Identification of the Essential Role of Viral Bcl-2 for Kaposi's Sarcoma-Associated Herpesvirus Lytic Replication.
Kaposi's sarcoma-associated herpesvirus (KSHV) evades host defenses through tight suppression of autophagy by targeting each step of its signal transduction: by viral Bcl-2 (vBcl-2) in vesicle nucleation, by viral FLIP (vFLIP) in vesicle elongation, and by K7 in vesicle maturation. By exploring the roles of KSHV autophagy-modulating genes, we found, surprisingly, that vBcl-2 is essential for KSHV lytic replication, whereas vFLIP and K7 are dispensable. Knocking out vBcl-2 from the KSHV genome resulted in decreased lytic gene expression at the mRNA and protein levels, a lower viral DNA copy number, and, consequently, a dramatic reduction in the amount of progeny infectious viruses, as also described in the accompanying article (A. Gelgor, I. Kalt, S. Bergson, K. F. Brulois, J. U. Jung, and R. Sarid, J Virol 89:5298-5307, 2015). More importantly, the antiapoptotic and antiautophagic functions of vBcl-2 were not required for KSHV lytic replication. Using a comprehensive mutagenesis analysis, we identified that glutamic acid 14 (E14) of vBcl-2 is critical for KSHV lytic replication. Mutating E14 to alanine totally blocked KSHV lytic replication but showed little or no effect on the antiapoptotic and antiautophagic functions of vBcl-2. Our study indicates that vBcl-2 harbors at least three important and genetically separable functions to modulate both cellular signaling and the virus life cycle.


OBJECTIVE
The present study shows for the first time that vBcl-2 is essential for KSHV lytic replication. Removal of the vBcl-2 gene results in a lower level of KSHV lytic gene expression, impaired viral DNA replication, and consequently, a dramatic reduction in the level of progeny production. More importantly, the role of vBcl-2 in KSHV lytic replication is genetically separated from its antiapoptotic and antiautophagic functions, suggesting that the KSHV Bcl-2 carries a novel function in viral lytic replication.
Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA.
Qiming
Q
Liang
Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA.
Brian
B
Chang
Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA.
Patrick
P
Lee
Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA.
Kevin F
KF
Brulois
Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA.
Jianning
J
Ge
Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA.
Mude
M
Shi
Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA Diagnostics Division, Infectious Disease Research, Abbott Laboratories, Abbott Park, Illinois, USA.
Mary A
MA
Rodgers
Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA.
Pinghui
P
Feng
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon, Republic of Korea.
Byung-Ha
BH
Oh
Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA.
Chengyu
C
Liang
Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA jaeujung@med.usc.edu.
Jae U
JU
Jung
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D017209:Apoptosis; D001343:Autophagy; D001483:Base Sequence; D002460:Cell Line; D004261:DNA Replication; D004279:DNA, Viral / Q000235:genetics; D015870:Gene Expression; D055786:Gene Knockout Techniques; D016679:Genome, Viral; D057809:HEK293 Cells; D019288:Herpesvirus 8, Human / Q000235:genetics / Q000472:pathogenicity / Q000502:physiology*; D054884:Host-Pathogen Interactions; D006801:Humans; D024101:Mitochondrial Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015513:Oncogene Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D014764:Viral Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D014779:Virus Replication / Q000235:genetics / Q000502:physiology*
C452184:Bcl-2 protein, Human herpesvirus 8; D004279:DNA, Viral; C468524:K7 protein, Human herpesvirus 8; D024101:Mitochondrial Proteins; D015513:Oncogene Proteins; D014764:Viral Proteins; C481275:viral FLIP protein, Human herpesvirus 8