Abstract 2965

Poster Board II-941

The long-term expression of human herpesvirus 8 (HHV-8) K1 produces hyperplasia of lymph nodes, splenomegaly, and lymphomas in mice. The mechanism of how K1 causes hyperplasia and lymphomas. The immunoreceptor tyrosine-based activation motif (ITAM) of K1 was shown previously to activate of Akt and nuclear factor kappa B (NF-kB). However, we have recently shown that K1 suppresses Fas-mediated apoptosis through its extracellular immunoglobulin-like domain and that K1-transfected mice survive a lethal dose of agonistic anti-Fas antibody (Jo2). We thus hypothesized that development of hyperplasia and lymphomas in K1-expressing mice is driven by altered Fas signaling.

Examination of mice expressing K1 via a ubiquitous promoter showed that K1 transgenic mice (n=10) had 90% developed lymphoid hyperplasia (at least 3 lymph nodes >3 mm) and 60% developed lymphomas after 18 months, while all (26) control nontransgenic mice remained free of lymph node hyperplasia, splenomegaly, and lymphoma. Some K1 mice developed liver or mesenteric tumors (4 of 10 mice). The spleens of 78% of K1 mice were enlarged at 18 months and were on average 3.5 times heavier than spleens of non-K1 transgenic control mice. Hematoxylin and eosin staining of spleen sections showed lymphocyte expansion in the periarteriolar lymphocyte sheath with disruption of normal spleen architecture. Anti-kappa and anti-lambda light chain antibodies revealed the presence of monoclonal foci in 3 out of 3 K1 mice (average 6 foci per single section of spleen), but no foci were present in 4 control non-transgenic mice. Moreover, K1 protein was expressed in approximately 10% of splenic cells after staining with anti-K1 antibody 2H5. To test the hypothesis that expression of K1 protein confers resistant to Fas-mediated apoptosis, splenic cells of 6-month-old K1 mice (n=3) and matched controls (n=3) were isolated and incubated with 50 ng/mL of agonistic anti-Fas antibody Jo2. At 12 hours of treatment, only 4±1% of splenocytes from K1 mice versus 17±2% of control splenocytes showed morphology of apoptosis (P<0.01). At 24 hours of treatment, the difference was even more significant (11±0.6% vs. 50±6%, P<0.005). Splenocytes of K1 mice were indeed more resistant to Jo2-induced apoptosis compared with splenocytes from age-matched control mice. Of mice inoculated with a lethal dose of Jo2 antibody, 3 out of 12 K1 transgenic (30%) and 13 out of 22 control mice (60%) died (P<0.05), further confirming the protective effect of K1 against Fas-mediated apoptosis.

We mapped the region that K1 uses to bind to Fas to an immunoglobulin (Ig) chain-like domain by expressing deletion mutants of K1. Overexpression of an Ig domain-containing protein CD79b competed with K1-Fas binding in a dose-dependent manner. Two 20-amino acid peptides (N251, N253) representing the Ig domain of K1 competed with K1-Fas binding in immunoprecipitation/immunoblotting analysis. The N251 and N253 peptides (100 mM) enhanced anti-Fas antibody (CH-11, 50 ng/mL)-induced apoptosis of BJAB lymphoma cells that expressed K1 but not that of vector-transfected BJAB cells. K1 splenocytes incubated with N251 and N253 showed enhanced rates of Fas-mediated apoptosis over control peptide-treated K1, and peptides did not enhance the apoptosis rates of control non-K1 expressing splenocytes.

This analysis indicates a key role of K1 in lymphoid hyperplasia and lymphoma and that K1 direct binding to Fas occurs in vivo, which is effectively targeted by competing peptides to restore apoptosis.


No relevant conflicts of interest to declare.

Author notes


Asterisk with author names denotes non-ASH members.