The erythropoietic stress response is essential for survival and recovery from anemias, cardio-pulmonary disease, chemotherapy and bone-marrow transplantation. It consists of a dramatic expansion in erythropoietic tissue leading to increased erythropoietic rate. Its principal mediator is the hormone erythropoietin, whose receptor, EpoR, is expressed by erythroid progenitors. The molecular targets of EpoR signaling during stress are largely unknown, and the spectrum of progenitors it regulates during stress is not fully defined. Our long-term goal is to elucidate the molecular mechanisms critical to the erythropoeitic stress response. A major obstacle had been the lack of specific cell-surface markers identifying differentiation stage-specific erythroid progenitors in vivo as they are undergoing stress. We have developed a flow- cytometric assay utilizing cell-surface CD71 and Ter119 that allows us to recognize stage-specific erythroblasts directly in hematopoeitic tissue. Using this assay, we found that the EpoR-activated transcription factor StatS is a key regulator of early erythroblast survival. Mice lacking StatS are anemic and have a deficient stress response. Therefore, our principal hypothesis suggests that early erythroblast survival directly regulates erythropoietic rate and the stress response. The present proposal focuses on the death- receptor Fas and its ligand, Fast. Using the flow-cytometric CD71/Ter119 assay, we found that both are expressed by approximately 50% of early erythroblasts in spleen. Fas expression was inversely related to early erythroblast survival and frequency in tissue. Further, chronic stress in mice or acute Epo administration result in down-regulation of Fas and FasL from the early erythroblast cell surface.
Our specific aims are: (1) Investigate Fas and FasL function in early erythroblasts in vivo. We will generate Fas and FasL mutant mice on an immune-deficient background free of autoimmune hemolysis, to test the hypothesis that early erythroblast Fas negatively regulates basal erythropoiesis. (2) Identify the molecular mechanisms responsible for down-regulation of erythroblast Fas and FasL during stress, by comparing early erythroblasts freshly isolated from mouse models of stress to equivalent cells isolated from control mice. 3) We will use EpoR+/- and erythroid-specific StatS-deficient mice to study the role of EpoR and StatS signaling in stress- induced erythroblast Fas/ FasL down-regulation. We will also test whether the deficient stress response of these mice is rescued when bred onto a Fas or FasL-mutant background. Relevance: In order to survive and recover from blood loss, anemia, or therapeutic procedures such as bone-marrow transplantation and chemotherapy, our bodies need to be able to generate red blood cells at a much higher rate than normal. The proposed work aims to understand mechanisms regulating the rapid production of red cells. This may contribute to future therapies of anemias, including anemia and fatigue associated with cancer.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL084168-05
Application #
7813877
Study Section
Hematopoiesis Study Section (HP)
Program Officer
Qasba, Pankaj
Project Start
2006-04-01
Project End
2012-03-31
Budget Start
2010-04-01
Budget End
2012-03-31
Support Year
5
Fiscal Year
2010
Total Cost
$394,469
Indirect Cost
Name
University of Massachusetts Medical School Worcester
Department
Pediatrics
Type
Schools of Medicine
DUNS #
603847393
City
Worcester
State
MA
Country
United States
Zip Code
01655
Koulnis, Miroslav; Porpiglia, Ermelinda; Porpiglia, P Alberto et al. (2012) Contrasting dynamic responses in vivo of the Bcl-xL and Bim erythropoietic survival pathways. Blood 119:1228-39
Porpiglia, Ermelinda; Hidalgo, Daniel; Koulnis, Miroslav et al. (2012) Stat5 signaling specifies basal versus stress erythropoietic responses through distinct binary and graded dynamic modalities. PLoS Biol 10:e1001383
Shearstone, Jeffrey R; Pop, Ramona; Bock, Christoph et al. (2011) Global DNA demethylation during mouse erythropoiesis in vivo. Science 334:799-802
Kang, Byoung Heon; Xia, Fang; Pop, Ramona et al. (2011) Developmental control of apoptosis by the immunophilin aryl hydrocarbon receptor-interacting protein (AIP) involves mitochondrial import of the survivin protein. J Biol Chem 286:16758-67
Koulnis, Miroslav; Liu, Ying; Hallstrom, Kelly et al. (2011) Negative autoregulation by Fas stabilizes adult erythropoiesis and accelerates its stress response. PLoS One 6:e21192
Koulnis, Miroslav; Pop, Ramona; Porpiglia, Ermelinda et al. (2011) Identification and analysis of mouse erythroid progenitors using the CD71/TER119 flow-cytometric assay. J Vis Exp :
Pop, Ramona; Shearstone, Jeffrey R; Shen, Qichang et al. (2010) A key commitment step in erythropoiesis is synchronized with the cell cycle clock through mutual inhibition between PU.1 and S-phase progression. PLoS Biol 8:
Socolovsky, Merav; Murrell, Michael; Liu, Ying et al. (2007) Negative autoregulation by FAS mediates robust fetal erythropoiesis. PLoS Biol 5:e252
Socolovsky, Merav (2007) Molecular insights into stress erythropoiesis. Curr Opin Hematol 14:215-24