Regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase gene expression in FRTL-5 cells. II. Down-regulation by v-K-ras oncogene.
3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity and mRNA levels were significantly reduced in FRTL-5 cells transformed with the Kirsten-Moloney sarcoma virus (KiMol); these cells have lost thyrotropin dependence and express high levels of p21ras. FRTL-5 cells, transformed with a temperature-sensitive mutant of the v-K-ras oncogene (Ats cells: 33 degrees C, permissive; 39 degrees C, nonpermissive), showed significant reduction of HMG-CoA reductase expression when exposed to 33 degrees C. In KiMol cells, as well as in Ats cells at 33 degrees C, the transcription driven by cAMP-responsive element was probed by measuring chloramphenicol acetyl transferase (CAT) levels after transfection with a chimeric plasmid containing the reporter gene linked to the rat reductase promoter. Basal CAT activity in KiMol cells transfected with wild-type promoter was lower than in FRTL-5 cells but was increased by forskolin to the levels attained in thyrotropin-stimulated FRTL-5 cells. Forskolin failed to increase CAT activity in KiMol cells transfected with the plasmid harboring a reductase promoter in which the cAMP-responsive element octamer was mutated to a nonpalindromic sequence. The effect of v-K-ras could be mimicked in FRTL-5 cells by tetradecanoyl phorbol acetate and reverted in KiMol and Ats cells, expressing active Ras protein, by increasing intracellular cAMP and/or by protein kinase C inhibition. The data are consistent with the contention that v-K-ras, through protein kinase C and depletion of intracellular cAMP, is inhibitory for the protein kinase A pathway. This is the first demonstration that active v-K-ras down-regulates HMG-CoA reductase expression.
Perillo
B
B
Dipartimento di Biologia e Patologia Cellulare e Molecolare L. Califano, Università Federico II, Napoli, Italy.
Tedesco
I
I
Laezza
C
C
Santillo
M
M
Romano
A
A
Aloj
S M
SM
Bifulco
M
M
D000085:Acetates / Q000378:metabolism; D051698:Activating Transcription Factor 2; D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D002460:Cell Line; D002461:Cell Line, Transformed; D002471:Cell Transformation, Neoplastic; D005576:Colforsin / Q000494:pharmacology; D017362:Cyclic AMP Response Element-Binding Protein / Q000276:immunology / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D011905:Genes, ras; D006903:Hydroxymethylglutaryl CoA Reductases / Q000096:biosynthesis* / Q000378:metabolism; D007700:Kinetics; D007708:Kirsten murine sarcoma virus / Q000235:genetics; D016350:Leucine Zippers; D008980:Moloney murine sarcoma virus / Q000235:genetics; D016296:Mutagenesis; D011493:Protein Kinase C / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D013696:Temperature; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013961:Thyroid Gland / Q000201:enzymology; D014157:Transcription Factors
D000085:Acetates; D051698:Activating Transcription Factor 2; D000906:Antibodies; D017362:Cyclic AMP Response Element-Binding Protein; D012333:RNA, Messenger; D014157:Transcription Factors; D005576:Colforsin; D006903:Hydroxymethylglutaryl CoA Reductases; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate