Tmod1 expression is upregulated while Tmod3 is downregulated during erythroid differentiation. (A) mRNA levels of Tmods during in vitro erythroid differentiation from human CD34⁺ cells, determined by qRT-PCR on days 7, 10, and 14, normalized to ACTB mRNA. (B) Western blots of Tmod1 and Tmod3 proteins during in vitro erythroid differentiation of CD34⁺ cells. Cell pellets were collected on days 3, 6, 8, 10, 13, and 16 after initiation of differentiation, solubilized, and analyzed by western blotting (4.1R was used as positive control for erythroid differentiation, and total actin [C4 antibody] and glyceraldehyde-3-phosphate dehydrogenase proteins were used as loading controls). (C-D) qRT-PCR analysis of Tmod1 and Tmod3 mRNAs in highly pure erythroblasts at distinct developmental stages isolated by FACS from (C) in vitro CD34⁺cell-erythroid differentiation cultures, or (D) mouse bone marrow, normalized to ACTB mRNA. Stages were: proerythroblasts (Pro-E), early basophilic erythroblasts (early Baso), late basophilic erythroblasts (late Baso), basophilic erythroblasts (Baso-E), polychromatic erythroblasts (Poly-E), and orthochromatic erythroblasts (Ortho-E).