E5a gene of human papillomavirus type 11 is required for initiation but not for maintenance of transformation in NIH 3T3 cells.
We have previously shown that the E5a gene of human papilloma virus type 11 (HPV-11/HPV-6c) is a transforming oncogene. In order to dissect the biological consequences of E5a gene expression we utilized the lac operator/repressor system to manipulate E5a gene expression. Cells were cotransfected with the lac repressor gene and the E5a gene that had been inserted downstream of a simian virus 40 (SV40) promoter containing the lac operator sequence. The expression of E5a gene could therefore be repressed by binding of lac repressor to the lac operator sequence in proximity to this SV40 regulatory region. The transfected cells were cultured in the presence of the inducer IPTG and under G418 selection. IPTG derepressed E5a gene expression by binding to the repressor and reducing its affinity for the lac operator sequence. In these studies, we found that E5a-transformed cells still maintained the transformed phenotype as judged by growth density, cell morphology and anchorage-independent growth when E5a gene expression was repressed. We also found that c-jun expression was induced 3 h after E5a expression was induced by IPTG and c-jun expression was not shut down after repression of E5a expression. This is the first demonstration that the E5a gene of HPV-11 initiates transformation of NIH 3T3 cells but is dispensable for maintenance of the transformed phenotype.
Chen
S L
SL
Department of Microbiology and Immunology, National Defense Medical Center, Taipei, Taiwan, Republic of China.
Tsao
Y P
YP
Lee
J W
JW
Liu
H S
HS
Yang
C M
CM
Tsao
L T
LT
D016475:3T3 Cells; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D002472:Cell Transformation, Viral; D029968:Escherichia coli Proteins; D005814:Genes, Viral / Q000235:genetics*; D007544:Isopropyl Thiogalactoside; D056968:Lac Repressors; D051379:Mice; D009856:Oncogene Proteins, Viral / Q000096:biosynthesis* / Q000235:genetics; D009857:Oncogenes / Q000235:genetics*; D027383:Papillomaviridae / Q000235:genetics*; D010641:Phenotype; D016755:Proto-Oncogene Proteins c-jun / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D011994:Recombinant Proteins; D012097:Repressor Proteins / Q000235:genetics; D014162:Transfection
D001426:Bacterial Proteins; C081917:E5A protein, Human papillomavirus type 11; D029968:Escherichia coli Proteins; D056968:Lac Repressors; D009856:Oncogene Proteins, Viral; D016755:Proto-Oncogene Proteins c-jun; D012333:RNA, Messenger; D011994:Recombinant Proteins; D012097:Repressor Proteins; D007544:Isopropyl Thiogalactoside