Mechanism of Fas signaling regulation by human herpesvirus 8 K1 oncoprotein.
BACKGROUND
Herpesvirus 8 (HHV-8) oncoprotein K1 is linked to lymphoproliferation and suppression of apoptosis mediated by the Fas death receptor. Expression of K1 in transgenic mice induces accumulation of lymphoid tissue cells and lymphoma.


METHODS
To examine how K1 and Fas interact to suppress apoptosis, K1-Fas binding was studied in human embryonic kidney (HEK) and lymphoma (BJAB) cells that expressed wild-type K1 or a K1 Ig domain deletion mutant and were treated with Fas ligand (FasL) or an agonistic Fas antibody, using immunoprecipitation and Western blot analysis. Cleavage of caspase-3 and apoptosis was compared in liver samples from mice that were transfected with empty vector vs with plasmids expressing wild-type K1 or a K1 Ig deletion mutant and treated with agonistic Fas antibody for 7 hours. These studies used immunohistochemical staining and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay. All statistical tests were two-sided.


RESULTS
Immunoprecipitation and Western blot analysis of transfected HEK and BJAB cells revealed that wild-type K1 but not Ig-deleted K1 binds to Fas and prevents Fas activation by FasL or by an agonistic Fas antibody. More mice that were transfected with wild-type K1 (7 of 10) than mice transfected with empty vector (3 of 13) or the K1 Ig deletion mutant (0 of 6) survived treatment with the agonistic Fas antibody. Compared with vector-transfected mice, livers of wild-type K1-transfected mice contained fewer cells in which caspase-3 was cleaved (87.6% vs 58.0%, difference = 29.6%, 95% confidence interval [CI] = 19.2% to 40.0%; P = .003) and fewer apoptotic cells (83.7% vs 34.2%, difference = 49.5%, 95% CI = 39.8% to 59.2%; P = .003).


CONCLUSIONS
K1 blocks Fas signaling by directly binding to Fas through the Ig-like domain of K1 and preventing binding of FasL. The relative resistance of cancer cells to Fas-mediated apoptosis may be due to the inhibition of Fas by Ig domain-containing proteins.
Berkova
Zuzana
Z
Department of Lymphoma and Myeloma, The University of Texas MD Anderson Cancer Center, Houston, 77054, USA.
Wang
Shu
S
Wise
Jillian F
JF
Maeng
Hoyoung
H
Ji
Yuan
Y
Samaniego
Felipe
F
D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D053200:Fas-Associated Death Domain Protein / Q000378:metabolism*; D005434:Flow Cytometry; D017353:Gene Deletion; D006801:Humans; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D020287:In Situ Nick-End Labeling; D007668:Kidney / Q000166:cytology / Q000196:embryology / Q000378:metabolism*; D008099:Liver / Q000378:metabolism / Q000473:pathology; D008198:Lymph Nodes / Q000378:metabolism* / Q000473:pathology*; D008223:Lymphoma / Q000378:metabolism*; D051379:Mice; D009154:Mutation; D010957:Plasmids; D015398:Signal Transduction; D014162:Transfection; D014764:Viral Proteins / Q000378:metabolism*
D053200:Fas-Associated Death Domain Protein; C111600:K1 protein, Human herpesvirus 8; D014764:Viral Proteins