Pitx3 promoter directs Cre-recombinase specifically in a human neuroblastoma cell line.
The Pitx3 gene is a homeobox transcription factor. This gene is expressed only in midbrain dopaminergic-neurons in the central nervous system, where its expression is important for development and survival of mesencephalic-dopaminergic neurons. The promoter region of the Pitx3 gene is not yet completely delimited. We used the Cre-loxP system to demonstrate the efficiency and specificity of a 4.2-kbp sequence in the 5'-flanking region of the Pitx3-gene promoter inserted in the 5'-terminus of the Cre-recombinase gene. A Cre-recombinase-reporter assay indicated that this 5'-flanking region possesses promoter activity. The cell-specific gene regulation of the Pitx3 promoter in neurons was demonstrated by a reverse-transcription polymerase chain reaction (RT-PCR) and Western blot detection of Cre-recombinase in SH-SY5Y cells but not in MCF7 cells. Functionality of the Cre-recombinase was confirmed in vitro. The Pitx3-promoter-Cre cassette here described can be used to develop transgenic mice with the specific expression of Cre-recombinase in midbrain-dopaminergic neurons to elucidate the gene function in which the conventional knockout leads to an early lethal phenotype. Such specific expression of the Pitx3 promoter may be used for gene therapy studies of Parkinson's disease.
Departamento de Histología, Facultad de Medicina, Universidad Autónoma de Nuevo León (UANL), Monterrey, NL, Mexico.
Diana L
DL
Castillo-Carranza
Humberto
H
Rodríguez-Rocha
Roberto
R
Montes-de-Oca-Luna
Julio
J
Sepúlveda-Saavedra
Héctor R
HR
Martínez
Yolanda
Y
López-Vidal
Odila
O
Saucedo-Cárdenas
D024506:5' Flanking Region / Q000235:genetics; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D015972:Gene Expression Regulation, Neoplastic; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D019426:Integrases / Q000235:genetics* / Q000378:metabolism*; D009447:Neuroblastoma / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011995:Recombination, Genetic / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic
D018398:Homeodomain Proteins; D014157:Transcription Factors; C433629:homeobox protein PITX3; C045073:Cre recombinase; D019426:Integrases