Full-length isoforms of Kaposi's sarcoma-associated herpesvirus latency-associated nuclear antigen accumulate in the cytoplasm of cells undergoing the lytic cycle of replication

H. Jacques Garrigues, Kellie Howard, Serge Barcy, Minako Ikoma, Ashlee Moses, Gail H. Deutsch, David Wu, Keiji Ueda, Timothy M. Rose

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

The latency-associated nuclear antigen (LANA) of the Kaposi's sarcomaassociated herpesvirus (KSHV) performs a variety of functions to establish and maintain KSHV latency. During latency, LANA localizes to discrete punctate spots in the nucleus, where it tethers viral episomes to cellular chromatin and interacts with nuclear components to regulate cellular and viral gene expression. Using highly sensitive tyramide signal amplification, we determined that LANA localizes to the cytoplasm in different cell types undergoing the lytic cycle of replication after de novo primary infection and after spontaneous, tetradecanoyl phorbol acetate-, or open reading frame 50 (ORF50)/replication transactivator (RTA)-induced activation. We confirmed the presence of cytoplasmic LANA in a subset of cells in lytically active multicentric Castleman disease lesions. The induction of cellular migration by scratch-wounding confluent cell cultures, culturing under subconfluent conditions, or induction of cell differentiation in primary cultures upregulated the number of cells permissive for primary lytic KSHV infection. The induction of lytic replication was characterized by high-level expression of cytoplasmic LANA and nuclear ORF59, a marker of lytic replication. Subcellular fractionation studies revealed the presence of multiple isoforms of LANA in the cytoplasm of ORF50/RTA-activated Vero cells undergoing primary infection. Mass spectrometry analysis demonstrated that cytoplasmic LANA isoforms were full length, containing the N-terminal nuclear localization signal. These results suggest that trafficking of LANA to different subcellular locations is a regulated phenomenon, which allows LANA to interact with cellular components in different compartments during both the latent and the replicative stages of the KSHV life cycle.

Original languageEnglish (US)
Article numbere01532-17
JournalJournal of Virology
Volume91
Issue number24
DOIs
StatePublished - Dec 1 2017

Fingerprint

Human herpesvirus 8
nuclear antigens
Human Herpesvirus 8
Protein Isoforms
Cytoplasm
cytoplasm
Herpesviridae
cells
Trans-Activators
Open Reading Frames
open reading frames
latency-associated nuclear antigen
Herpesviridae Infections
Nuclear Localization Signals
nuclear localization signals
Vero Cells
Viral Genes
Infection
Life Cycle Stages
cell movement

Keywords

  • Activation
  • Cytoplasm
  • KSHV
  • LANA
  • Mass spectrometry
  • Migration
  • Viral replication

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Insect Science
  • Virology

Cite this

Full-length isoforms of Kaposi's sarcoma-associated herpesvirus latency-associated nuclear antigen accumulate in the cytoplasm of cells undergoing the lytic cycle of replication. / Garrigues, H. Jacques; Howard, Kellie; Barcy, Serge; Ikoma, Minako; Moses, Ashlee; Deutsch, Gail H.; Wu, David; Ueda, Keiji; Rose, Timothy M.

In: Journal of Virology, Vol. 91, No. 24, e01532-17, 01.12.2017.

Research output: Contribution to journalArticle

Garrigues, H. Jacques ; Howard, Kellie ; Barcy, Serge ; Ikoma, Minako ; Moses, Ashlee ; Deutsch, Gail H. ; Wu, David ; Ueda, Keiji ; Rose, Timothy M. / Full-length isoforms of Kaposi's sarcoma-associated herpesvirus latency-associated nuclear antigen accumulate in the cytoplasm of cells undergoing the lytic cycle of replication. In: Journal of Virology. 2017 ; Vol. 91, No. 24.
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AU - Garrigues, H. Jacques

AU - Howard, Kellie

AU - Barcy, Serge

AU - Ikoma, Minako

AU - Moses, Ashlee

AU - Deutsch, Gail H.

AU - Wu, David

AU - Ueda, Keiji

AU - Rose, Timothy M.

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AB - The latency-associated nuclear antigen (LANA) of the Kaposi's sarcomaassociated herpesvirus (KSHV) performs a variety of functions to establish and maintain KSHV latency. During latency, LANA localizes to discrete punctate spots in the nucleus, where it tethers viral episomes to cellular chromatin and interacts with nuclear components to regulate cellular and viral gene expression. Using highly sensitive tyramide signal amplification, we determined that LANA localizes to the cytoplasm in different cell types undergoing the lytic cycle of replication after de novo primary infection and after spontaneous, tetradecanoyl phorbol acetate-, or open reading frame 50 (ORF50)/replication transactivator (RTA)-induced activation. We confirmed the presence of cytoplasmic LANA in a subset of cells in lytically active multicentric Castleman disease lesions. The induction of cellular migration by scratch-wounding confluent cell cultures, culturing under subconfluent conditions, or induction of cell differentiation in primary cultures upregulated the number of cells permissive for primary lytic KSHV infection. The induction of lytic replication was characterized by high-level expression of cytoplasmic LANA and nuclear ORF59, a marker of lytic replication. Subcellular fractionation studies revealed the presence of multiple isoforms of LANA in the cytoplasm of ORF50/RTA-activated Vero cells undergoing primary infection. Mass spectrometry analysis demonstrated that cytoplasmic LANA isoforms were full length, containing the N-terminal nuclear localization signal. These results suggest that trafficking of LANA to different subcellular locations is a regulated phenomenon, which allows LANA to interact with cellular components in different compartments during both the latent and the replicative stages of the KSHV life cycle.

KW - Activation

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KW - LANA

KW - Mass spectrometry

KW - Migration

KW - Viral replication

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