TRPC3 Is the Erythropoietin-regulated Calcium Channel in Human Erythroid Cells

Abstract

2؉ ] i was reduced. Our data demonstrate that Epo modulates TRPC3 activation through a PLC␥-mediated process that requires interaction of PLC␥ and IP 3 R with TRPC3. They also show that TRPC3 Tyr 226 is critical in Epo-dependent activation of TRPC3. These data demonstrate a redundancy of TRPC channel activation mechanisms by widely different agonists. Erythropoietin (Epo)2 is a glycoprotein that is required for proliferation and differentiation of erythroid cells (1, 2). The erythropoietin receptor (Epo-R) is a member of the cytokine receptor superfamily, members of which share many signal transduction pathways (3). Epo has been shown to stimulate a dose-dependent increase in [Ca 2ϩ ] i that is mediated through a voltage-independent ion channel (4 -6). In electrophysiological studies of normal human erythroid progenitor-derived cells, Epo stimulation increased calcium channel mean open time 2.5-fold and open probability 10-fold (5). To identify specific channels activated by erythropoietin, members of the transient receptor potential (TRP) protein superfamily were studied, because these channels have characteristics similar to those observed in electrophysiological studies of human erythroblasts (5). We determined that TRPC2, TRPC3, and TRPC6 are expressed on primary erythroid cells and that erythropoietin stimulated calcium influx through murine TRPC2 but not TRPC6 (7-9). Erythropoietin modulated calcium influx through TRPC2 through signaling mechanisms dependent on complex formation between TRPC2, Epo-R, phospholipase C␥ (PLC␥), and the inositol 1,4,5-trisphosphate receptor (IP 3 R), activation of PLC␥, and interaction of TRPC2 with IP 3 R (10). However, because TRPC2 is a pseudogene in humans (11), we hypothesized that the function of TRPC2 is provided by a different calcium-permeable channel in human erythroid cells.Calcium is a universal intracellular second messenger that influences many cell functions and in erythroid cells has an important role in colony growth and in terminal stages of differentiation (6,(12)(13)(14). The erythropoietin receptor also has been shown to activate Ca 2ϩ influx in other cell types. In myoblasts, Epo stimulated expansion of the progenitor population during differentiation and an increase in [Ca 2ϩ ] i dependent on extracellular calcium influx (15). In neuronal cell lines, Epo stimulated an increase in cell viability and an increase in 45 Ca 2ϩ uptake (16,17). Determination of the mechanisms through which the erythropoietin receptor modulates Ca 2ϩ influx is important in understanding regulation of erythroid proliferation and differentiation as well as the role of Epo-R expression in nonerythroid tissues and is likely to be applicable to other cytokine receptor pathways.

Identification and Functional Characterization of Ion Channels in CD34+ Hematopoietic Stem Cells from Human Peripheral Blood

“…Here we report that TRPM2, TRPC6, and TRPV2-like currents are functionally expressed in CD34 + HSCs. Although previous reports using CD34 + cells have discussed the expression of various TRP mRNAs and have conducted Ca 2+ influx experiments (den Dekker et al, 2001;Hirschler-Laszkiewicz et al, 2009;Tong et al, 2008;Zhang et al, 2006), our direct demonstration of ion channel currents is unprecedented.…”

Role of Helix-Loop-Helix Proteins during Differentiation of Erythroid Cells

“…The concentration of calcium is low in early erythroid progenitor cells, increases for a short period of time, and then decreases thereafter, reaching its lowest concentration in mature red blood cells. Other reports suggest that there may be several short periods during the differentiation of red blood cells that are characterized by a transient increase in the intracellular calcium concentration (93,131). Nevertheless, changes in the calcium concentration during erythroid cell differentiation may be important for the regulation of Myc, Tal1, and USF.…”

Protein kinase D-HDAC5 signaling regulates erythropoiesis and contributes to erythropoietin cross-talk with GATA1

“…25,26 In addition, signaling elements upstream of PKD, PLC-␥1, and PKC-⑀ are activated by erythropoietin and have been implicated in erythropoietic regulation. 31,32 PLC-␥1 null mice display embryonic lethality at day 9.5-10.5 of development with complete absence of erythroid progenitors but no deficit of granulocyte or monocyte progenitors. 33 The mechanistic basis for their unusual phenotype of lineage-selective erythroid aplasia has not been elucidated.…”