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Related Experiment Videos

Nucleoprotein complexes in simian virus 40-infected cells.

M White, R Eason

    Journal of Virology
    |October 1, 1971
    PubMed
    Summary

    Newly synthesized simian virus 40 (SV40) deoxyribonucleic acid (DNA) in infected cells forms protein-associated nucleoprotein complexes. These complexes, not free DNA, are extracted using Triton X-100, indicating a crucial role for protein in SV40 DNA replication.

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    Area of Science:

    • Molecular Virology
    • Cell Biology
    • Biochemistry

    Background:

    • Simian virus 40 (SV40) is a DNA tumor virus extensively used to study viral replication and DNA metabolism.
    • Understanding the intracellular state of viral DNA during replication is crucial for deciphering viral life cycles.

    Purpose of the Study:

    • To investigate the physical state of newly synthesized SV40 deoxyribonucleic acid (DNA) within infected African green monkey kidney cells (BSC-1).
    • To determine if SV40 DNA exists as free molecules or is associated with proteins during replication.

    Main Methods:

    • Infection of BSC-1 cells with SV40 and pulse-labeling with (3)H-thymidine to label newly synthesized DNA.
    • Extraction of infected cells using Triton X-100 and analysis of SV40 DNA sedimentation coefficients in sucrose gradients.
    • Confirmation of protein association using Pronase digestion and (14)C-amino acid labeling.

    Main Results:

    • Newly synthesized (3)H-SV40 DNA exhibited a sedimentation coefficient of 44S, significantly different from purified SV40 DNA (21S).
    • This higher sedimentation coefficient was attributed to the association of SV40 DNA with proteins, confirmed by Pronase sensitivity and amino acid labeling.
    • Replicating SV40 DNA molecules (25S) were also found as nucleoprotein complexes in Triton extracts, quantitatively similar to deoxycholate extracts.

    Conclusions:

    • Newly synthesized SV40 DNA in infected cells exists predominantly as nucleoprotein complexes, not as free DNA.
    • These findings suggest that protein association is integral to the state of SV40 DNA during its replication cycle within the host cell.

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