Unraveling crucial roles of homeobox gene Hlx in hematopoiesis and leukemogenesis

描述(由申请人提供)：尽管干细胞的分离在过去几十年中取得了显著进展，但对调控干细胞自我更新和谱系承诺的确切机制的了解仍然有限。在造血过程中，造血干细胞(HSC)的后代致力于分化为特定的细胞系，最终产生终末分化的细胞。转录因子已经被认为能够驱动一组特定的谱系特异性靶标的表达 基因，指示前体细胞采取一定的分化程序。转录因子活性的失调在白血病中起着重要作用，这意味着这些基因可能成为血液和其他形式癌症治疗干预的靶点。当我们在小鼠急性髓系白血病(AML)模型中分析纯化的白血病前造血干细胞和祖细胞(HSPC)时，我们发现一个新的非簇同源盒基因HLX上调了4倍，这表明HLX可能参与了健康的造血和恶性转化。我们的初步研究表明，HLX的过表达破坏了健康的髓系分化，并在体外赋予了无限的系列克隆性和对造血干/祖细胞的粒-单核细胞分化障碍。此外，HLX的过度表达会导致表型HSC的丧失，并在体内持续存在扩大的、异常的髓系祖细胞群体。我们还发现，HLX调控着一个对HSPC的谱系承诺和髓系分化至关重要的基因网络。值得注意的是，我们发现HLX在大多数AML患者中过度表达，并且HLX表达是AML患者生存的最强预测因素之一。我们还发现，HLX下调抑制了小鼠和人AML细胞的体外生长。该项目旨在了解HLX是如何在HSC和髓系分化过程中调节这些关键功能的。为了确定HLX在干细胞和祖细胞的谱系承诺中的作用，以及在髓系分化和急性髓系白血病细胞中的作用，我们将利用遗传小鼠模型、干细胞移植试验和体内和体外靶向降低HLX的水平。为了阐明HLX的作用机制，我们将研究我们通过转录图谱确定的下游途径，并进行染色质免疫沉淀，以建立HLX作为能够直接调节其靶基因的转录因子。

DESCRIPTION (provided by applicant): While isolation of stem cells has advanced dramatically in the last few decades, understanding of the precise mechanisms that regulate the self-renewal and lineage commitment of a stem cell is still limited. During hematopoiesis, progeny of hematopoietic stem cells (HSC) become committed to differentiate into specific cell lineages to ultimately generate terminally differentiated cells. Transcription factors have been recognized for their ability to drive expression of a characteristic set of lineage-specific target genes, instructing a precursor cell to adopt a certain differentiation program. Dysregulation of transcription factor activity has an important role in leukemia, implicating these genes as potential targets for therapeutic intervention in blood, and other forms of cancer. When we analyzed purified pre-leukemic hematopoietic stem and progenitor cells (HSPC) in a murine acute myeloid leukemia (AML) model, we found 4-fold upregulation of a novel non-clustered homeobox gene, H2.0-like homeobox (Hlx), suggesting that Hlx may be involved in healthy hematopoiesis and malignant transformation. Our preliminary studies indicate that overexpression of Hlx disrupts healthy myeloid differentiation and confers unlimited serial clonogenicity and a myelomonocytic differentiation block to hematopoietic stem and progenitor cells in vitro. Furthermore, overexpression of Hlx causes loss of phenotypic HSC and persistence of an expanded, aberrant myeloid progenitor population in vivo. We also find that Hlx regulates a network of genes important for lineage commitment and myeloid differentiation of HSPC. Strikingly, we find that Hlx is overexpressed in the majority of patients with AML, and that Hlx expression is one of the strongest predictors of AML patient survival. We also find that Hlx downregulation inhibits growth of murine and human AML cells in vitro. This project aims to understand how Hlx is regulating these critical functions in HSC and during myeloid differentiation. To characterize the roles of Hlx in lineage commitment of stem and progenitor cells, as well as in myeloid differentiation and acute myeloid leukemia cells, we will utilize genetic murine models, stem cell transplantation assays and targeted reduction of Hlx levels in vivo and in vitro. To elucidate the mechanism of action of Hlx, we will study downstream pathways we have identified by transcriptional profiling and perform chromatin-immunoprecipitation to establish Hlx as a transcription factor capable of directly regulating its target genes.