Immunoreceptor tyrosine-based activation motif-dependent signaling by Kaposi's sarcoma-associated herpesvirus K1 protein: Effects on lytic viral replication

The Kaposi's sarcoma-associated herpesvirus (KSHV) K1 gene encodes a polype ptide bearing an immuno-receptor tyrosine-based activation motif (ITAM) tha t is constitutively active for ITAM-based signal transduction, Although ect opic overexpression of K1 in cultured fibroblasts can lead to growth transf ormation, in vivo this gene is primarily expressed in lymphoid cells underg oing lytic infection. Here we have examined function of K1 in the setting o f lytic replication, through the study of K1 mutants lacking functional ITA Ms. Expression of such mutants in BJAB cells cotransfected with wild-type K 1 results in dramatic inhibition of K1 signal transduction, as judged by im paired activation of Syk kinase and phospholipase C-gamma2 as well as by di minished expression of a luciferase reporter gene dependent upon K1-induced calcium and Ras signaling. Thus, the mutants behave as dominantly acting i nhibitors of K1 function. To assess the role of K1 in lytic replication, we introduced these K1 mutants into BCBL-1 cells, a B-cell lymphoma line late ntly infected with KSHV, and induced lyric replication by ectopic expressio n of the KSHV ORF50 transactivator. Expression of lytic cycle genes was dim inished up to 80% in the presence of a K1 dominant negative mutant. These i nhibitory effects could be overridden by tetradecanoyl phorbol acetate trea tment, indicating that inhibition was not due to irreversible cell injury a nd suggesting that other signaling events could bypass the block. We conclu de that ITAM-dependent signaling by K1 is not absolutely required for lytic reactivation but functions to modestly augment lyric replication in B cell s, the natural reservoir of KSHV.