the results, we designed a series of nested transcriptional fusion reporter constructs that spanned various regions upstream of pslA. The representative constructs are displayed in Fig. 2A. As shown in Fig. 2B, the reporters containing the promoter region from our study (TR1→DN1 and TR1→DN3) had significantly higher transcriptional activity compared with the reporter containing only the promoter reported in the Overhage study (TR2→DN1 and TR5→DN1). While it is possible that psl has two distinct promoters, we conclude that the majority of the psl transcripts initiate at 148 bp upstream of pslA ORF (designated as +1 in this paper).

Bottom Line: In this study, we demonstrate that the alternative σ-factor RpoS is a positive transcriptional regulator of psl gene expression.Furthermore, we show that psl mRNA has an extensive 5' untranslated region, to which the post-transcriptional regulator RsmA binds and represses psl translation.This constitutes a novel mechanism for translational repression by this family of regulators.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of Washington, Seattle, WA 98195, USA.

Show MeSH