Thrombocytopenia-associated mutations in the ANKRD26 regulatory region induce MAPK hyperactivation
Dominique Bluteau, … , Remi Favier, Hana Raslova
Dominique Bluteau, … , Remi Favier, Hana Raslova
Published January 16, 2014
Citation Information: J Clin Invest. 2014;124(2):580-591. https://doi.org/10.1172/JCI71861.
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Research Article Hematology

Thrombocytopenia-associated mutations in the ANKRD26 regulatory region induce MAPK hyperactivation

Abstract

Point mutations in the 5′ UTR of ankyrin repeat domain 26 (ANKRD26) are associated with familial thrombocytopenia 2 (THC2) and a predisposition to leukemia. Here, we identified underlying mechanisms of ANKRD26-associated thrombocytopenia. Using megakaryocytes (MK) isolated from THC2 patients and healthy subjects, we demonstrated that THC2-associated mutations in the 5′ UTR of ANKRD26 resulted in loss of runt-related transcription factor 1 (RUNX1) and friend leukemia integration 1 transcription factor (FLI1) binding. RUNX1 and FLI1 binding at the 5′ UTR from healthy subjects led to ANKRD26 silencing during the late stages of megakaryopoiesis and blood platelet development. We showed that persistent ANKRD26 expression in isolated MKs increased signaling via the thrombopoietin/myeloproliferative leukemia virus oncogene (MPL) pathway and impaired proplatelet formation by MKs. Importantly, we demonstrated that ERK inhibition completely rescued the in vitro proplatelet formation defect. Our data identify a mechanism for development of the familial thrombocytopenia THC2 that is related to abnormal MAPK signaling.

Authors

Dominique Bluteau, Alessandra Balduini, Nathalie Balayn, Manuela Currao, Paquita Nurden, Caroline Deswarte, Guy Leverger, Patrizia Noris, Silverio Perrotta, Eric Solary, William Vainchenker, Najet Debili, Remi Favier, Hana Raslova

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Figure 7

Inhibition of sustained MAPK pathway in MKs of patients leads to the correction of PPT formation defect.

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Inhibition of sustained MAPK pathway in MKs of patients leads to the cor...
In vitro MK differentiation was induced from control or patient peripheral blood CD34+ progenitors in the presence of TPO. At day 8, the DMSO (–PD98059) or the MAPK inhibitor PD98059 (+PD98059) was added to the cultures performed in liquid medium. The percentage of PPT-forming MKs was estimated by counting MKs exhibiting 1e or more cytoplasmic processes with areas of constriction at different days of culture. A total of 200 cells per well were counted. The histograms show 1 experiment for each THC2 patient with its respective control performed in triplicate. Data represent mean ± SD of triplicate. (A–C) Histograms show the percentage of PPT bearing MKs before and after inhibition of MAPK pathway with PD98059 inhibitor. (A) PD3_2 and PD5_III4 patient with their respective control, (B) PD5_III2 and PD5_IV1 patients, (C) PD3_3 patient and its respective control. (D and E) Representative microscopic images of PPT formation by control and patient MKs before and after addition of PD98059 inhibitor. (D) PD5_III2 and PD5_IV1 patients. (E) PD3_3 patient and its respective control.