Gene expression promoted by the SV40 DNA targeting sequence and the hypoxia-responsive element under normoxia and hypoxia

Gene expression promoted by the SV40 DNA targeting sequence and the hypoxia-responsive element under normoxia and hypoxia

Autor Sacramento, Chester Bittencourt Autor UNIFESP Google Scholar
Moraes, Jane Zveiter de Autor UNIFESP Google Scholar
Denapolis, P.m.a. Autor UNIFESP Google Scholar
Han, Sang Won Autor UNIFESP Google Scholar
Instituição Universidade Federal de São Paulo (UNIFESP)
Resumo The main objective of the present study was to find suitable DNA-targeting sequences (DTS) for the construction of plasmid vectors to be used to treat ischemic diseases. The well-known Simian virus 40 nuclear DTS (SV40-DTS) and hypoxia-responsive element (HRE) sequences were used to construct plasmid vectors to express the human vascular endothelial growth factor gene (hVEGF). The rate of plasmid nuclear transport and consequent gene expression under normoxia (20% O2) and hypoxia (less than 5% O2) were determined. Plasmids containing the SV40-DTS or HRE sequences were constructed and used to transfect the A293T cell line (a human embryonic kidney cell line) in vitro and mouse skeletal muscle cells in vivo. Plasmid transport to the nucleus was monitored by real-time PCR, and the expression level of the hVEGF gene was measured by ELISA. The in vitro nuclear transport efficiency of the SV40-DTS plasmid was about 50% lower under hypoxia, while the HRE plasmid was about 50% higher under hypoxia. Quantitation of reporter gene expression in vitro and in vivo, under hypoxia and normoxia, confirmed that the SV40-DTS plasmid functioned better under normoxia, while the HRE plasmid was superior under hypoxia. These results indicate that the efficiency of gene expression by plasmids containing DNA binding sequences is affected by the concentration of oxygen in the medium.
Palavra-chave SV40-DTS
Hypoxia-responsive element
Transfection efficiency
Normoxia
Hypoxia
Ischemic diseases
Idioma Inglês
Financiador Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)
Número do financiamento FAPESP: 06/59630-0
Data de publicação 2010-08-01
Publicado em Brazilian Journal of Medical and Biological Research. Associação Brasileira de Divulgação Científica, v. 43, n. 8, p. 722-727, 2010.
ISSN 0100-879X (Sherpa/Romeo, fator de impacto)
Publicador Associação Brasileira de Divulgação Científica
Extensão 722-727
Fonte http://dx.doi.org/10.1590/S0100-879X2010007500064
Direito de acesso Acesso aberto Open Access
Tipo Artigo
Web of Science WOS:000283261800005
SciELO S0100-879X2010000800005 (estatísticas na SciELO)
Endereço permanente http://repositorio.unifesp.br/handle/11600/5881

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