Extended Data Fig. 4: Influence of the rs112233623-T allele on cyclin D3 expression, growth and cell cycle in erythroid cultures.
From: Reduced cyclin D3 expression in erythroid cells protects against malaria
a, Western blot assay showing cyclin D3 expression in erythroblasts derived from individuals homozygous for the WT rs112233623-C allele (CC, n = 4) versus the derived rs112233623-T allele (TT, n = 5); human β-actin was run on the same gel as loading control. Band intensities in the immunoblots were quantified using ImageJ software using the same detection time point for both signals (1 min) and the differences were expressed as ratio of Cyclin D3 to β-actin means for each group. For gel source data, see Supplementary Fig. 1. b, Growth curves of cultured erythroid progenitor cells derived from individuals homozygous for the WT rs112233623-C allele (CC, red line) versus derived rs112233623-T allele (TT, blue line). The mean ± s.e.m is shown, (for each genotype, n = 3 biologically independent experiments). A two-sided Student’s t-test was used (*P < 0.05; ns, not significant). The statistical results for all comparisons are provided in Supplementary Table 3. c,d, Analysis of erythroblasts derived from individuals homozygous for the WT rs112233623-C allele (c) or the derived rs112233623-T allele (d) at day 7 of erythroid differentiation. In each panel, from left to right: representative flow-cytometry plots with propidium iodide staining, showing the distribution of cell-cycle phases (G0–G1, S, G2–M); dot plots of GPA+ gated erythroblasts showing expression of Band3 and integrin α4 (data are mean ± s.e.m., n = 4 per genotype); and cytospin preparations stained with May–Grünwald–Giemsa (scale bars, 20 μm; 50× magnification).
