Intrabodies targeting the Kaposi sarcoma-associated herpesvirus latency antigen inhibit viral persistence in lymphoma cells.
Kaposi sarcoma-associated herpesvirus (KSHV) latency-associated nuclear antigen-1 (LANA1) is essential for the maintenance and segregation of viral episomes in KSHV latently infected B cells. We report development of intracellular, rabbit-derived antibodies generated by phage display technology, which bind to N-terminal LANA1 epitopes and neutralize the chromosome-binding activity of LANA1. Although these cloned single-chain variable fragments (scFvs) show relatively low binding affinities for the LANA1 viral antigen in in vitro assays, they nonetheless outcompete KSHV-seropositive human sera for LANA1 epitope binding. In heterologous cells, intracellular intrabody expression inhibits LANA1-dependent plasmid maintenance of both an artificial plasmid containing KSHV LANA1 binding sequences and a bacterial artificial chromosome containing the entire KSHV genome. In KSHV naturally infected primary effusion lymphoma cells, intracellular intrabody expression causes a reduction or loss of the typical LANA1 punctate, nuclear pattern. This morphologically apparent LANA1 dispersion correlates to loss of viral episome by molecular analysis. These data suggest a novel approach to antiherpes viral therapy and confirm LANA1 is critical target for neutralization of KSHV viral latency.
Corte-Real
Sofia
S
URIA-Centro de Patogénese Molecular, Faculty of Pharmacy, University of Lisbon, 1649-019 Lisbon, Portugal.
Collins
Chris
C
Aires da Silva
Frederico
F
Simas
J Pedro
JP
Barbas
Carlos F
CF
Chang
Yuan
Y
Moore
Patrick
P
Goncalves
Joao
J
D000595:Amino Acid Sequence; D000818:Animals; D000956:Antigens, Viral / Q000235:genetics / Q000276:immunology*; D002460:Cell Line; D019288:Herpesvirus 8, Human / Q000502:physiology; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000276:immunology / Q000494:pharmacology*; D008223:Lymphoma / Q000821:virology*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics / Q000276:immunology*; D019151:Peptide Library; D011817:Rabbits; D012514:Sarcoma, Kaposi / Q000276:immunology / Q000821:virology*; D014162:Transfection; D017735:Virus Latency / Q000187:drug effects*
D000956:Antigens, Viral; D007135:Immunoglobulin Variable Region; D009687:Nuclear Proteins; D019151:Peptide Library; C109540:latency-associated nuclear antigen