The lytic activation of KSHV during keratinocyte differentiation is dependent upon a suprabasal position, the loss of integrin engagement, and calcium, but not the interaction of cadherins

We previously found that KSHV (HHV-8) lytic activation occurs during differentiation of oral keratinocytes in organotypic raft cultures. To further investigate the spatial and temporal aspects of KSHV lytic activation and the roles of integrins, cadherins, and calcium, we used rKSHV.219-infected primary oral keratinocytes in submerged, suspension, and direct suprabasal plating, models of differentiation. We found that early keratinocyte differentiation did not activate lytic KSHV in cells attached to a substratum, with activation only occurring in suprabasal cells. Temporally, KSHV lytic expression occurred between the expression of early and late differentiation markers. Keratinocytes differentiated in suspension culture, which mimics substratum loss that occurs with stratification, activated lytic KSHV. This lytic activation was inhibited by integrin engagement, showing that integrins are a control point for KSHV reactivation. A role for cadherins was not found. Elevated extracellular calcium was necessary, but not sufficient, for lytic activation.

Research Highlights► Early differentiation does not activate lytic gene expression in attached keratinocytes. ► KSHV lytic activation during differentiation requires a suprabasal position. ► KSHV lytic activation occurs prior to expression of late markers of differentiation. ► Integrin engagement controls KSHV lytic activation. ► Elevated extracellular calcium is necessary, but not sufficient, to activate KSHV.