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J Virol, July 1998, p. 5481-5492, Vol. 72, No. 7
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Phosphorylation of the Human Cytomegalovirus 86-Kilodalton Immediate-Early Protein IE2

Noam Y. Harel and James C. Alwine*

Graduate Group of Cell and Molecular Biology and Department of Microbiology, Cancer Center, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6142

Received 16 January 1998/Accepted 31 March 1998

We have investigated the phosphorylation state of the human cytomegalovirus 86-kDa immediate-early (IE) protein IEP86 from transfected and infected cells. We show that multiple domains of IEP86 are phosphorylated by cellular kinases, both in vitro and in vivo. Our data suggest that serum-inducible kinases play a significant role in cell-mediated IE protein phosphorylation and that a member of the mitogen-activated protein (MAP) kinase (MAPK) family, extracellular regulated kinase 2 (ERK2), phosphorylates several domains of IEP86 in vitro. Alanine substitution mutagenesis was performed on specific serines or threonines (T27, S144, T233/S234, and T555) found in consensus MAP kinase motifs. Analysis of these mutations showed that T27 and T233/S234 are the major sites for serum-inducible kinases and are the major ERK2 sites in vitro. S144 appeared to be phosphorylated in a serum-independent manner in vitro. All of the mutations except T555 eliminated specific phosphorylation in vivo. In transient transfection analyses, IEP86 isoforms containing mutations in S144 and, especially, T233/S234 displayed increased transcriptional activation relative to the wild type, suggesting that phosphorylation at these sites in wild-type IEP86 may result in reduction of its transcriptional activation ability.


* Corresponding author. Mailing address: 560 Clinical Research Building, 415 Curie Blvd., University of Pennsylvania, Philadelphia, PA 19104-6142. Phone: (215) 898-3256. Fax: (215) 573-3888. E-mail: alwine{at}mail.med.upenn.edu.


J Virol, July 1998, p. 5481-5492, Vol. 72, No. 7
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.



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Copyright © 1998 by the American Society for Microbiology. All rights reserved.