Mouse mammary tumor virus DNA in infected rat cells: characterization of unintegrated forms
Abstract:
Mouse mammary tumor virus (MMTV) DNA in chronically infected rat hepatoma cells is maintained in both the integrated and unintegrated state. Fractionation of DNA by the procedure of Hirt (1967) as well as by sedimentation through alkaline sucrose suggests that about two thirds of the viral DNA is associated with high molecular weight cell DNA. The remainder of the viral DNA is unintegrated and is present primarily as linear or open circular duplexes consisting of a genome-length strand complementary to the viral RNA ("minus" strand) and "plus" strands of subgenomic length. Approximately 20% of the unintegrated MMTV DNA is present as double-stranded, covently closed circles (form I) with a molecular weight of 6 X 10(6) daltons. Form I viral DNA is found primarily in the nucleus, whereas the open forms are both nuclear and cytoplasmic.
Insights
Mouse mammary tumor virus (MMTV) DNA exists in both integrated and unintegrated forms within infected rat hepatoma cells. Most MMTV DNA associates with host cell DNA, while the rest exists as circular or linear forms, with some as closed circles in the nucleus.
Area of Science:
- Virology
- Molecular Biology
- Cancer Research
Background:
- Mouse mammary tumor virus (MMTV) is a retrovirus known to cause mammary tumors in mice.
- Understanding the state and location of viral DNA within host cells is crucial for comprehending viral replication and pathogenesis.
- Chronic infection of host cells can lead to complex interactions between viral and cellular genomes.
Purpose of the Study:
- To investigate the physical state and intracellular localization of Mouse mammary tumor virus (MMTV) DNA in chronically infected rat hepatoma cells.
- To characterize the forms of unintegrated MMTV DNA present in infected cells.
- To determine the association of MMTV DNA with host cell DNA.
Main Methods:
- DNA extraction using Hirt fractionation.
- DNA sedimentation analysis through alkaline sucrose gradients.
- Characterization of viral DNA forms (linear, open circular, covalently closed circular).
- Subcellular fractionation to determine nuclear and cytoplasmic localization.
Main Results:
- MMTV DNA exists in both integrated and unintegrated forms in infected rat hepatoma cells.
- Approximately two-thirds of MMTV DNA is associated with high molecular weight host cell DNA.
- The remaining MMTV DNA is unintegrated, primarily as linear or open circular duplexes.
- About 20% of unintegrated MMTV DNA exists as covalently closed circular molecules (Form I) with a molecular weight of 6 x 10^6 daltons.
- Form I MMTV DNA is predominantly nuclear, while open forms are found in both nuclear and cytoplasmic fractions.
Conclusions:
- Chronic MMTV infection in rat hepatoma cells results in a heterogeneous distribution of viral DNA.
- The association of MMTV DNA with host cell DNA suggests potential mechanisms for viral genome maintenance and replication.
- The presence of distinct forms of unintegrated MMTV DNA, particularly nuclear covalently closed circles, indicates specific molecular events occurring during chronic infection.
