Functional imaging of viral transcription factories using 3D fluorescence microscopy

Christopher P. Chen, Frank Chuang, Yoshihiro Izumiya

Research output: Contribution to journalArticle

2 Scopus citations

Abstract

It is well known that spatial and temporal regulation of genes is an integral part of governing proper gene expression. Consequently, it is invaluable to understand where and when transcription is taking place within nuclear space and to visualize the relationship between episomes infected within the same cell's nucleus. Here, both immunofluorescence (IFA) and RNA-FISH have been combinedto identify actively transcribing Kaposi’s sarcoma-associated herpesvirus (KSHV) episomes. By staining KSHV latency-associated nuclear antigen (LANA), it is possible to locate where viral episomes exist within the nucleus. In addition, by designing RNA-FISH probes to target the intron region of a viral gene, which is expressed only during productive infection, nascent RNA transcripts can be located. Using this combination of molecular probes, it is possible to visualize the assembly of large viral transcription factories and analyze the spatial regulation of viral gene expression during KSHV reactivation. By including anti-RNA polymerase II antibody staining, one can also visualize the association between RNA polymerase II (RNAPII) aggregation and KSHV transcription during reactivation.

Original languageEnglish (US)
Article numbere56832
JournalJournal of Visualized Experiments
Volume2018
Issue number131
DOIs
StatePublished - Jan 18 2018

Keywords

  • Active transcription
  • Issue 131
  • Kaposi’s sarcoma-associated herpesvirus
  • KSHV
  • Microbiology
  • RNA polymerase II
  • RNA-FISH
  • Transcription
  • Viral transcription factory

ASJC Scopus subject areas

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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