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Journal of Virology, November 1999, p. 9555-9567, Vol. 73, No. 11
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Markers for trans-Golgi Membranes and the Intermediate Compartment Localize to Induced Membranes with Distinct Replication Functions in Flavivirus-Infected Cellsdagger

Jason M. Mackenzie,1,* Malcolm K. Jones,2 and Edwin G. Westaway1

Sir Albert Sakzewski Virus Research Centre, Royal Children's Hospital, Herston, Brisbane, Australia 4029,1 and Centre for Microscopy and Microanalysis, University of Queensland, St. Lucia, Brisbane, Australia 40722

Received 19 April 1999/Accepted 23 July 1999

Replication of the flavivirus Kunjin virus is associated with virus-induced membrane structures within the cytoplasm of infected cells; these membranes appear as packets of vesicles associated with the sites of viral RNA synthesis and as convoluted membranes (CM) and paracrystalline arrays (PC) containing the components of the virus-specified protease (E. G. Westaway, J. M. Mackenzie, M. T. Kenney, M. K. Jones, and A. A. Khromykh, J. Virol. 71:6650-6661, 1997). To determine the cellular origins of these membrane structures, we compared the immunolabelling patterns of several cell markers in relation to these sites by immunofluorescence and immunoelectron microscopy. A marker for the trans-Golgi membranes and the trans-Golgi network, 1,4-galactosyltransferase (GalT), was redistributed to large foci in the cytoplasm of Kunjin virus-infected cells, partially coincident with immunofluorescent foci associated with the putative sites of viral RNA synthesis. As determined by immunoelectron microscopy, the induced vesicle packets contained GalT, whereas the CM and PC contained a specific protein marker for the intermediate compartment (ERGIC53). A further indicator of the role of cellular organelles in their biogenesis was the observation that the Golgi apparatus-disrupting agent brefeldin A prevented further development of immunofluorescent foci of induced membranes if added before the end of the latent period but that once formed, these membrane foci were resistant to brefeldin A dispersion. Reticulum membranes emanating from the induced CM and PC were also labelled with the rough endoplasmic reticulum marker anti-protein disulfide isomerase and were obviously redistributed during infection. This is the first report identifying trans-Golgi membranes and the intermediate compartment as the apparent sources of the flavivirus-induced membranes involved in events of replication.

* Corresponding author. Mailing address: Sir Albert Sakzewski Virus Research Centre, Royal Children's Hospital, Herston, Brisbane, Australia 4029. Phone: 61 7 3253 1569. Fax: 61 7 3253 1401. E-mail: mackenzi{at}biosci.uq.oz.au.

dagger SASVRC publication 100.

Journal of Virology, November 1999, p. 9555-9567, Vol. 73, No. 11
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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