CRP-Cyclic AMP Dependent Inhibition of the Xylene-Responsive σ54-Promoter Pu in Escherichia coli
The expression of σ54-dependent Pseudomonas putida Pu promoter is activated by XylR activator when cells are exposed to a variety of aromatic inducers. In this study, the transcriptional activation of the P. putida Pu promoter was recreated in the heterologous host Escherichia coli. Here we show tha...
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| Veröffentlicht in: | PloS one 2014-01, Vol.9 (1), p.e86727 |
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| description | The expression of σ54-dependent Pseudomonas putida Pu promoter is activated by XylR activator when cells are exposed to a variety of aromatic inducers. In this study, the transcriptional activation of the P. putida Pu promoter was recreated in the heterologous host Escherichia coli. Here we show that the cAMP receptor protein (CRP), a well-known carbon utilization regulator, had an inhibitory effect on the expression of Pu promoter in a cAMP-dependent manner. The inhibitory effect was not activator specific. In vivo KMnO4 and DMS footprinting analysis indicated that CRP-cAMP poised the RNA polymerase at Pu promoter, inhibiting the isomerization step of the transcription initiation even in the presence of an activator. Therefore, the presence of PTS-sugar, which eliminates cAMP, could activate the poised RNA polymerase at Pu promoter to transcribe. Moreover, the activation region 1 (AR1) of CRP, which interacts directly with the αCTD (C-terminal domain of α-subunit) of RNA polymerase, was found essential for the CRP-mediated inhibition at Pu promoter. A model for the above observations is discussed. |
| doi_str_mv | 10.1371/journal.pone.0086727 |
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In this study, the transcriptional activation of the P. putida Pu promoter was recreated in the heterologous host Escherichia coli. Here we show that the cAMP receptor protein (CRP), a well-known carbon utilization regulator, had an inhibitory effect on the expression of Pu promoter in a cAMP-dependent manner. The inhibitory effect was not activator specific. In vivo KMnO4 and DMS footprinting analysis indicated that CRP-cAMP poised the RNA polymerase at Pu promoter, inhibiting the isomerization step of the transcription initiation even in the presence of an activator. Therefore, the presence of PTS-sugar, which eliminates cAMP, could activate the poised RNA polymerase at Pu promoter to transcribe. Moreover, the activation region 1 (AR1) of CRP, which interacts directly with the αCTD (C-terminal domain of α-subunit) of RNA polymerase, was found essential for the CRP-mediated inhibition at Pu promoter. A model for the above observations is discussed.</description><identifier>ISSN: 1932-6203</identifier><identifier>EISSN: 1932-6203</identifier><identifier>DOI: 10.1371/journal.pone.0086727</identifier><identifier>PMID: 24466213</identifier><language>eng</language><publisher>San Francisco: Public Library of Science</publisher><subject>Amino acids ; Binding sites ; Biology ; Cyclic AMP ; Deoxyribonucleic acid ; DNA ; DNA-directed RNA polymerase ; E coli ; Escherichia coli ; Footprinting ; Genes ; Genomes ; Inhibition ; Isomerization ; Laboratories ; Life sciences ; Pathogens ; Plasmids ; Proteins ; Pseudomonas putida ; Regulation ; Ribonucleic acid ; RNA ; RNA polymerase ; Signal transduction ; Stem cells ; Studies ; Sugar ; Transcription activation ; Transcription initiation ; Xylene</subject><ispartof>PloS one, 2014-01, Vol.9 (1), p.e86727</ispartof><rights>2014 Zhang, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License: http://creativecommons.org/licenses/by/4.0/ (the “License”), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.</rights><rights>2014 Zhang, et al 2014 Zhang, et al</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c2497-519296dd3a5168ef7ae66c96b8791c265290b1b9426b5c5c9bac4f5d2d5d54a63</citedby><cites>FETCH-LOGICAL-c2497-519296dd3a5168ef7ae66c96b8791c265290b1b9426b5c5c9bac4f5d2d5d54a63</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC3900584/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC3900584/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,727,780,784,864,885,2928,23866,27924,27925,53791,53793,79600,79601</link.rule.ids></links><search><contributor>Chatterji, Dipankar</contributor><creatorcontrib>Zhang, Yuan-Tao</creatorcontrib><creatorcontrib>Jiang, Feng</creatorcontrib><creatorcontrib>Tian, Zhe-Xian</creatorcontrib><creatorcontrib>Huo, Yi-Xin</creatorcontrib><creatorcontrib>Sun, Yi-Cheng</creatorcontrib><creatorcontrib>Wang, Yi-Ping</creatorcontrib><title>CRP-Cyclic AMP Dependent Inhibition of the Xylene-Responsive σ54-Promoter Pu in Escherichia coli</title><title>PloS one</title><description>The expression of σ54-dependent Pseudomonas putida Pu promoter is activated by XylR activator when cells are exposed to a variety of aromatic inducers. In this study, the transcriptional activation of the P. putida Pu promoter was recreated in the heterologous host Escherichia coli. Here we show that the cAMP receptor protein (CRP), a