Transcriptional regulatory and response mapping of the rat Ha-ras upstream sequence using primary mammary epithelial cells.
The rat Ha-ras upstream sequence (-2876 to +986 bp relative to the most 5' transcriptional start site) was transcriptionally mapped at a gross level. Ha-ras upstream sequence and 5' unidirectional deletion reporter constructs were transfected via particle bombardment into primary cultures of rat mammary epithelial cells. Analyses of Ha-ras reporter expression show that a fragment extending from -2876 to -2110 bp contains a positive regulatory sequence. The majority of Ha-ras expression is attributed to this sequence, since its deletion results in a 4-fold decrease in expression. The Ha-ras gene was also assessed for autoregulation using similar co-transfection experiments. Wild-type and activated (codon 12 G-->A transition) Ha-ras expression vectors, transcriptionally driven by Ha-ras upstream sequence, were co-transfected with Ha-ras upstream sequence deletion reporter constructs. The activated Ha-ras gene product induced its own transcription 2-fold, targeting the regulatory region between -2119 and -313 bp.
Ariazi
E A
EA
Department of Human Oncology, University of Wisconsin-Madison 53792, USA.
Thompson
T A
TA
Burkholder
J K
JK
Yang
N S
NS
Gould
M N
MN
D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D002874:Chromosome Mapping; D004848:Epithelium / Q000502:physiology; D005260:Female; D017353:Gene Deletion; D005809:Genes, Regulator; D011905:Genes, ras; D008321:Mammary Glands, Animal / Q000502:physiology*; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic; D051381:Rats; D011920:Rats, Inbred WF; D017422:Sequence Analysis, DNA / Q000379:methods; D014158:Transcription, Genetic; D014162:Transfection