Molecular cloning and characterization of endogenous SV40 DNA from human HBL-100 cells.
The human HBL-100 cell line harbours SV40 DNA integrated in tandem at a unique site. The SV40 T-antigen expressed in these cells is defective in a function essential to the replication of the viral genome. The integrated SV40 sequences were molecularly cloned in a bacteriophage, and a subclone (plasmid pSVHBI) containing a complete SV40 DNA was isolated. As compared to SV40 wild-type strain 776, sequence analysis of pSVHBI early region revealed the presence of several DNA alterations. Among these, a point mutation at position 3199, predicting a change at amino-acid 540 of arginine to isoleucine, was shown by marker rescue to be responsible for the deficiency of T-antigen. This novel mutation further delimits one of the T-antigen domains involved in SV40 DNA replication. Transfection experiments demonstrated that the transforming activity of the SV40 genome from HBL-100 cells is still preserved. Moreover, several transformed human cell clones thus obtained could be permanently established in culture.
Saint-Ruf
C
C
UPR6, Institut de Recherches Scientifiques sur le Cancer, Villejuif, France.
Nardeux
P
P
Cebrian
J
J
Lacasa
M
M
Lavialle
C
C
Cassingena
R
R
D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D004261:DNA Replication; D004279:DNA, Viral / Q000032:analysis*; D006801:Humans; D009154:Mutation; D013539:Simian virus 40 / Q000235:genetics*; D014162:Transfection
D004279:DNA, Viral