RUNX1, but not its familial platelet disorder mutants, synergistically activates PF4gene expression in combination with ETS family proteins

Abstract

WC, Doi T. RUNX1, but not its familial platelet disorder mutants, synergistically activates PF4 gene expression in combination with ETS family proteins. J Thromb Haemost 2013; 11: 1742-50.Summary. Background: Familial platelet disorder (FPD) is a rare autosomal dominant disease characterized by thrombocytopenia and abnormal platelet function. Causal mutations have been identified in the gene encoding runtrelated transcription factor 1 (RUNX1) of FPD patients. Objectives: To elucidate the role of RUNX1 in the regulation of expression of platelet factor 4 (PF4) and to propose a plausible mechanism underlying RUNX1-mediated induction of the FPD phenotype. Methods: We assessed whether RUNX1 and its mutants, in combination with E26 transformation-specific-1 (ETS-1), Core-binding factor subunit beta (CBFb), and Friend leukemia virus integration 1 (FLI-1), cooperatively regulate PF4 expression during megakaryocytic differentiation. In an embryonic stem cell differentiation system, expression levels of endogenous and exogenous RUNX1 and PF4 were determined by real-time RT-PCR. Promoter activation by the transcription factors were evaluated by reporter gene assays with HepG2 cells. DNA binding activity and protein interaction were analyzed by electrophoretic mobility shift assay and immunoprecipitation assay with Cos-7 cells, respectively. Protein localization was analyzed by immunocytochemistry and Western blotting with Cos-7 cells. Results: We demonstrated that RUNX1 activates endogenous PF4 expression in megakaryocytic differentiation. RUNX1, but not its mutants, in combination with ETS-1 and CBFb, or FLI-1, synergistically activated the PF4 promoter. Each RUNX1 mutant harbors various functional abnormalities, including loss of DNA-binding activity, abnormal subcellular localization, and/or alterations of binding affinities for ETS-1, CBFb, and FLI-1. Conclusions: RUNX1, but not its mutants, strongly and synergistically activates PF4 expression along with ETS family proteins. Furthermore, loss of the RUNX1 transcriptional activation function is induced by various functional abnormalities.

MiR-18a increased the permeability of BTB via RUNX1 mediated down-regulation of ZO-1, occludin and claudin-5

“…As mentioned before, overexpression of RUNX1 upregulated the expressions of tight junction related proteins ZO-1, occludin and claudin-5. RUNX1 can also modulate its own level by upregulating the expression of Smad6 to target itself for proteasome degradation [44]. It was found that RUNX1, but not its mutants, synergistically activated the PF4 promoter in combination with ETS-1 and CBFβ, or FLI-1 [45].…”

Broader expression of the mouse platelet factor 4‐cre transgene beyond the megakaryocyte lineage

“…Nevertheless, the regulation of Pf4 expression outside megakaryocytes has received little attention. In the megakaryocytic lineage, it has been suggested that RUNX1, in combination with ETS‐1 and CBFb or FLI‐1, synergistically activates the Pf4 promoter . These transcription factors are active in various non‐hematopoietic cells, especially members of the ETS family, which have been reported to control epithelial maturation in the intestine .…”

Transcription factor defects causing platelet disorders

“…43 More recently, RUNX1 was demonstrated to activate PF4 synergistically with the transcription factors CBF, ETS-1 and FLI-1 during megakaryocytic differentiation. 44 Two preliminary studies reported that the genes encoding RAB1B (RAB1B), a low molecular weight GTPase that is essential for vesicle transport between the endoplasmic reticulum and the Golgi apparatus, and Pallidin (PLDN), which is involved in granule vesicle biogenesis, were both downregulated in RUNX1 haplodeficiency, and shown to be direct targets for RUNX1 regulation, providing possible mechanisms for the defective granule biogenesis and secretion observed in RUNX1 deficiency. 45,46…”