well-known carbon utilization regulator, had an inhibitory effect on the expression of Pu promoter in a cAMP-dependent manner. The inhibitory effect was not activator specific. In vivo KMnO4 and DMS footprinting analysis indicated that CRP-cAMP poised the RNA polymerase at Pu promoter, inhibiting the isomerization step of the transcription initiation even in the presence of an activator. Therefore, the presence of PTS-sugar, which eliminates cAMP, could activate the poised RNA polymerase at Pu promoter to transcribe. Moreover, the activation region 1 (AR1) of CRP, which interacts directly with the αCTD (C-terminal domain of α-subunit) of RNA polymerase, was found essential for the CRP-mediated inhibition at Pu promoter. A model for the above observations is discussed.</description><subject>Amino acids</subject><subject>Binding sites</subject><subject>Biology</subject><subject>Cyclic AMP</subject><subject>Deoxyribonucleic acid</subject><subject>DNA</subject><subject>DNA-directed RNA polymerase</subject><subject>E coli</subject><subject>Escherichia coli</subject><subject>Footprinting</subject><subject>Genes</subject><subject>Genomes</subject><subject>Inhibition</subject><subject>Isomerization</subject><subject>Laboratories</subject><subject>Life sciences</subject><subject>Pathogens</subject><subject>Plasmids</subject><subject>Proteins</subject><subject>Pseudomonas putida</subject><subject>Regulation</subject><subject>Ribonucleic acid</subject><subject>RNA</subject><subject>RNA polymerase</subject><subject>Signal transduction</subject><subject>Stem cells</subject><subject>Studies</subject><subject>Sugar</subject><subject>Transcription activation</subject><subject>Transcription 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AMP Dependent Inhibition of the Xylene-Responsive σ54-Promoter Pu in Escherichia coli</title><author>Zhang, Yuan-Tao ; Jiang, Feng ; Tian, Zhe-Xian ; Huo, Yi-Xin ; Sun, Yi-Cheng ; Wang, Yi-Ping</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c2497-519296dd3a5168ef7ae66c96b8791c265290b1b9426b5c5c9bac4f5d2d5d54a63</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2014</creationdate><topic>Amino acids</topic><topic>Binding sites</topic><topic>Biology</topic><topic>Cyclic AMP</topic><topic>Deoxyribonucleic acid</topic><topic>DNA</topic><topic>DNA-directed RNA polymerase</topic><topic>E coli</topic><topic>Escherichia coli</topic><topic>Footprinting</topic><topic>Genes</topic><topic>Genomes</topic><topic>Inhibition</topic><topic>Isomerization</topic><topic>Laboratories</topic><topic>Life sciences</topic><topic>Pathogens</topic><topic>Plasmids</topic><topic>Proteins</topic><topic>Pseudomonas 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Yuan-Tao</au><au>Jiang, Feng</au><au>Tian, Zhe-Xian</au><au>Huo, Yi-Xin</au><au>Sun, Yi-Cheng</au><au>Wang, Yi-Ping</au><au>Chatterji, Dipankar</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>CRP-Cyclic AMP Dependent Inhibition of the Xylene-Responsive σ54-Promoter Pu in Escherichia coli</atitle><jtitle>PloS one</jtitle><date>2014-01-23</date><risdate>2014</risdate><volume>9</volume><issue>1</issue><spage>e86727</spage><pages>e86727-</pages><issn>1932-6203</issn><eissn>1932-6203</eissn><abstract>The expression of σ54-dependent Pseudomonas putida Pu promoter is activated by XylR activator when cells are exposed to a variety of aromatic inducers. In this study, the transcriptional activation of the P. putida Pu promoter was recreated in the heterologous host Escherichia coli. Here we show that the cAMP receptor protein (CRP), a well-known carbon utilization regulator, had an inhibitory effect on the expression of Pu promoter in a cAMP-dependent manner. The inhibitory effect was not activator specific. In vivo KMnO4 and DMS footprinting analysis indicated that CRP-cAMP poised the RNA polymerase at Pu promoter, inhibiting the isomerization step of the transcription initiation even in the presence of an activator. Therefore, the presence of PTS-sugar, which eliminates cAMP, could activate the poised RNA polymerase at Pu promoter to transcribe. Moreover, the activation region 1 (AR1) of CRP, which interacts directly with the αCTD (C-terminal domain of α-subunit) of RNA polymerase, was found essential for the CRP-mediated inhibition at Pu promoter. A model for the above observations is discussed.</abstract><cop>San Francisco</cop><pub>Public Library of Science</pub><pmid>24466213</pmid><doi>10.1371/journal.pone.0086727</doi><oa>free_for_read</oa></addata></record> |
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| subjects | Amino acids Binding sites Biology Cyclic AMP Deoxyribonucleic acid DNA DNA-directed RNA polymerase E coli Escherichia coli Footprinting Genes Genomes Inhibition Isomerization Laboratories Life sciences Pathogens Plasmids Proteins Pseudomonas putida Regulation Ribonucleic acid RNA RNA polymerase Signal transduction Stem cells Studies Sugar Transcription activation Transcription initiation Xylene |
| title | CRP-Cyclic AMP Dependent Inhibition of the Xylene-Responsive σ54-Promoter Pu in Escherichia coli |
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