The human cytomegalovirus IE86 protein can block cell cycle progression after inducing transition into the S phase of permissive cells

Eain A. Murphy, Daniel Streblow, Jay Nelson, Mark F. Stinski

Research output: Contribution to journalArticle

111 Citations (Scopus)

Abstract

Human cytomegalovirus (HCMV) infection of permissive cells has been reported to induce a cell cycle halt. One or more viral proteins may be involved in halting progression at different stages of the cell cycle. We investigated how HCMV infection, and specifically IE86 protein expression, affects the cell cycles of permissive and nonpermissive cells. We used a recombinant virus that expresses the green fluorescent protein (GFP) to determine the effects of HCMV on the cell cycle of permissive cells. Fluorescence by GFP allowed us to select for only productively infected cells. Replication-defective adenovirus vectors expressing the IE72 or IE86 protein were also used to efficiently transduce 95% or more of the cells. The adenovirus-expressed IE86 protein was determined to be functional by demonstrating negative autoregulation of the major immediate-early promoter and activation of an early vital promoter in the context of the vital genome. To eliminate adenovirus protein effects, plasmids expressing GFP for fluorescent selection of only transfected cells and wild-type IE86 protein or a mutant IE86 protein were tested in permissive and nonpermissive cells. HCMV infection induced the entry of U373 cells into the S phase. All permissive cells infected with HCMV were blocked in cell cycle progression and could not divide. After either transduction or transfection and IE86 protein expression, the number of all permissive or nonpermissive cell types in the S phase increased significantly, but the cells could no longer divide. The IE72 protein did not have a significant effect on the S phase. Since IE86 protein inhibits cell cycle progression, the IE2 gene in a human fibroblast IE86 protein-expressing cell line was sequenced. The IE86 protein in these retrovirus-transduced cells has mutations in a critical region of the vital protein. The locations of the mutations and the function of the IE86 protein in controlling cell cycle progression are discussed.

Original languageEnglish (US)
Pages (from-to)7108-7118
Number of pages11
JournalJournal of Virology
Volume74
Issue number15
DOIs
StatePublished - Aug 2000

Fingerprint

Human herpesvirus 5
S Phase
interphase
cell cycle
Cell Cycle
proteins
cells
Proteins
Cytomegalovirus Infections
Adenoviridae
Green Fluorescent Proteins
green fluorescent protein
Cytomegalovirus
Cytomegalovirus IE2 protein
Retroviridae Proteins
Retroviridae
protein synthesis
promoter regions
mutation
autoregulation

ASJC Scopus subject areas

  • Immunology

Cite this

The human cytomegalovirus IE86 protein can block cell cycle progression after inducing transition into the S phase of permissive cells. / Murphy, Eain A.; Streblow, Daniel; Nelson, Jay; Stinski, Mark F.

In: Journal of Virology, Vol. 74, No. 15, 08.2000, p. 7108-7118.

Research output: Contribution to journalArticle

@article{e7d17f389e29442d86b95ec5bfe1fe93,
title = "The human cytomegalovirus IE86 protein can block cell cycle progression after inducing transition into the S phase of permissive cells",
abstract = "Human cytomegalovirus (HCMV) infection of permissive cells has been reported to induce a cell cycle halt. One or more viral proteins may be involved in halting progression at different stages of the cell cycle. We investigated how HCMV infection, and specifically IE86 protein expression, affects the cell cycles of permissive and nonpermissive cells. We used a recombinant virus that expresses the green fluorescent protein (GFP) to determine the effects of HCMV on the cell cycle of permissive cells. Fluorescence by GFP allowed us to select for only productively infected cells. Replication-defective adenovirus vectors expressing the IE72 or IE86 protein were also used to efficiently transduce 95{\%} or more of the cells. The adenovirus-expressed IE86 protein was determined to be functional by demonstrating negative autoregulation of the major immediate-early promoter and activation of an early vital promoter in the context of the vital genome. To eliminate adenovirus protein effects, plasmids expressing GFP for fluorescent selection of only transfected cells and wild-type IE86 protein or a mutant IE86 protein were tested in permissive and nonpermissive cells. HCMV infection induced the entry of U373 cells into the S phase. All permissive cells infected with HCMV were blocked in cell cycle progression and could not divide. After either transduction or transfection and IE86 protein expression, the number of all permissive or nonpermissive cell types in the S phase increased significantly, but the cells could no longer divide. The IE72 protein did not have a significant effect on the S phase. Since IE86 protein inhibits cell cycle progression, the IE2 gene in a human fibroblast IE86 protein-expressing cell line was sequenced. The IE86 protein in these retrovirus-transduced cells has mutations in a critical region of the vital protein. The locations of the mutations and the function of the IE86 protein in controlling cell cycle progression are discussed.",
author = "Murphy, {Eain A.} and Daniel Streblow and Jay Nelson and Stinski, {Mark F.}",
year = "2000",
month = "8",
doi = "10.1128/JVI.74.15.7108-7118.2000",
language = "English (US)",
volume = "74",
pages = "7108--7118",
journal = "Journal of Virology",
issn = "0022-538X",
publisher = "American Society for Microbiology",
number = "15",

