Mutation of the Putative Catalytic Site of LUNA Abrogates HCMV Reactivation from Latency
(A) HFFs were infected at an MOI of 0.1 with WT Merlin, LUNA catalytic dead mutant (LUNAFUN-MUT), or WT virus and the growth measured over 14 days. Supernatants were titered for infectious virus production every 2 days.
(B) Western blotting on mock (M), LUNASHORT (KO), WT, revertant (Rev), or LUNA functional mutant (FM) at 24 and 72 hpi for viral protein expression.
(C and D) CD34+ cells latently infected with WT or a catalytic dead mutant of LUNA (LUNAFUN-MUT) were differentiated to mature DCs. Induction of IE gene expression (C) or reactivation of virus by infectious center formation in co-cultures (D) was quantified by qRT-PCR or immuno-staining, respectively. Data are mean ± SD and represent triplicate analyses performed in three independent experiments. ∗∗p < 0.01 and ∗∗∗p < 0.001.