Lung cell-specific expression of the murine surfactant protein A (SP-A) gene is mediated by interactions between the SP-A promoter and thyroid transcription factor-1.
Cis-acting elements determining lung epithelial cell-selective transcription of the murine surfactant protein A (SP-A) gene were identified between nucleotide positions -255 and -57. This region of the murine SP-A gene contained nucleotide sequences consistent with thyroid transcription factor-1 (TTF-1) binding motifs. An SP-A-CAT plasmid containing the TTF-1 binding sites was transcriptionally active in mouse lung epithelial (MLE-15) cells but not in HeLa, 3T3, or H441 cells. However, transcription of the SP-A-CAT construct was activated after cotransfection of HeLa cells with a vector expressing recombinant TTF-1, pCMV-TTF-1. Recombinant TTF-1 homeodomain protein bound to four distinct binding sites located between nucleotides -166 to -117 [corrected]. Proteins in nuclear extracts of MLE-15 cells bound TTF-1 binding sites and were supershifted by TTF-1 antibody. Mutations of three of the TTF-1 binding sites in this region reduced expression of the SP-A-CAT construct in transfected MLE-15 cells and reduced transactivation in HeLa cells. TTF-1 interacts with complex protein/DNA binding sites located in the 5'-flanking region of the murine SP-A gene enhancing lung epithelial cell-specific expression in vitro.
Bruno
M D
MD
Division of Pulmonary Biology, Children's Hospital Medical Center, Cincinnati, Ohio 45229-3039.
Bohinski
R J
RJ
Huelsman
K M
KM
Whitsett
J A
JA
Korfhagen
T R
TR
D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D008168:Lung / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000502:physiology*; D011401:Promoter Regions, Genetic; D011510:Proteolipids / Q000235:genetics*; D037662:Pulmonary Surfactant-Associated Protein A; D037661:Pulmonary Surfactant-Associated Proteins; D011663:Pulmonary Surfactants / Q000235:genetics*; D000074482:Thyroid Nuclear Factor 1; D014157:Transcription Factors / Q000502:physiology*; D015533:Transcriptional Activation
C000614412:NKX2-1 protein, human; C000614414:Nkx2-1 protein, mouse; D009687:Nuclear Proteins; D011510:Proteolipids; D037662:Pulmonary Surfactant-Associated Protein A; D037661:Pulmonary Surfactant-Associated Proteins; D011663:Pulmonary Surfactants; D000074482:Thyroid Nuclear Factor 1; D014157:Transcription Factors