}

TY - JOUR

T1 - The human cytomegalovirus IE86 protein can block cell cycle progression after inducing transition into the S phase of permissive cells

AU - Murphy, Eain A.

AU - Streblow, Daniel

AU - Nelson, Jay

AU - Stinski, Mark F.

PY - 2000/8

Y1 - 2000/8

N2 - Human cytomegalovirus (HCMV) infection of permissive cells has been reported to induce a cell cycle halt. One or more viral proteins may be involved in halting progression at different stages of the cell cycle. We investigated how HCMV infection, and specifically IE86 protein expression, affects the cell cycles of permissive and nonpermissive cells. We used a recombinant virus that expresses the green fluorescent protein (GFP) to determine the effects of HCMV on the cell cycle of permissive cells. Fluorescence by GFP allowed us to select for only productively infected cells. Replication-defective adenovirus vectors expressing the IE72 or IE86 protein were also used to efficiently transduce 95% or more of the cells. The adenovirus-expressed IE86 protein was determined to be functional by demonstrating negative autoregulation of the major immediate-early promoter and activation of an early vital promoter in the context of the vital genome. To eliminate adenovirus protein effects, plasmids expressing GFP for fluorescent selection of only transfected cells and wild-type IE86 protein or a mutant IE86 protein were tested in permissive and nonpermissive cells. HCMV infection induced the entry of U373 cells into the S phase. All permissive cells infected with HCMV were blocked in cell cycle progression and could not divide. After either transduction or transfection and IE86 protein expression, the number of all permissive or nonpermissive cell types in the S phase increased significantly, but the cells could no longer divide. The IE72 protein did not have a significant effect on the S phase. Since IE86 protein inhibits cell cycle progression, the IE2 gene in a human fibroblast IE86 protein-expressing cell line was sequenced. The IE86 protein in these retrovirus-transduced cells has mutations in a critical region of the vital protein. The locations of the mutations and the function of the IE86 protein in controlling cell cycle progression are discussed.

AB - Human cytomegalovirus (HCMV) infection of permissive cells has been reported to induce a cell cycle halt. One or more viral proteins may be involved in halting progression at different stages of the cell cycle. We investigated how HCMV infection, and specifically IE86 protein expression, affects the cell cycles of permissive and nonpermissive cells. We used a recombinant virus that expresses the green fluorescent protein (GFP) to determine the effects of HCMV on the cell cycle of permissive cells. Fluorescence by GFP allowed us to select for only productively infected cells. Replication-defective adenovirus vectors expressing the IE72 or IE86 protein were also used to efficiently transduce 95% or more of the cells. The adenovirus-expressed IE86 protein was determined to be functional by demonstrating negative autoregulation of the major immediate-early promoter and activation of an early vital promoter in the context of the vital genome. To eliminate adenovirus protein effects, plasmids expressing GFP for fluorescent selection of only transfected cells and wild-type IE86 protein or a mutant IE86 protein were tested in permissive and nonpermissive cells. HCMV infection induced the entry of U373 cells into the S phase. All permissive cells infected with HCMV were blocked in cell cycle progression and could not divide. After either transduction or transfection and IE86 protein expression, the number of all permissive or nonpermissive cell types in the S phase increased significantly, but the cells could no longer divide. The IE72 protein did not have a significant effect on the S phase. Since IE86 protein inhibits cell cycle progression, the IE2 gene in a human fibroblast IE86 protein-expressing cell line was sequenced. The IE86 protein in these retrovirus-transduced cells has mutations in a critical region of the vital protein. The locations of the mutations and the function of the IE86 protein in controlling cell cycle progression are discussed.

UR - http://www.scopus.com/inward/record.url?scp=0033928395&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0033928395&partnerID=8YFLogxK

U2 - 10.1128/JVI.74.15.7108-7118.2000

DO - 10.1128/JVI.74.15.7108-7118.2000

M3 - Article

VL - 74

SP - 7108

EP - 7118

JO - Journal of Virology

JF - Journal of Virology

SN - 0022-538X

IS - 15

ER -