Role of far upstream repressor elements controlling proto-Ha-ras gene transcription.
The far upstream region of the rat Ha-ras gene has been characterized to determine whether possible repressor sequences may control the low level of Ha-ras gene transcription from its TATA-less, GC-rich strong promoter. The chloramphenicol acetyl transferase (CAT) gene under the control of the 3.8-kb Ha-ras upstream promoter was minimally expressed in HeLa cells. Surprisingly, CAT gene expression was increased by the deletion of a 0.7-kb BglII fragment containing non-coding exon minus 2 and TATA box promoter elements located 1.7 kb upstream of the GC-rich strong promoter. Far upstream (CA)25 repeats also appeared to repress Ha-ras gene activity. Sequences within the 0.7-kb BglII fragment suppressed CAT gene expression when placed upstream of a heterologous thymidine kinase (tk) gene promoter. Repressor activity was further localized to a 160-bp AvrII-BglII sub-fragment. Gel shift assays identified two sequence-specific DNA binding proteins. The results demonstrated for the first time that far upstream repressor sequences control normal transcription of the Ha-ras proto-oncogene.
Chakraborty
A K
AK
Indian Institute of Chemical Biology (CSIR), Calcutta, 700032, India.
Hodgson
C P
CP
D000818:Animals; D001426:Bacterial Proteins; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D015245:Deoxyribonuclease BamHI / Q000378:metabolism; D015247:Deoxyribonuclease HindIII / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D005786:Gene Expression Regulation; D011905:Genes, ras / Q000235:genetics*; D011401:Promoter Regions, Genetic; D051381:Rats; D012097:Repressor Proteins / Q000502:physiology*; D017422:Sequence Analysis, DNA; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D012097:Repressor Proteins; D015500:Chloramphenicol O-Acetyltransferase; D015245:Deoxyribonuclease BamHI; D015247:Deoxyribonuclease HindIII; C024404:BglII endonuclease; C064053:CCTAGG-specific type II deoxyribonucleases; D015252:Deoxyribonucleases, Type II Site-Specific