Details

Autor(en) / Beteiligte

• PEARSON-WHITE, SONIA HELEN

Titel

MUTATIONAL ANALYSIS OF THE SIMIAN VIRUS 40 LARGE AND SMALL T ANTIGEN GENES

Ort / Verlag

ProQuest Dissertations & Theses

Erscheinungsjahr

1984

Quelle

ProQuest Dissertations & Theses A&I

Beschreibungen/Notizen

• The SV40 large and small T antigen genes were subjected to point and insertion mutagenesis at pre-selected target sites within the genes. Target sites included the coding region common to both large and small T antigens, the small t antigen unique coding region, splice junctions for large and small t antigens, and a region of large T with a putative involvement in DNA replication function and origin-binding. Four mutants with changes in the small t antigen gene or donor splice junction had a partial defect in DNA replication; in two of these mutants DNA replication was reduced by about an order of magnitude. One of the small t antigen mutants most defective for DNA replication, bc1158, has a single G to A transition at the donor splice junction, that does not alter coding sequence. This mutant produced no small t messenger RNA, demonstrating the significance of the G residue at the donor splice junction edge. A second mutant, bc1157, carries a C to G transversion located 7 base pairs upstream from the normal donor splice junction. This creates a sequence homologous to the normal splice junction sequence for a stretch of 5 base pairs, which is utilized as an alternate splice junction, again demonstrating the importance of the local sequence at splice junctions. An insertion mutant, in1153, with a single isoleucine codon inserted into the putative replication region of the large T antigen gene, lacked replication activity. However, the ability to transform non-permissive cells was retained, indicating that the replication and transforming functions are separable. Partially purified in1153 T antigen retained some ability to bind specifically to the origin of DNA replication, but the activity was reduced relative to wild type T antigen by about ten-fold.