Fig. 8.

TET3 is a miR-150 target involved in monocyte subset differentiation. a Hybridization analysis of 3′-UTR of TET3 with mature sequences of miR-150 using RNA-hybrid software in human and mouse. Structures with minimum free energy are shown. b Expression of TET3 (normalized to B2M) in monocytes generated by culture of human CD34⁺ cells transduced with hsa-miR-150-GFP compared to control-transduced cells (SCR). Results are expressed as TET3 expression fold-change in hsa-miR-150-overexpressing cells relative to control (SCR, normalized to 1). Mean ± SEM of three independent experiments. Paired t test, *P < 0.05. c Immunoblot analysis of TET3 expression in four healthy donor (CTL) and seven CMML patient monocytes. ADAPTIN and ACTIN are the loading controls. TET3/ADAPTIN or TET3/ACTIN ratio are shown below. Blots have been cropped for clarity. d Heat map of TET2 and TET3 gene expression, compared to a selection of differentially expressed genes in classical and nonclassical monocyte subsets as measured by RNA-seq (N = 4 healthy donors). The expression of genes characterizing these two subsets is shown. e Representative flow cytometry analysis of CD11b⁺ or CD43⁺ monocyte populations gated on Ly6C expression in peripheral blood of a wild-type (WT) and a Tet3^−/− (KO) mice. f Monocyte subsets measured by flow cytometry in WT and Tet3^−/− mice. N = 3, mean ± SEM, unpaired t test, *P < 0.05. One representation of three independent experiments is shown. g Heat map of differentially expressed genes in Ly6C^high and Ly6C^low monocyte subsets as measured by RNA-seq of two series of wild-type littermates (N = 3 wild-type mice for Mir150^−/− littermates, and 2 or 3 wild-type mice for Tet3^−/− littermates)