DYRK1A regulates B cell acute lymphoblastic leukemia through phosphorylation of FOXO1 and STAT3
Rahul S. Bhansali, … , Sébastien Malinge, John D. Crispino
Rahul S. Bhansali, … , Sébastien Malinge, John D. Crispino
Published January 4, 2021
Citation Information: J Clin Invest. 2021;131(1):e135937. https://doi.org/10.1172/JCI135937.
View: Text | PDF
Research Article Oncology

DYRK1A regulates B cell acute lymphoblastic leukemia through phosphorylation of FOXO1 and STAT3

Abstract

DYRK1A is a serine/threonine kinase encoded on human chromosome 21 (HSA21) that has been implicated in several pathologies of Down syndrome (DS), including cognitive deficits and Alzheimer’s disease. Although children with DS are predisposed to developing leukemia, especially B cell acute lymphoblastic leukemia (B-ALL), the HSA21 genes that contribute to malignancies remain largely undefined. Here, we report that DYRK1A is overexpressed and required for B-ALL. Genetic and pharmacologic inhibition of DYRK1A decreased leukemic cell expansion and suppressed B-ALL development in vitro and in vivo. Furthermore, we found that FOXO1 and STAT3, transcription factors that are indispensable for B cell development, are critical substrates of DYRK1A. Loss of DYRK1A-mediated FOXO1 and STAT3 signaling disrupted DNA damage and ROS regulation, respectively, leading to preferential cell death in leukemic B cells. Thus, we reveal a DYRK1A/FOXO1/STAT3 axis that facilitates the development and maintenance of B-ALL.

Authors

Rahul S. Bhansali, Malini Rammohan, Paul Lee, Anouchka P. Laurent, Qiang Wen, Praveen Suraneni, Bon Ham Yip, Yi-Chien Tsai, Silvia Jenni, Beat Bornhauser, Aurélie Siret, Corinne Fruit, Alexandra Pacheco-Benichou, Ethan Harris, Thierry Besson, Benjamin J. Thompson, Young Ah Goo, Nobuko Hijiya, Maria Vilenchik, Shai Izraeli, Jean-Pierre Bourquin, Sébastien Malinge, John D. Crispino

×

Figure 4

DYRK1A phosphorylates FOXO1 and regulates late cell-cycle progression.

Options: View larger image (or click on image) Download as PowerPoint
DYRK1A phosphorylates FOXO1 and regulates late cell-cycle progression. (...
(A) Western blot results of an in vitro kinase assay with GST-DYRK1A, GFP-FOXO1, and ATP-γ-S. Alkylated reaction products were analyzed for thiophosphate esters. (B) Western blot showing p-FOXO1 (Ser326) and total FOXO1 protein in pre-B cells after 4 hours of treatment with EHT 1610 (i and ii indicate separate blots from the same cell extracts). (C) Western blot showing DYRK1A, p-FOXO1 (Ser326), and total FOXO1 protein in Dyrk1a+/+ or Dyrk1afl/fl pre-B cells after transduction with MIGR1-Cre (i and ii indicate separate blots from the same cell extracts). (D) GFP immunofluorescence of primary murine pre-B cells transfected with pcDNA-GFP-FOXO1 (WT, S329A, or S329E) for 48 hours. Cells were treated with DMSO or EHT 1610 for 4 hours and 10 μg/mL Hoechst 33342 for 1 hour prior to imaging. Images are representative of 4 independent experiments. Scale bars: 2.5 μm. (E) Representative flow cytometric plots of pre-B cell distribution in early G2-M (4N, pY15+) or late G2-M (4N, pY15–) after a 48-hour treatment with EHT 1610, AS1842856, or both using p-CDK1 (Tyr15) versus DNA content. Numbers indicate the percentage of live cells in each gate. The early/late G2-M ratio is indicated in red. (F) Quantification of the early/late G2-M ratio based on gating in E. Data indicate the mean ± SD. (G) Quantification of CellTrace Violet dye dilution assay data after a 48-hour treatment with EHT 1610, AS1842856, or both. Data indicate the mean ± SD. (H) Quantification of the percentage of γ-H2AX+ cells in primary murine pre-B cells treated with EHT 1610, AS1842856 (100 nM, 500 nM, or 1000 nM represented by a scale bar), or both for 48 hours. Data indicate the mean ± SD. (I) Quantification of flow cytometric analysis of apoptosis using annexin V staining in primary murine pre-B cells treated with EHT 1610, AS1842856 (100 nM, 500 nM, or 1000 nM represented by a scale bar), or both for 48 hours. Data indicate the mean ± SD. (J) Gadd45a and Foxo1 mRNA expression following qRT-PCR in primary murine pre-B cells after treatment with EHT 1610, AS1842856, or both. Data indicate the mean ± SD (from triplicate wells of a representative sample). n = 3 biological replicates (A–C and E–J). Significance was determined by ANOVA with post hoc Bonferroni’s correction (F–J). *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001.