Identification of cJun-responsive genes in Rat-1a cells using multiple techniques: increased expression of stathmin is necessary for cJun-mediated anchorage-independent growth.
cJun is a major component of the transcription factor AP-1 and mediates a diverse set of biologic properties including proliferation, differentiation, and apoptosis. To identify cJun-responsive genes, we inducibly expressed cJun in Rat-1a cells and observed two distinct phenotypes: changes in cellular morphology with adherent growth and anchorage-independent growth. The biologic effects of cJun were entirely reversible demonstrating that they require the continued presence of cJun. To determine the genes, which mediate the biologic effects of cJun, we employed multiple methods including differential gene analysis, suppression subtractive hybridization, and cDNA microarrays. We identified 38 cJun-responsive genes including three uncharacterized genes under adherent and/or nonadherent conditions. Half of the known 36 genes were cytoskeleton- and adhesion-related genes, suggesting a major role of cJun in the regulation of the genes related to cell morphology. As proof of the principle that this approach could identify genes whose upregulation was necessary for nonadherent growth, we investigated one gene, stathmin whose upregulation by cJun was observed only under these conditions. Although overexpression of stathmin did not result in nonadherent growth, inhibition of stathmin protein expression by antisense oligonucleotides in cJun-induced Rat-1a cells prevented nonadherent growth. These results suggest that stathmin plays an essential role in anchorage-independent growth by cJun and may be a potential target for specific inhibitors for AP-1-dependent processes involved in carcinogenesis.
Cell and Cancer Biology Department, Center For Cancer Research, National Cancer Institute, Rockville, MD 20850, USA.
Ichiro
I
Kinoshita
Virna
V
Leaner
Motoo
M
Katabami
Ramon G
RG
Manzano
Paul
P
Dent
Anita
A
Sabichi
Michael J
MJ
Birrer
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002455:Cell Division; D002460:Cell Line; D017931:DNA Primers; D004317:Doxorubicin / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D017930:Genes, Reporter; D016758:Genes, jun; D049452:Green Fluorescent Proteins; D007700:Kinetics; D008156:Luciferases / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics; D008868:Microtubule Proteins; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010750:Phosphoproteins / Q000235:genetics*; D011401:Promoter Regions, Genetic; D016755:Proto-Oncogene Proteins c-jun / Q000235:genetics*; D051381:Rats; D011916:Rats, Inbred F344; D050777:Stathmin; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection
D017931:DNA Primers; D008164:Luminescent Proteins; D008868:Microtubule Proteins; D010750:Phosphoproteins; D016755:Proto-Oncogene Proteins c-jun; D050777:Stathmin; D014157:Transcription Factors; D049452:Green Fluorescent Proteins; D004317:Doxorubicin; D008156:Luciferases