Expansion of NKG2C⁺ NK cells in response to fibroblasts infected with HCMV deletion mutants. (Ai) The expression of total class I molecules (HLA-I) and HLA-E was analyzed by flow cytometry in MRC-5 fibroblasts. The expression of HLA-E was also undetectable in AD169-infected MRC-5 cells (not shown). (Aii) Staining of an HLA-E⁺ cell line (.221-AEH) is shown for comparison. (Aiii) Expression of HLA-I in MRC-5 cells infected for 72 hours with either AD169, HB5 (ΔUS2-6), or HB5-ΔUS2-11 mutants; HLA expression in cells infected with HB5-ΔUS40-42 or HB5-ΔUS14-20 was comparable to HB5 (not shown). Production of type I IFN upon HCMV infection up-regulates HLA class I expression.⁴³  In this experiment, the presence of a fraction of bystander noninfected cells among AD169-treated cells provides a suitable internal control. Numbers correspond to the mean fluorescence intensity (MFI) of the histograms. (B) PBLs from an HCMV⁺ donor were cultured in parallel with MRC-5 cells infected (MOI = 1) with the wild-type AD169 HCMV strain or different deletion mutants generated from the HB5 BACmid clone (ΔUS2-6). Two-color flow cytometry analysis was performed by day 10. Data are representative of 6 different experiments. The proportion of NKG2C⁺ cells in mock-infected cultures (not shown) was 10.5%.