Cyclin D1 antisense RNA destabilizes pRb and retards lung cancer cell growth.
To investigate the role of cyclin D1 in the regulation of lung cancer cell growth, we created five stably transfected cell lines carrying a cyclin D1 antisense construct. The transfected cells exhibited a marked decrease in the rate of cell growth, in contrast to the original lines (A549 and NCI-H441). The expression of several cell cycle-regulating proteins, including cyclin A, the cyclin-dependent kinases (cdk) 2 and cdk4, in addition to cyclin D1 itself, was markedly decreased. The expression of one cdk inhibitor, p21WAF1/CIP1, increased in the A549-derived cell lines. A specific target of cyclin D1 activity, the growth-suppressing product of the retinoblastoma gene, pRb, exhibited decreased expression and a decreased level of phosphorylation in the transfected cells. Decreased expression of pRb due to a significant increase in its turnover rate suggested that the stability of the protein may depend on phosphorylation by cyclin D1-dependent cdk activity. In addition to the impact on pRb stability, decreased expression of cyclin D1 induced susceptibility to cell death after withdrawal of exogenous growth factors in the antisense transfected cell lines, a response that was not observed in the original cancer cell lines. We conclude that abrogation of cyclin D1 overexpression in lung cancer cells disrupts several key pathways that are required for uncontrolled cell growth and induces those that lead to cell death after growth factor deprivation. Therefore, we speculate that use of antisense cyclin D1 expression in appropriate gene vectors could be a useful method for retarding lung cancer cell growth in accessible tumors such as those of the lung epithelium.
Driscoll
B
B
Deparment of Surgery, Childrens Hospital Los Angeles Research Institute, University of Southern California School of Medicine 90027, USA.
Wu
L
L
Buckley
S
S
Hall
F L
FL
Anderson
K D
KD
Warburton
D
D
D042846:CDC2-CDC28 Kinases; D016923:Cell Death / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D016895:Culture Media, Serum-Free; D019925:Cyclin A / Q000096:biosynthesis; D019938:Cyclin D1 / Q000235:genetics*; D056742:Cyclin D2; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D050759:Cyclin-Dependent Kinase Inhibitor p21; D018844:Cyclin-Dependent Kinases / Q000096:biosynthesis; D016213:Cyclins / Q000096:biosynthesis; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006801:Humans; D007700:Kinetics; D008175:Lung Neoplasms; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D017346:Protein Serine-Threonine Kinases / Q000096:biosynthesis; D011518:Proto-Oncogene Proteins; D016372:RNA, Antisense / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D014162:Transfection; D014407:Tumor Cells, Cultured
C080336:CCND2 protein, human; C493956:CDKN1A protein, human; D016895:Culture Media, Serum-Free; D019925:Cyclin A; D056742:Cyclin D2; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D011518:Proto-Oncogene Proteins; D016372:RNA, Antisense; D011994:Recombinant Proteins; D016160:Retinoblastoma Protein; D019938:Cyclin D1; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; C495900:CDK4 protein, human; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D018844:Cyclin-Dependent Kinases