Pluripotent stem cells reveal erythroid-specific activities of the GATA1 N-terminus
Marta Byrska-Bishop, … , Mitchell J. Weiss, Stella T. Chou
Marta Byrska-Bishop, … , Mitchell J. Weiss, Stella T. Chou
Published January 26, 2015
Citation Information: J Clin Invest. 2015;125(3):993-1005. https://doi.org/10.1172/JCI75714.
View: Text | PDF
Research Article

Pluripotent stem cells reveal erythroid-specific activities of the GATA1 N-terminus

Abstract

Germline GATA1 mutations that result in the production of an amino-truncated protein termed GATA1s (where s indicates short) cause congenital hypoplastic anemia. In patients with trisomy 21, similar somatic GATA1s-producing mutations promote transient myeloproliferative disease and acute megakaryoblastic leukemia. Here, we demonstrate that induced pluripotent stem cells (iPSCs) from patients with GATA1-truncating mutations exhibit impaired erythroid potential, but enhanced megakaryopoiesis and myelopoiesis, recapitulating the major phenotypes of the associated diseases. Similarly, in developmentally arrested GATA1-deficient murine megakaryocyte-erythroid progenitors derived from murine embryonic stem cells (ESCs), expression of GATA1s promoted megakaryopoiesis, but not erythropoiesis. Transcriptome analysis revealed a selective deficiency in the ability of GATA1s to activate erythroid-specific genes within populations of hematopoietic progenitors. Although its DNA-binding domain was intact, chromatin immunoprecipitation studies showed that GATA1s binding at specific erythroid regulatory regions was impaired, while binding at many nonerythroid sites, including megakaryocytic and myeloid target genes, was normal. Together, these observations indicate that lineage-specific GATA1 cofactor associations are essential for normal chromatin occupancy and provide mechanistic insights into how GATA1s mutations cause human disease. More broadly, our studies underscore the value of ESCs and iPSCs to recapitulate and study disease phenotypes.

Authors

Marta Byrska-Bishop, Daniel VanDorn, Amy E. Campbell, Marisol Betensky, Philip R. Arca, Yu Yao, Paul Gadue, Fernando F. Costa, Richard L. Nemiroff, Gerd A. Blobel, Deborah L. French, Ross C. Hardison, Mitchell J. Weiss, Stella T. Chou

×

Figure 5

PCA identifies lineage signatures and reveals heterogeneity within progenitor cell populations.

Options: View larger image (or click on image) Download as PowerPoint
PCA identifies lineage signatures and reveals heterogeneity within proge...
Single cells within CD43+CD41+CD235+ hematopoietic progenitor populations derived from WT and mutant iPSCs were purified by flow cytometry and analyzed for gene expression by qPCR. For comparison, committed erythroid (CD41–CD235+), myeloid (CD45+CD18+), and megakaryocytic (CD41+CD42+) cells generated from WT iPSCs were purified and analyzed in parallel. (A) Numbers of analyzed single cells of each genotype (total 755 single cells). WT lineage committed cells: myeloid (myelo), megakaryocytic, and erythroid (ery). CD43+CD41+CD235+ progenitor genotypes: euploid/WT GATA1, euploid/GATA1s, T21/WT GATA1, T21/GATA1s. (B) PCA on 170 committed erythroid (red), myeloid (green), and megakaryocytic (blue) cells based on expression patterns of 94 genes (right plot). PC loadings obtained from this analysis (right) were used to project 274 GATA1s (purple) and 311 WT GATA1 (black) progenitor cells onto PC1 and PC2 (left). (C) Smooth kernel density estimate of PC1 scores of GATA1s (purple line) and WT GATA1 progenitors (black line). Purple and black vertical lines represent mean PC1 scores. Shaded areas around the mean correspond to 90% confidence interval for the mean. *P = 10–13 (Mann-Whitney U test). (D) Violin plots showing distributions of single-cell expression levels of 3 genes that were upregulated (left) and 3 genes that were downregulated (right) in GATA1s vs. WT GATA1 progenitors in 5 cell types analyzed (FDR < 0.05; Mann-Whitney U test followed by BH-FDR correction). lfc, lfc of mean expression between GATA1s and WT GATA1 progenitors.