This Article
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Castellino, A. M.
Right arrow Articles by Pipas, J. M.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Castellino, A. M.
Right arrow Articles by Pipas, J. M.

 Previous Article  |  Next Article 

J. Virol., 10 1997, 7549-7559, Vol 71, No. 10
Copyright © 1997, American Society for Microbiology

trans-Dominant and non-trans-dominant mutant simian virus 40 large T antigens show distinct responses to ATP

AM Castellino, P Cantalupo, IM Marks, JV Vartikar, KW Peden and JM Pipas
Department of Biological Sciences, University of Pittsburgh, Pennsylvania 15260, USA.

Simian virus 40 (SV40) DNA replication requires the coordinated action of multiple biochemical activities intrinsic to the virus-encoded large tumor antigen (T antigen). We report the preliminary biochemical characterization of the T antigens encoded by three SV40 mutants, 5030, 5031, and 5061, each of which have altered residues within or near the ATP binding pocket. All three mutants are defective for viral DNA replication in cultured cell lines. However, while 5030 and 5031 can be complemented in vivo by providing a wild-type T antigen in trans, 5061 exhibits a strong trans-dominant-negative phenotype. In order to determine the basis for their replication defects and to explore the mechanisms of trans dominance, we purified the T antigens encoded by each of these mutants and examined their activities in vitro. The 5061 T antigen had no measurable ATPase activity and failed to hexamerize in response to ATP, and its affinity for the SV40 origin of DNA replication (ori) DNA was not increased by ATP. In contrast, the 5030 and 5031 T antigens exhibited at least some ATPase activity and both readily formed hexamers in the presence of ATP. These mutants differed in that 5030 was very defective in an ori-dependent unwinding assay while 5031 retained significant activity. Both the 5030 and 5031 T antigens bound to ori-containing DNA, but the binding was less efficient than that of wild-type T antigen and was not affected by the presence of ATP. These results suggest that 5030 and 5031 are defective in some aspect of communication between the ATP binding and DNA binding domains and that the ability of ATP to induce T-antigen hexamerization is distinct from its action to increase the affinity for ori. Finally, all three mutants were defective for the ability to support SV40 DNA replication in vitro. Both the 5031 and 5061 T antigens inhibited wild- type-T-antigen-stimulated replication in vitro, while the 5030 T antigen did not. The fact that the 5031 T antigen was trans dominant in the in vitro assays but not in vivo indicates that the in vitro system does not accurately reflect events occurring in vivo.


This article has been cited by other articles:

  • Kumar, A., Meinke, G., Reese, D. K., Moine, S., Phelan, P. J., Fradet-Turcotte, A., Archambault, J., Bohm, A., Bullock, P. A. (2007). Model for T-Antigen-Dependent Melting of the Simian Virus 40 Core Origin Based on Studies of the Interaction of the Beta-Hairpin with DNA. J. Virol. 81: 4808-4818 [Abstract] [Full Text]  
  • Fewell, S. W., Smith, C. M., Lyon, M. A., Dumitrescu, T. P., Wipf, P., Day, B. W., Brodsky, J. L. (2004). Small Molecule Modulators of Endogenous and Co-chaperone-stimulated Hsp70 ATPase Activity. J. Biol. Chem. 279: 51131-51140 [Abstract] [Full Text]  
  • Zhang, Y.-H., Kooistra, K., Pietersen, A., Rohn, J. L., Noteborn, M. H. M. (2004). Activation of the Tumor-Specific Death Effector Apoptin and Its Kinase by an N-Terminal Determinant of Simian Virus 40 Large T Antigen. J. Virol. 78: 9965-9976 [Abstract] [Full Text]  
  • Ott, R. D., Wang, Y., Fanning, E. (2002). Mutational Analysis of Simian Virus 40 T-Antigen Primosome Activities in Viral DNA Replication. J. Virol. 76: 5121-5130 [Abstract] [Full Text]  
  • Sullivan, C. S., Gilbert, S. P., Pipas, J. M. (2001). ATP-Dependent Simian Virus 40 T-Antigen-Hsc70 Complex Formation. J. Virol. 75: 1601-1610 [Abstract] [Full Text]  
  • Sullivan, C. S., Cantalupo, P., Pipas, J. M. (2000). The Molecular Chaperone Activity of Simian Virus 40 Large T Antigen Is Required To Disrupt Rb-E2F Family Complexes by an ATP-Dependent Mechanism. Mol. Cell. Biol. 20: 6233-6243 [Abstract] [Full Text]  
  • Timchenko, T., de Kouchkovsky, F., Katul, L., David, C., Vetten, H. J., Gronenborn, B. (1999). A Single Rep Protein Initiates Replication of Multiple Genome Components of Faba Bean Necrotic Yellows Virus, a Single-Stranded DNA Virus of Plants. J. Virol. 73: 10173-10182 [Abstract] [Full Text]  
  • Pfister, T., Wimmer, E. (1999). Characterization of the Nucleoside Triphosphatase Activity of Poliovirus Protein 2C Reveals a Mechanism by Which Guanidine Inhibits Poliovirus Replication. J. Biol. Chem. 274: 6992-7001 [Abstract] [Full Text]  
  • Higashino, F., Pipas, J. M., Shenk, T. (1998). Adenovirus E4orf6 oncoprotein modulates the function of the p53-related protein, p73. Proc. Natl. Acad. Sci. USA 95: 15683-15687 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»