Localization of varicella-zoster virus gene 21 protein in virus-infected cells in culture.
Although four varicella-zoster virus (VZV) genes have been shown to be transcribed in latently infected human ganglia, their role in the development and maintenance of latency is unknown. To study these VZV transcripts, we decided first to localize their expression products in productively infected cells. We began with VZV gene 21, whose open reading frame (ORF) is 3,113 bp. We cloned the 5' and 3' ends and the predicted antigenic segments of the ORF as 1292-, 1280-, and 880-bp DNA fragments, respectively, into the prokaryotic expression vector pGEX-2T. The three VZV 21 ORFs were expressed as approximately 75-, 73-, and 59-kDa glutathione S-transferase fusion proteins in Escherichia coli. To prepare polyclonal antibodies that would recognize all potential epitopes on the VZV gene 21 protein, rabbits were inoculated with a mixture of the three fusion proteins, and antisera were obtained and affinity purified. Immunohistochemical and immunoelectron microscopic analyses using these antibodies revealed VZV ORF 21 protein in both the nucleus and cytoplasm of VZV-infected cells. When these antibodies were applied to purified VZV nucleocapsids, intense staining was seen in their central cores.
Mahalingam
R
R
Departments of Neurology, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA. ravi.mahalingam@uchsc.edu
Lasher
R
R
Wellish
M
M
Cohrs
R J
RJ
Gilden
D H
DH
D000818:Animals; D000914:Antibodies, Viral / Q000096:biosynthesis; D015153:Blotting, Western; D002460:Cell Line; D002522:Chlorocebus aethiops; D004926:Escherichia coli / Q000378:metabolism; D015870:Gene Expression; D005814:Genes, Viral; D005982:Glutathione Transferase; D014645:Herpesvirus 3, Human / Q000235:genetics*; D006801:Humans; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D014764:Viral Proteins
D000914:Antibodies, Viral; C113455:ORF 21 protein, Varicella-Zoster virus; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D005982:Glutathione Transferase