title: Loss of p19(ARF) eliminates the requirement for the pRB-binding motif in simian virus 40 large T antigen-mediated transformation.
abstract: At least three domains of simian virus 40 large T antigen (TAg) participate in cellular transformation. The LXCXE motif of TAg binds to all members of the retinoblastoma protein (pRB) family of tumor suppressors. The N-terminal 70 residues of TAg have significant homology to the J domain of Hsp40/DnaJ and cooperate with the LXCXE motif to inactivate the pRB family. A bipartite C-terminal domain of TAg binds to p53 and thereby disrupts the ability of p53 to act as a sequence-specific transcription factor. The contribution of these three domains of TAg to cellular transformation was evaluated in cells that contained inactivating mutations in the pRB and p53 pathways. Cells that stably expressed wild-type or selected mutant forms of TAg were generated in mouse embryo fibroblasts (MEFs) containing homozygous deletions in the RB, INK4a, and ARF loci. It was determined that the J domain, the LXCXE motif, and the p53-binding domain of TAg were required for full transformation of wild-type and RB(-/-) MEFs. In contrast, INK4a(-/-) MEFs that lacked expression of p16(INK4a) and p19(ARF) and ARF(-/-) MEFs that lacked p19(ARF) but expressed p16(INK4a) acquired anchorage-independent growth when expressing wild-type TAg or mutant derivatives that disrupted either the pRB-binding or p53-binding domain. The expression and function of the pRB family members were not overly disrupted in ARF(-/-) MEFs expressing LXCXE mutants of TAg. These results suggest that inactivating mutations of p19(ARF) can relieve the requirement for the LXCXE motif in TAg-mediated transformation and that TAg may have additional functions in transformation.
authors.affiliation: Dana-Farber Cancer Institute and Harvard Medical School, Boston, Massachusetts 02115, USA.
authors.forename: H H
authors.initials: HH
authors.lastname: Chao
authors.forename: A M
authors.initials: AM
authors.lastname: Buchmann
authors.forename: J A
authors.initials: JA
authors.lastname: DeCaprio
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D000957:Antigens, Viral, Tumor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D002452:Cell Count; D018797:Cell Cycle Proteins; D002455:Cell Division; D002461:Cell Line, Transformed; D002471:Cell Transformation, Neoplastic / Q000235:genetics*; D003260:Contact Inhibition; D019941:Cyclin-Dependent Kinase Inhibitor p16; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D005347:Fibroblasts; D017353:Gene Deletion; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D051379:Mice; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism*; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors / Q000378:metabolism; D025803:Tumor Suppressor Protein p14ARF; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
chemical_list: D000957:Antigens, Viral, Tumor; C533431:Arid4a protein, mouse; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D019941:Cyclin-Dependent Kinase Inhibitor p16; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D011506:Proteins; D016160:Retinoblastoma Protein; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors; D025803:Tumor Suppressor Protein p14ARF; D016159:Tumor Suppressor Protein p53