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Vol. 10, Issue 6, 1705-1717, June 1999
Department of Biochemistry and Molecular Biophysics, Howard Hughes
Medical Institute, Columbia University, College of Physicians and
Surgeons, New York New York 10032
To identify cellular functions involved in the early phase of the
retroviral life cycle, somatic cell mutants were isolated after
selection for resistance to infection. Rat2 fibroblasts were treated
with chemical mutagens, and individual virus-resistant clones were
recovered after selection for resistance to infection. Two clones were
characterized in detail. Both mutant lines were resistant to infection
by both ecotropic and amphotropic murine viruses, as well as by human
immunodeficiency virus type 1 pseudotypes. One clone showed a strong
block to reverse transcription of the retroviral RNA, including
formation of the earliest DNA products. The second clone showed normal
levels of viral DNA synthesis but did not allow formation of the
circular DNAs normally found in the nucleus. Cell fractionation showed
that the viral preintegration complex was present in a form that could
not be extracted under conditions that readily extracted the complex
from wild-type cells. The results suggest that the DNA was trapped in a
nonproductive state and excluded from the nucleus of the infected cell.
The properties of these two mutant lines suggest that host gene
products play important roles both before and after reverse transcription.
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