RAB14 Regulates Human Erythropoiesis and Megakaryopoiesis

We recently reported that RAB GTPase 14 (RAB14) knockdown (KD) increased the frequency and total numbers of erythroid cells generated in vitro in response to erythropoietin (EPO) from either the TF1 human leukemia erythropoietic model cell line or from primary human CD34⁺ hematopoietic stem-progenitor cells (HSPCs). RAB14 overexpression (OE) had the opposite effect. Thus, RAB14 functions as an endogenous inhibitor of human erythropoiesis (Kim et al., Br. J. Haematol., 2015). In contrast to the greater cell numbers generated in the presence of EPO, RAB14 KD TF1 cells grown in standard culture media containing granulocyte-macrophage colony-stimulating factor (GM-CSF; as the only cytokine) generated fewer total cells, compared to empty vector-transduced control TF1 cells. Furthermore, RAB14 KD TF1 cells cultured in GM-CSF media generated greater numbers of erythroid (CD34^-/CD71⁺/CD235a⁺) cells, as compared to control TF1 cells, suggesting that RAB14 KD stimulated erythropoiesis even in the absence of EPO. Cells generated from RAB14 KD TF1 cells had higher GATA1 and lower GATA2 transcription factor expression, as compared to controls, demonstrating the cells had undergone the "GATA1/2 switch," a hallmark of erythropoiesis. Consistent with higher GATA1 levels, RAB14 KD TF1 cells generated cells with higher levels of b- and g-hemoglobins. Similarly, RAB14 KD in primary human CD34⁺ HSPCs generated greater numbers of erythroid cells, with or without exogenous EPO. RAB14 KD CD34⁺ HSPCs cultured in GM-CSF media generated fewer monocytic/granulocytic (CD13⁺/CD33⁺) cells, as compared to control CD34⁺ HSPCs. Interestingly, RAB14 OE CD34⁺ HSPCs cultured in thrombopoietin (TPO)-containing media generated higher numbers of megakaryocytic (CD34^-/CD41a⁺/CD42b⁺) cells, as compared to control CD34⁺ HSPCs. In summary, (1) RAB14 KD in TF1 cells or primary human CD34⁺ HSPCs increased erythropoiesis in the presence or absence of EPO, but reduced myeloid cell differentiation, probably via the GATA1/2 switch; and (2) RAB14 OE in CD34⁺ HSPCs increased megakaryopoiesis in the presence of TPO. Thus, RAB14 normally serves as an endogenous hematopoietic decision-maker, physiologically inhibiting erythropoiesis and stimulating megakaryopoiesis (and possibly, to a lesser extent, mono-granulopoiesis).