Decreased stability of transforming growth factor beta type II receptor mRNA in RER+ human colon carcinoma cells.
Transforming growth factor beta (TGF-beta) is a potent inhibitor of cell growth and tumor progression. Previous work has shown that loss of functional TGF-beta type II receptor (RII) due to a frameshift mutation in the 5' half of the RII gene leads to TGF-beta 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 HCT116 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.
Jiang
W
W
Department of Biochemistry and Molecular Biology, Medical College of Ohio, Toledo 43699, USA.
Tillekeratne
M P
MP
Brattain
M G
MG
Banerji
S S
SS
D002277:Carcinoma / Q000235:genetics*; D002467:Cell Nucleus / Q000378:metabolism; D018388:Codon, Terminator; D003110:Colonic Neoplasms / Q000235:genetics*; D019008:Drug Resistance, Neoplasm; D016368:Frameshift Mutation; D006801:Humans; D017346:Protein Serine-Threonine Kinases; D012333:RNA, Messenger / Q000378:metabolism*; D012334:RNA, Neoplasm / Q000378:metabolism*; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta / Q000096:biosynthesis / Q000235:genetics*; D011995:Recombination, Genetic; D015398:Signal Transduction; D014158:Transcription, Genetic; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D018388:Codon, Terminator; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D018125:Receptors, Transforming Growth Factor beta; D016212:Transforming Growth Factor beta; D017346:Protein Serine-Threonine Kinases; D000077294:Receptor, Transforming Growth Factor-beta Type II