Proteomics analysis of the nucleolus in adenovirus-infected cells.
Adenoviruses replicate primarily in the host cell nucleus, and it is well established that adenovirus infection affects the structure and function of host cell nucleoli in addition to coding for a number of nucleolar targeted viral proteins. Here we used unbiased proteomics methods, including high throughput mass spectrometry coupled with stable isotope labeling by amino acids in cell culture (SILAC) and traditional two-dimensional gel electrophoresis, to identify quantitative changes in the protein composition of the nucleolus during adenovirus infection. Two-dimensional gel analysis revealed changes in six proteins. By contrast, SILAC-based approaches identified 351 proteins with 24 proteins showing at least a 2-fold change after infection. Of those, four were previously reported to have aberrant localization and/or functional relevance during adenovirus infection. In total, 15 proteins identified as changing in amount by proteomics methods were examined in infected cells using confocal microscopy. Eleven of these proteins showed altered patterns of localization in adenovirus-infected cells. Comparing our data with the effects of actinomycin D on the nucleolar proteome revealed that adenovirus infection apparently specifically targets a relatively small subset of nucleolar antigens at the time point examined.
Lam
Yun W
YW
Department of Biology and Chemistry, City University of Hong Kong, 83 Tat Chee Avenue, Kowloon Tong, Hong Kong, China.
Evans
Vanessa C
VC
Heesom
Kate J
KJ
Lamond
Angus I
AI
Matthews
David A
DA
D000256:Adenoviridae / Q000254:growth & development* / Q000502:physiology; D002466:Cell Nucleolus / Q000187:drug effects / Q000378:metabolism* / Q000821:virology; D003609:Dactinomycin / Q000494:pharmacology; D015180:Electrophoresis, Gel, Two-Dimensional; D006367:HeLa Cells; D054884:Host-Pathogen Interactions; D006801:Humans; D007553:Isotope Labeling / Q000379:methods; D013058:Mass Spectrometry; D009687:Nuclear Proteins / Q000032:analysis*; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D020543:Proteome / Q000032:analysis; D040901:Proteomics / Q000379:methods*
D009687:Nuclear Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D020543:Proteome; D003609:Dactinomycin