Neonatal expression of RNA-binding protein IGF2BP3 regulates the human fetal-adult megakaryocyte transition
Kamaleldin E. Elagib, Chih-Huan Lu, Goar Mosoyan, Shadi Khalil, Ewelina Zasadzińska, Daniel R. Foltz, Peter Balogh, Alejandro A. Gru, Deborah A. Fuchs, Lisa M. Rimsza, Els Verhoeyen, Miriam Sansó, Robert P. Fisher, Camelia Iancu-Rubin, Adam N. Goldfarb
Kamaleldin E. Elagib, Chih-Huan Lu, Goar Mosoyan, Shadi Khalil, Ewelina Zasadzińska, Daniel R. Foltz, Peter Balogh, Alejandro A. Gru, Deborah A. Fuchs, Lisa M. Rimsza, Els Verhoeyen, Miriam Sansó, Robert P. Fisher, Camelia Iancu-Rubin, Adam N. Goldfarb
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Research Article Hematology

Neonatal expression of RNA-binding protein IGF2BP3 regulates the human fetal-adult megakaryocyte transition

Abstract

Hematopoietic transitions that accompany fetal development, such as erythroid globin chain switching, play important roles in normal physiology and disease development. In the megakaryocyte lineage, human fetal progenitors do not execute the adult morphogenesis program of enlargement, polyploidization, and proplatelet formation. Although these defects decline with gestational stage, they remain sufficiently severe at birth to predispose newborns to thrombocytopenia. These defects may also contribute to inferior platelet recovery after cord blood stem cell transplantation and may underlie inefficient platelet production by megakaryocytes derived from pluripotent stem cells. In this study, comparison of neonatal versus adult human progenitors has identified a blockade in the specialized positive transcription elongation factor b (P-TEFb) activation mechanism that is known to drive adult megakaryocyte morphogenesis. This blockade resulted from neonatal-specific expression of an oncofetal RNA-binding protein, IGF2BP3, which prevented the destabilization of the nuclear RNA 7SK, a process normally associated with adult megakaryocytic P-TEFb activation. Knockdown of IGF2BP3 sufficed to confer both phenotypic and molecular features of adult-type cells on neonatal megakaryocytes. Pharmacologic inhibition of IGF2BP3 expression via bromodomain and extraterminal domain (BET) inhibition also elicited adult features in neonatal megakaryocytes. These results identify IGF2BP3 as a human ontogenic master switch that restricts megakaryocyte development by modulating a lineage-specific P-TEFb activation mechanism, revealing potential strategies toward enhancing platelet production.

Authors

Kamaleldin E. Elagib, Chih-Huan Lu, Goar Mosoyan, Shadi Khalil, Ewelina Zasadzińska, Daniel R. Foltz, Peter Balogh, Alejandro A. Gru, Deborah A. Fuchs, Lisa M. Rimsza, Els Verhoeyen, Miriam Sansó, Robert P. Fisher, Camelia Iancu-Rubin, Adam N. Goldfarb

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Figure 6

IGF2BP3 knockdown in neonatal megakaryocytes elicits adult phenotypic and molecular features.

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IGF2BP3 knockdown in neonatal megakaryocytes elicits adult phenotypic an...
(A–C) IGF2BP3 knockdown in neonatal megakaryocytes enhances enlargement, growth arrest, and polyploidization. CB progenitors transduced with lentiviral vectors were puromycin selected, cultured for 6 days in megakaryocyte medium, and analyzed by flow cytometry for FSC (A), PKH dye dilution (B), and PI staining of DNA (C). Analyses were on gated viable CD41+ cells. Graphs show mean ± SEM for 3 independent experiments. *P < 0.05; **P < 0.01, t test. (D and E) IGF2BP3 knockdown in neonatal megakaryocytes enhances P-TEFb activation and expression of target factors. Whole cell lysates of cells from A–C were immunoblotted for indicated factors as in Figure 2. Graphs show mean ± SEM for scanning densitometry values derived from 3 independent experiments, with signal normalization to tubulin, with control vector values set at 1. *P < 0.05; **P < 0.01; ***P < 0.005, t test. See also Supplemental Figures 8–10.