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Journal of Virology, November 2009, p. 11421-11428, Vol. 83, No. 22
0022-538X/09/$08.00+0     doi:10.1128/JVI.00762-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

The Endoplasmic Reticulum Chaperone BiP/GRP78 Is Important in the Structure and Function of the Human Cytomegalovirus Assembly Compartment{triangledown}

Nicholas J. Buchkovich,2 Tobi G. Maguire,2 Adrienne W. Paton,1 James C. Paton,1 and James C. Alwine2*

School of Molecular and Biomedical Science, University of Adelaide, Adelaide, South Australia 5005, Australia,1 Department of Cancer Biology, Abramson Family Cancer Research Institute, Cell and Molecular Biology Graduate Group, School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 191042

Received 14 April 2009/ Accepted 28 August 2009

We previously demonstrated that the endoplasmic reticulum (ER) chaperone BiP functions in human cytomegalovirus (HCMV) assembly and egress. Here, we show that BiP localizes in two cytoplasmic structures in infected cells. Antibodies to the extreme C terminus, which includes BiP's KDEL ER localization sequence, detect BiP in regions of condensed ER near the periphery of the cell. Antibodies to the full length, N terminus, or larger portion of the C terminus detect BiP in the assembly compartment. This inability of C-terminal antibodies to detect BiP in the assembly compartment suggests that BiP's KDEL sequence is occluded in the assembly compartment. Depletion of BiP causes the condensed ER and assembly compartments to dissociate, indicating that BiP is important for their integrity. BiP and pp28 are in association in the assembly compartment, since antibodies that detect BiP in the assembly compartment coimmunoprecipitate pp28 and vice versa. In addition, BiP and pp28 copurify with other assembly compartment components on sucrose gradients. BiP also coimmunoprecipitates TRS1. Previous data show that cells infected with a TRS1-deficient virus have cytoplasmic and assembly compartment defects like those seen when BiP is depleted. We show that a fraction of TRS1 purifies with the assembly compartment. These findings suggest that BiP and TRS1 share a function in assembly compartment maintenance. In summary, BiP is diverted from the ER to associate with pp28 and TRS1, contributing to the integrity and function of the assembly compartment.

* Corresponding author. Mailing address: Department of Cancer Biology, 314 Biomedical Research Building, 421 Curie Blvd., School of Medicine, University of Pennsylvania, Philadelphia, PA 19104-6142. Phone: (610) 565-5067. Fax: (610) 573-3888. E-mail: alwine{at}mail.med.upenn.edu

{triangledown} Published ahead of print on 9 September 2009.

Journal of Virology, November 2009, p. 11421-11428, Vol. 83, No. 22
0022-538X/09/$08.00+0     doi:10.1128/JVI.00762-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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