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REPLICATION

Function of Rta Is Essential for Lytic Replication of Murine Gammaherpesvirus 68

Ting-Ting Wu, Leming Tong, Tammy Rickabaugh, Samuel Speck, Ren Sun
Ting-Ting Wu
Department of Molecular and Medical Pharmacology, UCLA AIDS Institute, Jonsson Comprehensive Cancer Center, and Molecular Biology Institute, University of California at Los Angeles, Los Angeles, California 90095, and
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Leming Tong
Department of Molecular and Medical Pharmacology, UCLA AIDS Institute, Jonsson Comprehensive Cancer Center, and Molecular Biology Institute, University of California at Los Angeles, Los Angeles, California 90095, and
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Tammy Rickabaugh
Department of Molecular and Medical Pharmacology, UCLA AIDS Institute, Jonsson Comprehensive Cancer Center, and Molecular Biology Institute, University of California at Los Angeles, Los Angeles, California 90095, and
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Samuel Speck
Department of Pathology and Immunology and Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri
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Ren Sun
Department of Molecular and Medical Pharmacology, UCLA AIDS Institute, Jonsson Comprehensive Cancer Center, and Molecular Biology Institute, University of California at Los Angeles, Los Angeles, California 90095, and
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DOI: 10.1128/JVI.75.19.9262-9273.2001
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ABSTRACT

Rta, encoded primarily by open reading frame 50, is well conserved among gammaherpesviruses. It has been shown that the Rta proteins of Epstein Barr virus (EBV), Kaposi's sarcoma-associated herpesvirus (KSHV, or HHV-8), and murine gammaherpesvirus 68 (MHV-68; also referred to as γHV68) play an important role in viral reactivation from latency. However, the role of Rta during productive de novo infection has not been characterized in gammaherpesviruses. Since there are cell lines that can support efficient productive de novo infection by MHV-68 but not EBV or KSHV, we examined whether MHV-68 Rta plays a role in initiating viral lytic replication in productively infected cells. Rta, functioning as a transcriptional activator, can activate the viral promoter of early lytic genes. The amino acid sequence alignments of the Rta homologues suggest that the organizations of their functional domains are similar, with the DNA binding and dimerization domains at the N terminus and the trans-activation domain at the C terminus. We constructed two mutants of MHV-68 Rta, Rd1 and Rd2, with deletions of 112 and 243 amino acids from the C terminus, respectively. Rd1 and Rd2 could no longer trans-activate the promoter of MHV-68 gene 57, consistent with the deletions of theirtrans-activation domains at the C terminus. Furthermore, Rd1 and Rd2 were able to function as dominant-negative mutants, inhibiting trans-activation of wild-type Rta. To study whether Rd1 and Rd2 blocked viral lytic replication, purified virion DNA was cotransfected with Rd1 or Rd2 into fibroblasts. Expression of viral lytic proteins was greatly suppressed, and the yield of infectious viruses was reduced up to 104-fold. Stable cell lines constitutively expressing Rd2 were established and infected with MHV-68. Transcription of the immediate-early gene, rta, and the early gene, tk, of the virus was reduced in these cell lines. The presence of Rd2 also led to attenuation of viral lytic protein expression and virion production. The ability of Rta dominant-negative mutants to inhibit productive infection suggests that the trans-activation function of Rta is essential for MHV-68 lytic replication. We propose that a single viral protein, Rta, governs the initiation of MHV-68 lytic replication during both reactivation and productive de novo infection.

  • Copyright © 2001 American Society for Microbiology
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Function of Rta Is Essential for Lytic Replication of Murine Gammaherpesvirus 68
Ting-Ting Wu, Leming Tong, Tammy Rickabaugh, Samuel Speck, Ren Sun
Journal of Virology Oct 2001, 75 (19) 9262-9273; DOI: 10.1128/JVI.75.19.9262-9273.2001

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Function of Rta Is Essential for Lytic Replication of Murine Gammaherpesvirus 68
Ting-Ting Wu, Leming Tong, Tammy Rickabaugh, Samuel Speck, Ren Sun
Journal of Virology Oct 2001, 75 (19) 9262-9273; DOI: 10.1128/JVI.75.19.9262-9273.2001
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KEYWORDS

Gammaherpesvirinae
Immediate-Early Proteins
Trans-Activators

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