Ssx2ip regulates erythroid progenitor cell cycle progression and proliferation. (A) mRNA expression of cell cycle regulatory genes Cdk4, Cdk6, and Chek1 in GFP-sorted CD71⁺Ter119⁻ (R2) and CD71⁺Ter119⁺ (R3) cells infected with shControl or shSsx2ip-2-expressing retrovirus. (B) Representative flow cytometric gating of lineage-depleted spleen cells infected with shControl or shSsx2ip-2-expressing retrovirus, cultured for 3 days, and incubated with BrdU for 1 hour. Ter119⁺ cells were stained with DAPI, anti-CD71 and anti-BrdU antibodies. (C) Quantitation of percentage of G1/S/G2/M-phase cells (from Figure 5A) in CD71⁻Ter119⁻, CD71^lowTer119⁻, CD71^highTer119⁻, CD71^highTer119⁺, CD71^lowTer119⁺, and CD71⁻Ter119⁺ subpopulations. (D) Representative plots depicting ModFit analysis of cell proliferation generations (in different colors) after 48 hours using CellTrace Violet. R3 population is shown. (E) Quantitation of percentage of cells in each cell proliferation generation based on CellTrace Violet. (F) Proliferation index of R3 cells after Ssx2ip knockdown after 48 hours incubation with CellTrace Violet. Error bars represent SD. ∗P < .05 (two-tailed unpaired Student t test).