Early growth response transcription factor EGR‐1 regulates Gαq gene in megakaryocytic cells

Abstract

Summary. Background: Gaq (Gene GNAQ) plays a major role in platelet signal transduction but little is known regarding its transcriptional regulation. Objectives: We studied Gaq promoter activity using luciferase reporter gene assays in human erythroleukemia (HEL) cells treated with phorbol 12-myristate 13-acetate (PMA) for 24 h to induce megakaryocytic transformation. Methods and results: PMA-treated HEL cells showed enhanced Gaq expression. Reporter (luciferase) gene studies on 5¢ upstream construct (up to )116 bp from ATG) revealed a negative regulatory site at )238/)202 and two positive sites at )203/)138 and )1116/)731. The positive regulatory region )203/)138 contained overlapping Sp1/AP-2/ EGR-1 consensus sites. Gel shift studies on Gaq oligonucleotides 1 ()203/)175) and 2 ()174/)152) using HEL cell extracts demonstrated protein binding that was due to early growth response factor EGR-1 at two sites. Mutations in either EGR-1 site markedly decreased the gene activity, indicating functional relevance. Mutation of consensus E-Box motif ()185/)180) had no effect. Reduction in the expression of endogenous EGR-1 with antisense oligonucleotide to EGR-1 inhibited PMA-induced Gaq transcription. Correspondingly, Egr-1 deficient mouse platelets also showed 50% reduction in the Gaq expression relative to wild-type platelets. Conclusions: These studies suggest that Gaq gene is regulated during PMA-induced megakaryocytic differentiation by EGR-1, an early growth response transcription factor that regulates a wide array of genes and plays a major role in diverse activities, including cell proliferation, differentiation and apoptosis, and in vascular response to injury and atherosclerosis.

Developmental Control of NRAMP1 (SLC11A1) Expression in Professional Phagocytes

“…Each site constitutes a central motif in DNAse footprints obtained from CMP (mCD34), MEPs (CMK and K562 cells) or myelo-monocytic cells (CD14 + MNs, HL-60 and NB4). Both EGR1 and ELF1 can exert positive or negative regulatory effects on target gene expression in various myeloid lineages [79,80,81,82,83,84,85,86,87,88,89,90,91] so that either factor might inhibit full-length transcription of NRAMP1 in non-expressing cells (see Section 2.3.1.6). Lastly, transcriptional activity at DHS F10 was independently reported in K562 cells [49] thus supporting a regulatory role of this element in myeloid cells.…”

Plant Polyphenol Intake Alters Gene Expression in Canine Leukocytes

“…In platelets, a role has been attributed to PAR-1 in vascular function regulation, with dual relaxant/constrictor effects on the endothelium [48] . GNAQ, also an upregulated gene, has been intensively studied because of its relevant role in cardiovascular signal transduction [49,50] , mainly through its effect on transducing the information of plasma membrane receptors (thromboxane 2, ␣ 1 -adrenergic receptor, platelet-activating factor, thrombin) involved in platelet activation, vasoconstriction and vascular smooth hyperplasia/hypertrophy [51] .…”

Regulation of megakaryocytic differentiation of K562 cells by FosB, a member of the Fos family of AP-1 transcription factors

“…The situation in part stems from the restricted access to cells during various stages of differentiation. Cell line models such as myelogenous leukemia K562 cells [11] and signaling molecules such as PMA (also called TPA) have thus been used extensively to model megakaryopoiesis [11,12]. One of the established findings from such in vitro system is that PMAinduced ERK signaling activates AP-1 activity, which in turn mediates a subset of molecular events involved in megakaryocytic differentiation [13].…”