Modeling genetic modifiers of hematopoiesis with induced pluripotent stem cells

描述（由申请人提供）：了解基因如何相互作用以协调组织发育并导致人类疾病是生物医学中的一个基本问题。通过对唐氏综合征（DS，21三体，T21）和造血转录因子加塔-1的研究，我们被这个难题所吸引。DS引起多种造血异常，包括红细胞增多症、血小板减少症和两种相关的克隆性疾病：短暂性骨髓增生性疾病（TMD）和急性巨核细胞白血病（AMKL）。后一种疾病是多步骤进展的一部分，其需要体细胞GATA 1突变，导致称为加塔-1 s的83个氨基酸截短蛋白。GATA 1中类似的种系突变导致整倍体患者贫血。这些临床观察提出了几个有趣的问题：1）21号染色体（HSA 21）上的哪些基因调控造血？2)加塔-1氨基末端如何促进造血分化？3)相同的GATA 1突变如何导致T21和非T21患者的不同疾病？和4）T21和GATA 1突变如何独特地协同作用以引起骨髓增生？小鼠模型提供了重要的信息，但不能完全概括人类疾病。我们正在研究这些问题，使用人胎肝标本和诱导多能干细胞（iPSC）从DS，TMD和GATA 1相关贫血患者产生。初步研究表明，具有T21和加塔-1的iPSC表现出明显的造血异常，其概括了相关人类疾病的许多方面。现在，我们将对具有T21、加塔-1或两者的iPSC中的原始（卵黄囊型）和定形（胎肝型）造血进行系统表征。我们将操纵候选HSA 21基因在iPSCs中的表达，以确定那些负责DS相关血液异常的基因。同时，我们将使用我们的患者来源的iPSC和MEP样细胞系来研究 GATA 1突变如何失调造血基因表达，并通过努力鉴定与加塔-1氨基末端相互作用的蛋白质来研究相关机制。我们的研究将阐明GATA 1和HSA 21基因，单独和一起，调节造血。更一般地说，我们希望创造新的范例，其中其他造血疾病可以通过创建和操纵患者来源的iPSC来建模。

DESCRIPTION (provided by applicant): Understanding how genes interact to coordinate tissue development and cause human disease is a fundamental problem in biomedicine. We are drawn to this puzzle through studies of Down syndrome (DS, trisomy 21, T21) and the hematopoietic transcription factor GATA-1. DS causes multiple hematopoietic abnormalities including polycythemia, thrombocytopenia and two related clonal disorders: transient myeloproliferative disorder (TMD) and acute megakaryoblastic leukemia (AMKL). The latter disorders are part of a multi-step progression that requires somatic GATA1 mutations resulting in an 83 amino acid truncated protein termed GATA-1s. Similar germline mutations in GATA1 cause anemia in euploid patients. These clinical observations raise several interesting questions: 1) what genes on chromosome 21 (HSA21) regulate hematopoiesis? 2) How does the GATA-1 amino terminus facilitate hematopoietic differentiation? 3) How do the same GATA1 mutations cause different diseases in patients with and without T21? and 4) How do T21 and GATA1 mutations synergize uniquely to cause myeloproliferation? Murine models have provided important information, but do not fully recapitulate the human diseases. We are studying these problems using human fetal liver specimens and induced pluripotent stem cells (iPSCs) generated from patients with DS, TMD, and GATA1s-associated anemias. Preliminary studies indicate that iPSCs with T21 and GATA-1s exhibit distinct hematopoietic abnormalities that recapitulate many aspects of the associated human disorders. Now, we will perform systematic characterization of primitive (yolk sac-type) and definitive (fetal liver-type) hematopoiesis in iPSCs with T21, GATA-1s or both. We will manipulate the expression of candidate HSA21 genes in iPSCs to identify those responsible for DS-associated blood abnormalities. In parallel, we will use our patient-derived iPSCs and a MEP-like cell line to study how GATA1s mutations dysregulate hematopoietic gene expression and investigate the associated mechanisms through efforts to identify proteins that interact with the GATA-1 amino terminus. Our studies will elucidate how GATA1 and HSA21 genes, separately and together, modulate hematopoiesis. More generally, we hope to create new paradigms in which other hematopoietic diseases can be modeled through creation and manipulation of patient-derived iPSCs.