Transcriptomic and phospho-proteomic analyzes of erythroblasts expanded in vitro from normal donors and from patients with polycythemia vera

Todd Hricik, Giulia Federici, Ann Zeuner, Giuliana Alimena, Agostino Tafuri, Valentina Tirelli, Lilian Varricchio, Francesca Masiello, Fiorella Ciaffoni, Stefania Vaglio, Emanuel F. Petricoin, Gabriella Girelli, Ross L. Levine, Anna Rita F Migliaccio

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

Erythropoiesis is a tightly regulated process which becomes decoupled from its normal differentiation program in patients with polycythemia vera (PV). Somatic mutations in JAK2 are commonly associated with this myeloid proliferative disorder. To gain insight into the molecular events that are required for abnormally developing erythroid cells to escape dependence on normal growth signals, we performed in vitro expansion of mature erythroblasts (ERY) from seven normal healthy donors and from seven polycythemic patients in the presence of IL3, EPO, SCF for 10, 11, or 13 days. Normal ERYs required exposure to the glucocorticoid dexamethasone (Dex) for expansion, while PV-derived ERYs expanded in the absence of dexamethasone. RNA expression profiling revealed enrichment of two known oncogenes, GPR56 and RAB4a, in PV-derived ERYs along with reduced expression levels of transcription factor TAL1 (ANOVA FDR <0.05). While both normal and polycythemic-derived ERYs integrated signaling cascades for growth, they did so via different signaling pathways which are represented by their differential phospho-profiles. Our results show that normal ERYs displayed greater levels of phosphorylation of EGFR, PDGFRβ, TGFβ, and cKit, while PV-derived ERYs were characterized by increased phosphorylation of cytoplasmic kinases in the JAK/STAT, PI3K, and GATA1 pathways. Together these data suggest that PV erythroblast expansion and maturation may be maintained and enriched in the absence of dexamethasone through reduced TAL1 expression and by accessing additional signaling cascades. Members of this acquired repertoire may provide important insight into the pathogenesis of aberrant erythropoiesis in myeloproliferative neoplasms such as polycythemia vera.

Original languageEnglish
Pages (from-to)723-729
Number of pages7
JournalAmerican Journal of Hematology
Volume88
Issue number9
DOIs
Publication statusPublished - Sep 2013

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Erythroblasts
Polycythemia Vera
Proteomics
Tissue Donors
Dexamethasone
Erythropoiesis
Phosphorylation
Janus Kinases
Erythroid Cells
Growth
Phosphatidylinositol 3-Kinases
Oncogenes
Glucocorticoids
In Vitro Techniques
Analysis of Variance
Transcription Factors
RNA
Mutation
Neoplasms

ASJC Scopus subject areas

  • Hematology

Cite this

Transcriptomic and phospho-proteomic analyzes of erythroblasts expanded in vitro from normal donors and from patients with polycythemia vera. / Hricik, Todd; Federici, Giulia; Zeuner, Ann; Alimena, Giuliana; Tafuri, Agostino; Tirelli, Valentina; Varricchio, Lilian; Masiello, Francesca; Ciaffoni, Fiorella; Vaglio, Stefania; Petricoin, Emanuel F.; Girelli, Gabriella; Levine, Ross L.; Migliaccio, Anna Rita F.

In: American Journal of Hematology, Vol. 88, No. 9, 09.2013, p. 723-729.

Research output: Contribution to journalArticle

Hricik, T, Federici, G, Zeuner, A, Alimena, G, Tafuri, A, Tirelli, V, Varricchio, L, Masiello, F, Ciaffoni, F, Vaglio, S, Petricoin, EF, Girelli, G, Levine, RL & Migliaccio, ARF 2013, 'Transcriptomic and phospho-proteomic analyzes of erythroblasts expanded in vitro from normal donors and from patients with polycythemia vera', American Journal of Hematology, vol. 88, no. 9, pp. 723-729. https://doi.org/10.1002/ajh.23487
Hricik, Todd ; Federici, Giulia ; Zeuner, Ann ; Alimena, Giuliana ; Tafuri, Agostino ; Tirelli, Valentina ; Varricchio, Lilian ; Masiello, Francesca ; Ciaffoni, Fiorella ; Vaglio, Stefania ; Petricoin, Emanuel F. ; Girelli, Gabriella ; Levine, Ross L. ; Migliaccio, Anna Rita F. / Transcriptomic and phospho-proteomic analyzes of erythroblasts expanded in vitro from normal donors and from patients with polycythemia vera. In: American Journal of Hematology. 2013 ; Vol. 88, No. 9. pp. 723-729.
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abstract = "Erythropoiesis is a tightly regulated process which becomes decoupled from its normal differentiation program in patients with polycythemia vera (PV). Somatic mutations in JAK2 are commonly associated with this myeloid proliferative disorder. To gain insight into the molecular events that are required for abnormally developing erythroid cells to escape dependence on normal growth signals, we performed in vitro expansion of mature erythroblasts (ERY) from seven normal healthy donors and from seven polycythemic patients in the presence of IL3, EPO, SCF for 10, 11, or 13 days. Normal ERYs required exposure to the glucocorticoid dexamethasone (Dex) for expansion, while PV-derived ERYs expanded in the absence of dexamethasone. RNA expression profiling revealed enrichment of two known oncogenes, GPR56 and RAB4a, in PV-derived ERYs along with reduced expression levels of transcription factor TAL1 (ANOVA FDR <0.05). While both normal and polycythemic-derived ERYs integrated signaling cascades for growth, they did so via different signaling pathways which are represented by their differential phospho-profiles. Our results show that normal ERYs displayed greater levels of phosphorylation of EGFR, PDGFRβ, TGFβ, and cKit, while PV-derived ERYs were characterized by increased phosphorylation of cytoplasmic kinases in the JAK/STAT, PI3K, and GATA1 pathways. Together these data suggest that PV erythroblast expansion and maturation may be maintained and enriched in the absence of dexamethasone through reduced TAL1 expression and by accessing additional signaling cascades. Members of this acquired repertoire may provide important insight into the pathogenesis of aberrant erythropoiesis in myeloproliferative neoplasms such as polycythemia vera.",
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AU - Ciaffoni, Fiorella

AU - Vaglio, Stefania

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