Characterization of the lung Krüppel-like transcription factor gene and upstream regulatory elements.
We previously described the isolation and characterization of the cDNA for lung Krüppel-like factor (LKLF), a zinc finger transcription factor that is predominately expressed in the lung of adult mice. In this study, we report the complete structure and nucleotide sequence of the mouse LKLF gene, which is comprised of three exons and two small introns. Moreover, the identification of critical sequence elements required for expression is described using reporter constructs with the LKLF promoter transfected into LA-4 lung cells. Results from these constructs reveal an important region for transcriptional activity that lies between the -490/-72bp upstream sequence. This region contains two canonical Sp1 binding sites that affect expression levels in a non tissue-specific manner. In addition, using a base-pair mutagenesis strategy, a region from -157/-72bp was found to be necessary for upregulating expression. In transfection assays, mutations of the -138/-111bp region resulted in approximately 70-80% loss of promoter activity. This cis-element does not appear to correspond to any known transcription factor consensus sequence. Moreover, mutations within this cis-region disrupt the binding of a protein complex from nuclear extracts of various tissues.
Schrick
J J
JJ
Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati, Cincinnati, OH 45267-0524, USA.
Hughes
M J
MJ
Anderson
K P
KP
Croyle
M L
ML
Lingrel
J B
JB
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002460:Cell Line; D004586:Electrophoresis; D005091:Exons; D017353:Gene Deletion; D017930:Genes, Reporter; D051741:Kruppel-Like Transcription Factors; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011401:Promoter Regions, Genetic; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics*; D015534:Trans-Activators / Q000235:genetics*; D014158:Transcription, Genetic; D014162:Transfection; D015854:Up-Regulation
C498314:Klf2 protein, mouse; D051741:Kruppel-Like Transcription Factors; D015534:Trans-Activators