TY - JOUR
T1 - Decreased stability of transforming growth factor β type II receptor mRNA in RER+ human colon carcinoma cells
AU - Jiang, Wen
AU - Tillekeratne, Manoranjani P.M.
AU - Brattain, Michael G.
AU - Banerji, Sunandita S.
PY - 1997/12/2
Y1 - 1997/12/2
N2 - Transforming growth factor β (TGF-β) is a potent inhibitor of cell growth and tumor progression. Previous work has shown that loss of functional TGF-β type II receptor (RII) due to a frameshift mutation in the 5' half of the RII gene leads to TGF-β resistance in a highly progressed, RER+ human colon carcinoma cell line designated HCT116. Expression of this mutated RII gene was highly repressed in RER+ cell lines such as HCTI 16 and RKO, as analyzed by RNase protection assays. Nuclear run-on and RII promoter-reporter (CAT) assays showed that the transcriptional levels of the RII gene in these RER+ cells were not reduced, compared to RII-expressing cells. However, the half-lives of the RII mRNA, as analyzed by RNase protection assays following actinomycin D treatment, were significantly decreased. This suggested that the decreased expression of the RII gene mutant was due to decreased mRNA stability. Furthermore, RII mRNA from HCT116 transfected with wild-type RII had a longer half-life than the endogenous mutated RII mRNA. A dominant negative RII mutant, which encodes a similarly truncated RII protein as HCT116 but lacks the extensive 3' untranslated region of RII mRNA, gave the same half-life as endogenous wild-type RII mRNA. We conclude that the frameshift mutation which results in a premature stop codon in the 5' half of the mRNA transcript accounts for the reduced RII mRNA levels in RER+ cells.
AB - Transforming growth factor β (TGF-β) is a potent inhibitor of cell growth and tumor progression. Previous work has shown that loss of functional TGF-β type II receptor (RII) due to a frameshift mutation in the 5' half of the RII gene leads to TGF-β resistance in a highly progressed, RER+ human colon carcinoma cell line designated HCT116. Expression of this mutated RII gene was highly repressed in RER+ cell lines such as HCTI 16 and RKO, as analyzed by RNase protection assays. Nuclear run-on and RII promoter-reporter (CAT) assays showed that the transcriptional levels of the RII gene in these RER+ cells were not reduced, compared to RII-expressing cells. However, the half-lives of the RII mRNA, as analyzed by RNase protection assays following actinomycin D treatment, were significantly decreased. This suggested that the decreased expression of the RII gene mutant was due to decreased mRNA stability. Furthermore, RII mRNA from HCT116 transfected with wild-type RII had a longer half-life than the endogenous mutated RII mRNA. A dominant negative RII mutant, which encodes a similarly truncated RII protein as HCT116 but lacks the extensive 3' untranslated region of RII mRNA, gave the same half-life as endogenous wild-type RII mRNA. We conclude that the frameshift mutation which results in a premature stop codon in the 5' half of the mRNA transcript accounts for the reduced RII mRNA levels in RER+ cells.
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U2 - 10.1021/bi9717892
DO - 10.1021/bi9717892
M3 - Article
C2 - 9402753
AN - SCOPUS:0030734793
SN - 0006-2960
VL - 36
SP - 14786
EP - 14793
JO - Biochemistry
JF - Biochemistry
IS - 48
ER -