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Identification of a new pebp2 alpha A2 isoform from zebrafish runx2 capable of inducing osteocalcin gene expression in vitro

2005-08Journal article Open access
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dc.contributor.authorPinto, Jorge
dc.contributor.authorConceição, Natércia
dc.contributor.authorS B Viegas, Carla
dc.contributor.authorLeite, Ricardo
dc.contributor.authorHurst, L. D.
dc.contributor.authorKelsh, R. N.
dc.contributor.authorLeonor Cancela, M.
dc.date.accessioned2018-12-07T14:52:43Z
dc.date.available2018-12-07T14:52:43Z
dc.date.issued2005-08
dc.description.abstractIntroduction: RUNX2 (also known as CBFA1/Osf2/AML3/PEBP2 alpha A) is a transcription factor essential for bone formation in mammals, as well as for osteoblast and chondrocyte differentiation, through regulation of expression of several bone- and cartilage-related genes. Since its discovery, Runx2 has been the subject of intense studies, mainly focused in unveiling regulatory targets of this transcription factor in high vertebrates. However, no single study has been published addressing the role of Runx2 in bone metabolism of low vertebrates. While analyzing the zebrafish (Danio rerio) runx2 gene, we identified the presence of two orthologs of RUNX2, which we named runx2a and runx2b and cloned a pebp2 alpha A-like transcript of the runx2b gene, which we named pebp2 alpha A2. Materials and Methods: Zebrafish runx2b gene and cDNA were isolated by RT-PCR and sequence data mining. The 3D structure of runx2b runt domain was modeled using mouse Runx1 runt as template. The regulatory effect of pebp2 alpha A2 on osteocalcin expression was analyzed by transient co-transfection experiments using a luciferase reporter gene. Phylogenetic analysis of available Runx sequences was performed with TREE-PUZZLE 5.2. and MrBayes. Results and Conclusions: We showed that the runx2b gene structure is highly conserved between mammals and fish. Zebrafish runx2b has two promoter regions separated by a large intron. Sequence analysis suggested that the runx2b gene encodes three distinct isoforms, by a combination of alternative splicing and differential promoter activation, as described for the human gene. We have cloned a pebp2 alpha A-like transcript of the runx2b gene, which we named pebp2 alpha A2, and showed its high degree of sequence similarity with the mammalian pebp2 alpha A. The cloned zebrafish osteocalcin promoter was found to contain three putative runx2-binding elements, and one of them, located at -221 from the ATG, was capable of mediating pebp2 alpha A2 transactivation. In addition, cross-species transactivation was also confirmed because the mouse Cbfa1 was able to induce the zebrafish osteocalcin promoter, whereas the zebrafish pebp2 alpha A2 activated the murine osteocalcin promoter. These results are consistent with the high degree of evolutionary conservation of these proteins. The 3D structure of the runx2b runt domain was modeled based on the runt domain of mouse Runx1. Results show a high degree of similarity in the 3D configuration of the DNA binding regions from both domains, with significant differences only observed in non-DNA binding regions or in DNA-binding regions known to accommodate considerable structure flexibility. Phylogenetic analysis was used to clarify the relationship between the isoforms of each of the two zebrafish Runx2 orthologs and other Runx proteins. Both zebrafish runx2 genes clustered with other Runx2 sequences. The duplication event seemed, however, to be so old that, whereas Runx2b clearly clusters with the other fish sequences, it is unclear whether Runx2a clusters with Runx2 from higher vertebrates or from other fish.
dc.description.versioninfo:eu-repo/semantics/publishedVersion
dc.identifier.doi10.1359/JBMR.050318
dc.identifier.issn0884-0431
dc.identifier.urihttp://hdl.handle.net/10400.1/11176
dc.language.isoeng
dc.peerreviewedyes
dc.publisherAmerican Society for Bone and Mineral Research
dc.relationEffect of extracellular calcium on MGP gene expression - Role of MGP in extracellular matrix mineralization
dc.relationIDENTIFICATION AND MOLECULAR CHARACTERIZATION OF NUCLEAR FACTORS RESPONSIBLE FOR MODULATION OF MATRIX GLA PROTEIN GENE EXPRESSION IN XENOPUS LAEVIS
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/
dc.subjectAcute myeloid-leukemia
dc.subjectMatrix gla protein
dc.subjectFish Sparus-Aurata
dc.subjectBone morphogenetic protein-2
dc.subjectMultiple sequence alignment
dc.subjectTranscription factor
dc.subjectOsteoblast differentiation
dc.subjectDna-Binding
dc.subjectCleidocranial dysplasia
dc.subjectGenomic structure
dc.titleIdentification of a new pebp2 alpha A2 isoform from zebrafish runx2 capable of inducing osteocalcin gene expression in vitro
dc.typejournal article
dspace.entity.typePublication
oaire.awardTitleEffect of extracellular calcium on MGP gene expression - Role of MGP in extracellular matrix mineralization
oaire.awardTitleIDENTIFICATION AND MOLECULAR CHARACTERIZATION OF NUCLEAR FACTORS RESPONSIBLE FOR MODULATION OF MATRIX GLA PROTEIN GENE EXPRESSION IN XENOPUS LAEVIS
oaire.awardURIinfo:eu-repo/grantAgreement/FCT/POCI/POCTI%2FBCI%2F48748%2F2002/PT
oaire.awardURIinfo:eu-repo/grantAgreement/FCT//SFRH%2FBPD%2F9451%2F2002/PT
oaire.citation.endPage1453
oaire.citation.issue8
oaire.citation.startPage1440
oaire.citation.titleJournal of Bone and Mineral Research
oaire.citation.volume20
oaire.fundingStreamPOCI
person.familyNamePinto
person.familyNameConceição
person.familyNameViegas
person.familyNameLeite
person.familyNameCancela
person.givenNameJorge
person.givenNameNatércia
person.givenNameCarla
person.givenNameRicardo
person.givenNameM. Leonor
person.identifier447622
person.identifier.ciencia-id7C11-760D-F425
person.identifier.ciencia-idC414-F596-EEC6
person.identifier.ciencia-idF115-256E-C84D
person.identifier.orcid0000-0003-1501-458X
person.identifier.orcid0000-0002-5057-0912
person.identifier.orcid0000-0002-5765-3665
person.identifier.orcid0000-0002-9622-3895
person.identifier.orcid0000-0003-3114-6662
person.identifier.ridJ-7827-2013
person.identifier.ridN-6695-2014
person.identifier.ridB-3389-2008
person.identifier.scopus-author-id8656187600
person.identifier.scopus-author-id8656310300
person.identifier.scopus-author-id8362650900
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.nameFundação para a Ciência e a Tecnologia
project.funder.nameFundação para a Ciência e a Tecnologia
rcaap.rightsopenAccess
rcaap.typearticle
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