Interaction of mouse adenovirus type 1 early region 1A protein with cellular proteins pRb and p107
| Autor: | Katherine R. Spindler, Clayton W. Beard, Kimberley Smith, Amy Oberhauser Ball, Baoling Ying |
|---|---|
| Rok vydání: | 1996 |
| Předmět: |
Immunoprecipitation
Mutant Cell Cell Cycle Proteins Retinoblastoma-Like Protein p107 DNA-binding protein Retinoblastoma Protein Cell Line Mastadenovirus Mice Virology medicine Tumor Cells Cultured Animals Humans E2F biology Retinoblastoma protein Nuclear Proteins Molecular biology E2F Transcription Factors DNA-Binding Proteins medicine.anatomical_structure Cell culture Mutagenesis biology.protein Adenovirus E1A Proteins biological phenomena cell phenomena and immunity Carrier Proteins Transcription Factor DP1 Retinoblastoma-Binding Protein 1 Transcription Factors |
| Zdroj: | Virology. 224(1) |
| ISSN: | 0042-6822 |
| Popis: | We demonstrated functional associations between mouse adenovirus type 1 (MAV-1) early region 1A (E1A) protein and both the mouse retinoblastoma protein (pRb) and the mouse pRb-related protein, p107. Interactions between MAV-1 E1A and mouse pRb or mouse p107 proteins were examined in infected cell lysates using a mouse embryonic fibroblast cell line infected with wild-type and mutant MAV-1 viruses. Using a polyclonal antibody to MAV-1 E1A, exogenously added mouse pRb or mouse p107 was coimmunoprecipitated from wild-type, dIE105 (CR1 delta)-, and dIE106 (CR3 delta)-infected cell lysates. No coimmunoprecipitation was seen with cell lysates from dIE102 (CR2 delta) or pmE109, a mutant virus that produces no detectable E1A protein due to an ATG to TTG point mutation in the initiator methionine. Introduction of mouse pRb into SAOS-2 cells resulted in a flat and enlarged cell phenotype, whereas cotransfection of mouse pRb and MAV-1 E1A resulted in a significant reduction of flat cells, presumably due to E1A binding pRb. CR1 delta and CR2 delta E1A proteins were less effective at reducing the number of flat, enlarged cells induced by pRb expression than were the CR3 delta or wild-type E1A proteins. The reduced ability of these mutants to inactivate pRb relative to wild-type E1A correlated with their reduced ability to bind pRb in the in vitro coimmunoprecipitation experiments. As a measure of p107/MAV-1 E1A complex formation in MAV-1-infected cells, we used mobility shift assays to examine cell extracts for the presence of p107-containing E2F protein-DNA complexes. Mock-, dIE102-, and pmE109-infected cell extracts formed a p107-containing complex, whereas wild-type-infected cell extracts did not. Thus the formation of a p107-E2F complex in wild-type- or these mutant-infected extracts inversely correlated with the presence of E1A-p107 complexes identified in the vitro coimmunoprecipitation experiments. This is consistent with E1A-p107 complexes forming in wild-type MAV-1-infected cells. |
| Databáze: | OpenAIRE |
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