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Article: Transcriptional regulation of the promoter of the rat frizzled related protein gene by CREB

TitleTranscriptional regulation of the promoter of the rat frizzled related protein gene by CREB
Authors
Issue Date2003
PublisherNature Publishing Group. The Journal's web site is located at http://www.nature.com/onc
Citation
Oncogene, 2003, v. 22 n. 25, p. 3901-3910 How to Cite?
AbstractFrizzled related proteins (Frps) are secreted proteins structurally similar to frizzled receptors; they bind Wnt via the cysteine-rich domain and antagonize the Wnt signaling pathway. In this study, we have investigated the mechanisms regulating the transcriptional regulation of rat Frp (rFrp) promoter. From previous findings, we know that the transcriptional activation domain of rFrp resides in the region -202 to -144 relative to the transcription start site, and that it is essential for efficient promoter activity. The study presented here was designed to identify trans-acting factors that bind to this critical domain of the rFrp promoter and to elucidate the pathway involved in the regulation of rFrp expression. Electrophoretic mobility shift assay (EMSA) demonstrated that specific DNA-protein binding activities fall into two adjacent core sequences with (CTTTGGGGG) at -197 to -189 and (AGATGATGTAA) at -151 to -141 of the rFrp promoter. Reporter assay showed that these core sequences are both required for the activation of rFrp promoter. Mutation within either one or both core sequence drastically reduced the promoter activity. Southwestern blotting showed that the estimated molecular mass of the distinct binding protein to the (AGATGATGTAA) domain is about 43 kDa. Further EMSA suggested CREB as the trans-acting factor in the DNA-protein complex, which was out competed by CREB consensus oligonucleotides and supershifted by anti-CREB antibody. Overexpression of PKA and CREB also transactivated rFrp promoter, and dominant-negative CREB inhibited the promoter activity in transient reporter assays. More importantly, CREB, phosphorylated CREB and the adaptor protein CBP were found binding to the endogenous rFrp promoter using chromatin immunoprecipitation assay. Collectively, our results demonstrate the induction of rFrp promoter activity by PKA and CREB in vitro, and the binding of CREB and CBP to the rFrp promoter core motif in vivo.
Persistent Identifierhttp://hdl.handle.net/10722/88786
ISSN
2015 Impact Factor: 7.932
2015 SCImago Journal Rankings: 4.047
ISI Accession Number ID
References

 

DC FieldValueLanguage
dc.contributor.authorYam, JWPen_HK
dc.contributor.authorChan, KWen_HK
dc.contributor.authorHsiao, WLWen_HK
dc.date.accessioned2010-09-06T09:47:59Z-
dc.date.available2010-09-06T09:47:59Z-
dc.date.issued2003en_HK
dc.identifier.citationOncogene, 2003, v. 22 n. 25, p. 3901-3910en_HK
dc.identifier.issn0950-9232en_HK
dc.identifier.urihttp://hdl.handle.net/10722/88786-
dc.description.abstractFrizzled related proteins (Frps) are secreted proteins structurally similar to frizzled receptors; they bind Wnt via the cysteine-rich domain and antagonize the Wnt signaling pathway. In this study, we have investigated the mechanisms regulating the transcriptional regulation of rat Frp (rFrp) promoter. From previous findings, we know that the transcriptional activation domain of rFrp resides in the region -202 to -144 relative to the transcription start site, and that it is essential for efficient promoter activity. The study presented here was designed to identify trans-acting factors that bind to this critical domain of the rFrp promoter and to elucidate the pathway involved in the regulation of rFrp expression. Electrophoretic mobility shift assay (EMSA) demonstrated that specific DNA-protein binding activities fall into two adjacent core sequences with (CTTTGGGGG) at -197 to -189 and (AGATGATGTAA) at -151 to -141 of the rFrp promoter. Reporter assay showed that these core sequences are both required for the activation of rFrp promoter. Mutation within either one or both core sequence drastically reduced the promoter activity. Southwestern blotting showed that the estimated molecular mass of the distinct binding protein to the (AGATGATGTAA) domain is about 43 kDa. Further EMSA suggested CREB as the trans-acting factor in the DNA-protein complex, which was out competed by CREB consensus oligonucleotides and supershifted by anti-CREB antibody. Overexpression of PKA and CREB also transactivated rFrp promoter, and dominant-negative CREB inhibited the promoter activity in transient reporter assays. More importantly, CREB, phosphorylated CREB and the adaptor protein CBP were found binding to the endogenous rFrp promoter using chromatin immunoprecipitation assay. Collectively, our results demonstrate the induction of rFrp promoter activity by PKA and CREB in vitro, and the binding of CREB and CBP to the rFrp promoter core motif in vivo.en_HK
dc.languageengen_HK
dc.publisherNature Publishing Group. The Journal's web site is located at http://www.nature.com/oncen_HK
dc.relation.ispartofOncogeneen_HK
dc.subject.meshAnimalsen_HK
dc.subject.meshBinding Sitesen_HK
dc.subject.meshCells, Cultureden_HK
dc.subject.meshCyclic AMP Response Element-Binding Protein - genetics - physiologyen_HK
dc.subject.meshCyclic AMP-Dependent Protein Kinases - physiologyen_HK
dc.subject.meshElectrophoretic Mobility Shift Assayen_HK
dc.subject.meshGene Expression Regulationen_HK
dc.subject.meshGenes, Reporteren_HK
dc.subject.meshMacromolecular Substancesen_HK
dc.subject.meshPoint Mutationen_HK
dc.subject.meshPromoter Regions, Genetic - geneticsen_HK
dc.subject.meshProteins - geneticsen_HK
dc.subject.meshRatsen_HK
dc.subject.meshTranscriptional Activationen_HK
dc.titleTranscriptional regulation of the promoter of the rat frizzled related protein gene by CREBen_HK
dc.typeArticleen_HK
dc.identifier.openurlhttp://library.hku.hk:4550/resserv?sid=HKU:IR&issn=0950-9232&volume=22&issue=25&spage=3901&epage=3910&date=2003&atitle=Transcriptional+regulation+of+the+promoter+of+the+rat+frizzled+related+protein+gene+by+CREBen_HK
dc.identifier.emailYam, JWP:judyyam@pathology.hku.hken_HK
dc.identifier.authorityYam, JWP=rp00468en_HK
dc.description.naturelink_to_subscribed_fulltext-
dc.identifier.doi10.1038/sj.onc.1206489en_HK
dc.identifier.pmid12813463-
dc.identifier.scopuseid_2-s2.0-0038001068en_HK
dc.identifier.hkuros78573en_HK
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-0038001068&selection=ref&src=s&origin=recordpageen_HK
dc.identifier.volume22en_HK
dc.identifier.issue25en_HK
dc.identifier.spage3901en_HK
dc.identifier.epage3910en_HK
dc.identifier.isiWOS:000183612000009-
dc.publisher.placeUnited Kingdomen_HK

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