Characterization of the interactions of human papillomavirus type 16 E6 with p53 and E6-associated protein in insect and human cells.
Human papillomavirus (HPV) 16 E6 induces the degradation of the tumour suppressor protein p53 by the ubiquitin-dependent proteolysis pathway. In vitro, this process involves the formation of a trimolecular complex between E6, p53 and a cellular protein E6-associated protein (E6-AP). However, an analysis of their potential interactions in vivo has not been carried out. We have established a model for the expression and analysis of the interactions of these three proteins in insect cells, a eukaryotic system where potentially crucial modifications of the proteins will occur. In baculovirus-infected cells the degradation of p53 can occur. However, p53 is only degraded early in the infectious cycle due to a lack of ATP at later times. Consequently, substantial quantities of material can be produced in this system for further analysis. Evidence is also provided that, in vivo, E6 can interact with p53 in the absence of E6-AP and that E6-AP can interact with p53 in the absence of E6. Furthermore, analysis of the subcellular localization of the proteins using both biochemical fractionation and indirect immunofluorescence suggests that the degradation of p53 occurs in the perinuclear region of the cell.
Daniels
P R
PR
Department of Biology, University of York, UK.
Sanders
C M
CM
Maitland
N J
NJ
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D002478:Cells, Cultured; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D007313:Insecta / Q000378:metabolism; D008025:Ligases / Q000378:metabolism*; D009856:Oncogene Proteins, Viral / Q000378:metabolism*; D027383:Papillomaviridae / Q000378:metabolism; D012097:Repressor Proteins; D013347:Subcellular Fractions / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases
C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D012097:Repressor Proteins; D016159:Tumor Suppressor Protein p53; C471639:UBE3A protein, human; D044767:Ubiquitin-Protein Ligases; D008025:Ligases