PKCϵ controls protection against TRAIL in erythroid progenitors

Abstract

Apoptosis plays a central role in the regulation of the size of the hematopoietic stem cell pool as well as in the processes of cell differentiation along the various hematopoietic lineages. TRAIL is a member of the TNF family of cytokines with a known apoptogenic role against a variety of malignant cells and an emerging role in the modulation of normal hematopoiesis. Here we worked on the hypothesis that PKC⑀ could act as a switch of the cellular response to TRAIL during erythropoiesis.We demonstrate that EPO-induced erythroid CD34 cells are insensitive to the apoptogenic effect of TRAIL at day 0 due to the lack of specific receptor expression. From day 3 onward, erythroid cells express surface death receptors and become sensitive to TRAIL up to day 7/8 when, notwithstanding death-receptor expression, the EPO-driven up-regulation of PKC⑀ intracellular levels renders differentiating erythroid cells resistant to TRAIL likely via Bcl-2 up-regulation. Our conclusion is that in human CD34 cells, EPO promotes a series of events that, being finely regulated in their kinetics, restricts the sensitivity of these cells to TRAIL to a specific period of time, which therefore represents the "TRAIL window" for the negative regulation of erythroid-cell numbers. ( IntroductionIt is now well established that apoptosis plays a central role in the regulation of the size of the hematopoietic stem cell pool 1 as well as in the processes of cell differentiation along the various hematopoietic lineages. Fetal erythropoiesis can be negatively regulated by Fas ligand, 2,3 although bone marrow hematopoiesis does not appear to be affected by Fas deficiency. 3 However, Fas expression on hematopoietic stem cells or hematopoietic progenitors can be induced by certain cytokines such as IFN␥ or tumor necrosis factor ␣ (TNF␣), [3][4][5] reducing hematopoietic repopulating potential. Relatively little is known on the effects of other members of the TNF family on hematopoietic progenitors. We have demonstrated that TNF-related apoptosis-inducing ligand (TRAIL) acts as a negative regulator of adult erythropoiesis, selectively reducing the number of erythroblasts in liquid culture, as well as reducing the number and size of erythroid colonies in semisolid assays. 6 Recently, Secchiero et al demonstrated that TRAIL inhibited the generation of mature erythroblasts in liquid culture through the activation of an ERK 1/2-mediated signaling pathway. 7 TRAIL is a member of the TNF family of cytokines, which are structurally related proteins playing important roles in regulating cell death, immune response, and inflammation. 8,9 The unique feature of TRAIL, compared with other members of the TNF family, is its ability to induce apoptosis in a variety of malignant cells both in vitro and in vivo, displaying minimal toxicity on normal cells and tissues. 10,11 TRAIL interacts with 4 high-affinity transmembrane receptors belonging to the apoptosis-inducing TNF-receptor (R) family. TRAIL-R1 (DR4) and TRAIL-R2 (DR5) transduce apoptotic signals on binding of TR...

Downregulation of Bid is associated with PKCɛ-mediated TRAIL resistance

“…18,19,21,22,38 We also found that overexpression of PKCe did not alter the levels of TRAIL receptors or interfere with the recruitment of caspase-8 to the DISC. Overexpression of PKCe caused a decrease in Bid protein and mRNA in MCF-7 cells.…”

“…It has been reported that EPOdriven upregulation of PKCe level renders differentiating erythroid cells resistant to TRAIL likely via Bcl-2 upregulation. 21 We have shown that overexpression of PKCe increased Bcl-2 not only at the protein level but also at the mRNA level. In addition, knockdown of PKCe by siRNA decreased Bcl-2 level.…”

“…2,10,[15][16][17][18] Protein kinase C (PKC), a family of phospholipid-dependent serine/threonine kinases, is emerging as a critical regulator of cellular responses to TRAIL. 12,14,[18][19][20][21][22][23][24] The PKC family consists of 11 isoforms that are classified as conventional (a, b1, b2 and g), novel (d, e, Z and y) or atypical (z and i/l). 25 PKCs are involved in the regulation of cell proliferation, differentiation as well as cell death.…”

“…23 Furthermore, PKCe overexpression did not modify the cell surface expression of TRAIL receptors in erythromyeloid cell lines. 21 It has been reported that mitochondria constitute an important target in overcoming inherent resistance to TRAIL. 2,3,10,16,19,22,32 However, PKCe did not affect Bcl-2 and Bax in glioma cells.…”

Protein kinase Cε makes the life and death decision

“…In MCF-7 cells, however, overexpression of the PKCε regulatory domain decreased Bcl-2 protein levels and increased susceptibility to apoptosis induced by multiple factors although the effects of PKCα regulatory domain was more pronounced [133]. It has been reported that EPO-driven upregulation of PKCε levels renders differentiating erythroid cells resistant to TRAIL, likely via Bcl-2 upregulation [134].…”