Mechanisms of Stem Cell Factor and Erythropoietin Proliferative Co-signaling in FDC2-ER Cells

Abstract

Studies of hematopoietic progenitor cell development in vivo, ex vivo, and in factor-dependent cell lines have shown that c-kit promotes proliferation through synergistic effects with at least certain type 1 cytokine receptors, including the erythropoietin (Epo) receptor. Presently, c-kit is shown to efficiently support both mitogenesis and survival in the FDCP1 cell subline, FDC2. In this system, mitogenic synergy with c-kit was observed for ectopically expressed wild-type Epo receptors (wt-ER), an epidermal growth factor (EGF) receptor/Epo receptor chimera, and a highly truncated Epo receptor construct ER-Bx1. Thus, the Epo receptor cytoplasmic box 1 subdomain appears, at least in part, to mediate mitogenic synergy with c-kit. In studies of potential effectors of this response, Jak2 tyrosine phosphorylation was shown to be induced by Epo, but not by stem cell factor (SCF). In addition and in contrast to signaling in Mo7e and BM6 cell lines, in FDC2-ER cells SCF and Epo each were shown to rapidly activate Pim 1 gene expression. Recently, roles also have been suggested for the nuclear trans-factor GATA-1 in regulating progenitor cell proliferation. In FDC2-ER cells, the ectopic expression of GATA-1 had no detectable effect on Epo inhibition of apoptosis. However, GATA-1 expression did result in a selective and marked inhibition in mitogenic responsiveness to SCF and to a decrease in c-kit transcript expression. These studies of SCF and Epo signaling in FDC2–wt-ER cells serve to functionally map the ERB1 region as a c-kit–interactive domain, suggest that Pim1 might contribute to SCF and Epo mitogenic synergy and support the notion that SCF and Epo may act in opposing ways during red cell differentiation.

Synergy between erythropoietin and stem cell factor during erythropoiesis can be quantitatively described without co‐signaling effects

“…The model excludes the contribution of a supra-additive (synergistic) effect of SCF and EPO (in green) on the proliferation rate or on the death/survival rate, as well as co-signaling (in green). reanalysis of cell line systems in which both receptors have been constitutively expressed (Joneja et al, 1997;Tan et al, 2003) may provide definitive evidence of the contribution of a co-signaling mechanism, once the distinct cytokine effects on proliferation and survival have been taken into account.…”

“…Interactions between EPO and SCF have been found to impact erythroid cell output in vivo (de Haan et al, 1995a,b;Molineux et al, 1991;Roeder et al, 1998) and in vitro (Case et al, 2001;Quesniaux et al, 1992). In addition, this synergy appears to be a conserved mechanism of erythroid cell amplification since it has been observed in cultures of peripheral blood (Panzenbock et al, 1998), fetal liver, bone marrow (McNiece et al, 1991) and umbilical cord blood (Case et al, 2001) stem cells, as well as in cultures of multipotential cell lines (Broudy et al, 1993;Joneja et al, 1997).…”

“…It reflects the ability of individual cells to simultaneously respond to both cytokines and in many studies, it has been associated with crosstalk between signaling pathways. For instance, functional interactions between c-kit and EPOR or downstream signaling molecules have been identified in different cell types (Arcasoy and Jiang, 2005;Broudy et al, 1998;Joneja et al, 1997;Pircher et al, 2001;Sui et al, 1998;Wu et al, 1995Wu et al, , 1997 and in primary human cell cultures, a supra-additive activation of a mitogen-activated protein kinase (MAPK) in response to a combination of SCF and EPO was found to correlate with a synergistic erythroid cell expansion (Sui et al, 1998). However, a number of technical issues in these studies prevent them from providing definitive evidence on the contribution of SCF and EPO signaling crosstalk in adult erythropoiesis.…”

“…However, a number of technical issues in these studies prevent them from providing definitive evidence on the contribution of SCF and EPO signaling crosstalk in adult erythropoiesis. For instance, some studies have been performed with (i) established cell lines that do not show a synergistic expansion in response to SCF and EPO (Broudy et al, 1998;Jacobs-Helber et al, 1997;Kapur et al, 1998;Wu et al, 1995); (ii) cell lines that have been engineered to over-express c-kit or EPOR (Joneja et al, 1997;Li et al, 2003;Pircher et al, 2001;Wu et al, 1995); (iii) primary cells from fetal sources that have been engineered to over-express c-kit or EPOR (Wu et al, 1997); and (iv) bulk biochemical assays (e.g., Western blots) on samples that likely contained mixed cell populations (Arcasoy and Jiang, 2005;Boer et al, 2003;Joneja et al, 1997;Kapur and Zhang, 2001;Pircher et al, 2001;Sui et al, 1998).…”

Signaling Induced by Erythropoietin and Stem Cell Factor in UT‐7/Epo Cells: Transient versus Sustained Proliferation

“…In HCD57 cells which express endogenous EpoR and c-kit receptors and in 32D cells transfected with EpoR, initiation of EpoR signaling and proliferation following exposure to SCF in the absence of Epo is observed [19,20]. Furthermore, the cytoplasmic box 1 domain of EpoR in part mediates mitogenic synergy with c-kit in FDCP1 cells transfected with EpoR [21] which occurs in the absence of JAK2 activation. However, while SCF induces cell proliferation and components of the EpoR signaling cascade, long-term cell proliferation is not maintained resulting from incomplete activation of the EpoR signaling cascade.…”