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DNA polymerase alpha from the nuclear matrix of cells infected with simian virus 40
Abstract:
The nuclear matrix prepared from normal, simian virus 40 (SV40)-infected, and SV40-transformed cells contained DNA polymerase activities. Approximately 12% of the total DNA polymerase activities in isolated nuclei remained with the nuclear matrix. alpha-polymerase was the major matrix DNA polymerase activity as judged by sensitivity to various inhibitors: aphidicolin, dideoxy-TTP, and N-ethylmaleimide. Approximately 2-4 fold higher DNA polymerase activity was detected in matrices obtained from lytically infected and virus-transformed cells than that found in normal cells. In lytically infected cells, 30-50% of the matrix-bound DNA polymerase activity solubilized by sonication co-sedimented with majority of the matrix T-antigen, and was co-precipitated with anti-T sera. The results suggest that alpha-polymerase and viral T-antigen may form a functional complex in the matrix.
Insights
Simian virus 40 infection and transformation increase nuclear matrix DNA polymerase activity, primarily alpha-polymerase. This enzyme may form a functional complex with viral T-antigen within the matrix.
Area of Science:
- Molecular Biology
- Virology
- Cell Biology
Background:
- The nuclear matrix plays a crucial role in DNA replication and organization.
- Simian virus 40 (SV40) infection and transformation alter cellular processes, including DNA metabolism.
Purpose of the Study:
- To investigate DNA polymerase activities associated with the nuclear matrix in normal, SV40-infected, and SV40-transformed cells.
- To determine the specific type of DNA polymerase present in the nuclear matrix and its potential interaction with viral proteins.
Main Methods:
- Preparation of nuclear matrices from different cell types.
- Assay of DNA polymerase activities and characterization using specific inhibitors (aphidicolin, dideoxy-TTP, N-ethylmaleimide).
- Analysis of matrix-bound DNA polymerase activity and T-antigen co-sedimentation and co-precipitation.
Main Results:
- Approximately 12% of total nuclear DNA polymerase activity was associated with the nuclear matrix.
- Alpha-polymerase was identified as the major DNA polymerase activity in the nuclear matrix.
- Matrices from SV40-infected and -transformed cells exhibited 2-4 fold higher DNA polymerase activity compared to normal cells.
- A significant portion of matrix-bound DNA polymerase activity co-sedimented and co-precipitated with SV40 T-antigen in infected cells.
Conclusions:
- The nuclear matrix harbors significant DNA polymerase activity, predominantly alpha-polymerase.
- SV40 infection and transformation lead to elevated nuclear matrix DNA polymerase activity.
- Evidence suggests a functional association between alpha-polymerase and SV40 T-antigen within the nuclear matrix, potentially implicating them in viral DNA replication.