Characterization of the promoter regulatory region of the human pyruvate dehydrogenase beta gene.
A genomic clone (19 kb) harboring the intron-exon sequences and the promoter-regulatory region of the E1 beta gene of human pyruvate dehydrogenase complex was isolated by screening a placental genomic library. The nucleotide sequence of the promoter region (1245 bp) showed 18 differences (including mismatches, insertions, and deletions) as compared to the published sequence [Koike et al. (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 5594-5597]. The E1 beta promoter lacked a TATA box homology but contained initiator sequences (two) and Sp1 sites (three) which are frequently found in TATA-less promoters. The DNase I footprinting pattern of the promoter region with crude rat liver nuclear extracts showed at least seven regions of protein binding and nuclease protection (P1-P7). The DNase I protected regions contained consensus nucleotide sequences recognized by GATA-1, Sp1, IgNF-A, Lva, bicoid Q9, NF-kB, HNF-5, H4TF-1, WAP5, and ADH transription factors. Transient expression of chloramphenicol acetyltransferase (CAT) suggested the possible presence of negative elements located within the sequence from -2316 to -930, whereas deletion constructs containing -929 to +32 and -98 to +32 DNA sequences showed approximately 7- and 20-fold increases in CAT activity over the basal CAT activity. Additional studies indicated the presence of an orientation-dependent cis element (or elements) within the region from -282 to -397 that acts as an enhancer or a repressor upon a heterologous thymidine kinase promoter.(ABSTRACT TRUNCATED AT 250 WORDS)
Madhusudhan
K T
KT
Department of Biochemistry, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106.
Naik
S S
SS
Patel
M S
MS
D001483:Base Sequence; D001665:Binding Sites; D016384:Consensus Sequence; D004268:DNA-Binding Proteins / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D030481:Pyruvate Dehydrogenase (Lipoamide); D011768:Pyruvate Dehydrogenase Complex / Q000235:genetics*; D015183:Restriction Mapping; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; D011768:Pyruvate Dehydrogenase Complex; D014157:Transcription Factors; D030481:Pyruvate Dehydrogenase (Lipoamide)