Document Detail


Isolation and characterization of the promoter region of the rat kidney-type glutaminase gene.
MedLine Citation:
PMID:  11267668     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
A lambdaEMBL3 rat genomic library was screened to clone a phage that contained the promoter region of the kidney-type mitochondrial glutaminase gene. The resulting lambdaGA1 phage contained 13.7 kb of genomic DNA that was mapped by Southern blotting and restriction analysis. The 2.22 kb and 0.83 kb SacI fragments of lambdaGA1 were sequenced and the transcription initiation site was identified by RNase mapping. The reported sequence contains 2287 bp of the promoter, the entire exon 1 (542 bp), and 223 bp of the initial intron of the glutaminase gene. The initial exon contains 141 bp of 5'-nontranslated sequence and 401 bp of coding sequence that encodes the 72-amino acid mitochondrial targeting presequence and 61 amino acids from the N-terminus of the mature 66 kDa glutaminase subunit. Various segments of the GA promoter were cloned into a chloramphenicol acetyltransferase (CAT) expression vector. The resulting GA-CAT constructs were transfected into LLC-PK(1)-F(+) kidney cells to assess the promoter function of the isolated genomic DNA. The GA(-402)CAT construct produced a 10-fold greater CAT activity than the promoter-less pCAT vector. Analysis of various deletion constructs indicated that elements located between -402 and -63 bp must act in synergy with more proximal elements to create a functional promoter. The initial 402 bp segment lacks a TATA sequence but is GC-rich and contains two CCAAT boxes and two Sp1 sites.
Authors:
L Taylor; X Liu; W Newsome; R A Shapiro; M Srinivasan; N P Curthoys
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Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Biochimica et biophysica acta     Volume:  1518     ISSN:  0006-3002     ISO Abbreviation:  Biochim. Biophys. Acta     Publication Date:  2001 Mar 
Date Detail:
Created Date:  2001-03-27     Completed Date:  2001-06-28     Revised Date:  2008-11-21    
Medline Journal Info:
Nlm Unique ID:  0217513     Medline TA:  Biochim Biophys Acta     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  132-6     Citation Subset:  IM    
Affiliation:
Department of Biochemistry and Molecular Biology, Colorado State University, 80523-1870, Fort Collins, CO, USA.
Data Bank Information
Bank Name/Acc. No.:
GENBANK/AF302091
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MeSH Terms
Descriptor/Qualifier:
Amino Acid Sequence
Animals
Base Sequence
Chloramphenicol O-Acetyltransferase / genetics
DNA, Complementary
Genes, Reporter
Glutaminase / genetics*
Kidney / enzymology*
LLC-PK1 Cells
Molecular Sequence Data
Promoter Regions, Genetic*
Rats
Swine
Grant Support
ID/Acronym/Agency:
DK 37124/DK/NIDDK NIH HHS
Chemical
Reg. No./Substance:
0/DNA, Complementary; EC 2.3.1.28/Chloramphenicol O-Acetyltransferase; EC 3.5.1.2/Glutaminase

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