Journal Article

Reduced DOCK4 expression leads to erythroid dysplasia in myelodysplastic syndromes

John Anastasi, Andrew Artz, Thomas Quenon, Michelle M. Le Beau, Subhradip Karmakar, David L. Ebenezer, Madhu Unnikrishnan, Hui Liu, Jacqueline Boultwood, Ulrich Steidl, Matthias Bartenstein, Kathy L. McGraw, Vijay Yajnik, Ryan Duggan, Amit Verma, Todd Evans, Amittha Wickrema, Ting-Chun Liu, Sriram Sundaravel, Tushar Bhagat, Ingrid Torregroza, Yiting Yu, Andrea Pellagatti, Alan F. List

2015

Overview

Anemia is the predominant clinical manifestation of myelodysplastic syndromes (MDS). Genes that are aberrantly expressed and/or mutated that lead to the dysplastic erythroid morphology seen in −7/del(7q) MDS have not been identified. In this study, we show that reduced expression of dedicator of cytokinesis 4 ( DOCK4 ) causes dysplasia by disrupting the actin cytoskeleton in developing red blood cells. In addition, our identification of the molecular pathway that leads to morphological defects in this type of MDS provides potential therapeutic targets downstream of DOCK4 that can be exploited to reverse the dysplasia in the erythroid lineage. Furthermore, we developed a novel single-cell multispectral flow cytometry assay for evaluation of disrupted F-actin filaments, which can be used for potential early detection of dysplastic cells in MDS. Anemia is the predominant clinical manifestation of myelodysplastic syndromes (MDS). Loss or deletion of chromosome 7 is commonly seen in MDS and leads to a poor prognosis. However, the identity of functionally relevant, dysplasia-causing, genes on 7q remains unclear. Dedicator of cytokinesis 4 (DOCK4) is a GTPase exchange factor, and its gene maps to the commonly deleted 7q region. We demonstrate that DOCK4 is underexpressed in MDS bone marrow samples and that the reduced expression is associated with decreased overall survival in patients. We show that depletion of DOCK4 levels leads to erythroid cells with dysplastic morphology both in vivo and in vitro. We established a novel single-cell assay to quantify disrupted F-actin filament network in erythroblasts and demonstrate that reduced expression of DOCK4 leads to disruption of the actin filaments, resulting in erythroid dysplasia that phenocopies the red blood cell (RBC) defects seen in samples from MDS patients. Reexpression of DOCK4 in −7q MDS patient erythroblasts resulted in significant erythropoietic improvements. Mechanisms underlying F-actin disruption revealed that DOCK4 knockdown reduces ras-related C3 botulinum toxin substrate 1 (RAC1) GTPase activation, leading to increased phosphorylation of the actin-stabilizing protein ADDUCIN in MDS samples. These data identify DOCK4 as a putative 7q gene whose reduced expression can lead to erythroid dysplasia.

• Publisher: National Acad Sciences,National Academy of Sciences
• Subject: Actins - genetics / Actins - metabolism / Anemia / Animals / Bioassays / Biological Sciences / Blood diseases / Bone marrow / Calmodulin-Binding Proteins - genetics / Calmodulin-Binding Proteins - metabolism / Erythroblasts - metabolism / Erythroblasts - pathology / Female / Gene expression / Gene Expression Regulation / GTPase-Activating Proteins - biosynthesis / GTPase-Activating Proteins - genetics / Humans / Male / Myelodysplastic Syndromes - genetics / Myelodysplastic Syndromes - metabolism / Myelodysplastic Syndromes - pathology / Phosphorylation / PNAS Plus / rac1 GTP-Binding Protein - genetics / rac1 GTP-Binding Protein - metabolism / Toxins / Zebrafish - genetics / Zebrafish - metabolism / Zebrafish Proteins - genetics / Zebrafish Proteins - metabolism