HOX cluster intergenic non-coding RNAs in myeloid differentiation and function

描述(由申请人提供)：中性粒细胞和分化的髓系细胞异常丰富地表达非编码RNA(NcRNA)转录本，特别是来自同源盒(Hox)基因簇的基因间转录本。与已被充分研究的HOX基因在胚胎模式形成中的功能相比，人们对它们在造血中的作用知之甚少。我们已经确定了一个HOX区域ncRNA，HOTAIRM1，它在髓系细胞中特异表达，并调节近端Hoxa簇中的基因表达。我们建议对HOXA簇中的基因间转录本进行详细的研究，目的是确定HOTAIRM1和其他HOXA ncRNAs在调控HOX基因表达、髓系分化和成熟中性粒细胞功能中的功能。中心假说：HOTAIRM1和HOXA基因簇中的其他非编码基因间转录本调节发育重要的HOX基因的表达，从而调节髓系分化和功能。具体地说，我们将：1.研究HOTAIRM1在HOX基因表达调控中的作用。假设：HOTAIRM1在髓系分化过程中调节HOX基因的表达模式。每个子目标测试一个关于特定调控模式的假设：a.通过顺式或反式作用调节HOXA簇中的单个HOX基因；b.对5‘和3’HOXA簇基因组的差异调控；c.优先抑制或激活HOXB和非簇同源盒基因2.确定HOTAIRM1调控HOX基因表达的机制。以下每个子目标都测试了一个特定的、非排他性的假设，即正在研究的特定机制有助于HOTAIRM1的调节功能：a.HOTAIRM1的亚细胞定位和分子邻居；b.与HOTAIRM1相关的分子；c.转录活性的调节；d.染色质结构的调节；e.转录后调节mRNA的稳定性和翻译。3.检测HOXA簇基因间ncRNAs对髓系发育和功能的影响。A.HOTAIRM1基因敲除或过表达对髓系基因表达、分化和功能的影响。假设：HOTAIRM1通过控制髓系基因和下游效应器来调节髓系的承诺、分化和功能。B.敲除或过度表达额外的HoxA基因间ncRNAs的影响。假设：多个HOX区域基因间转录协同调节髓系基因的表达和功能。在髓系细胞中表达的其他HoxA基因间ncRNAs的特征；额外的HoxA基因间转录本的敲除和过度表达本研究将增加我们对髓系基因调控、分化和功能的基本理解。这些结果可能揭示干预髓系成熟障碍的潜在靶点，如骨髓发育不良和白血病，以及中性粒细胞介导的炎症障碍。 公共卫生相关性：拟议的研究应确定一种新的非编码RNA转录本在髓系分化和成熟的中性粒细胞功能中的作用。这些结果将增加我们对这些过程的基本了解，并可能揭示干预髓系成熟障碍(如骨髓发育不良和白血病)以及中性粒细胞介导的炎性疾病(如关节炎、炎症性肠病和缺血后组织损伤)的潜在靶点。

DESCRIPTION (provided by applicant): Neutrophils and differentiating myeloid cells are unusually rich in the expression of non-coding RNA (ncRNA) transcripts, particularly intergenic transcripts from the homeobox-containing (HOX) gene clusters. In contrast to the well-studied functions of HOX genes in embryonic pattern formation, their roles in hematopoiesis are less well understood. We have identified a HOX region ncRNA, HOTAIRM1, that is expressed specifically in myeloid cells and regulates the expression of genes in the proximal HOXA cluster. We propose a detailed study of intergenic transcripts in the HOXA cluster, with the objectives of determining the functions of HOTAIRM1 and other HOXA ncRNAs in the regulation of HOX gene expression, myeloid differentiation, and mature neutrophil function. Central hypothesis: HOTAIRM1 and other non-coding intergenic transcripts in the HOXA gene cluster regulate the expression of developmentally important HOX genes, thereby modulating myeloid differentiation and function. Specifically, we will: 1. Investigate the function of HOTAIRM1 in the regulation of HOX gene expression. Hypothesis: HOTAIRM1 regulates the pattern of HOX gene expression during myeloid differentiation. Each subaim tests a hypothesis regarding a specific regulatory pattern: A. Regulation of individual HOX genes within the HOXA cluster, by either cis or trans actions; B. Differential regulation of sets of 5' versus 3' HOXA cluster genes; C. Preferential repression or activation of HOXB and non-clustered homeobox genes 2. Determine the mechanisms of HOTAIRM1 regulation of HOX gene expression. Each of the following subaims tests a specific, non-exclusive hypothesis that the specific mechanism under examination contributes to the regulatory functions of HOTAIRM1: A. Subcellular localization and molecular neighbors of HOTAIRM1; B. Molecules associated with HOTAIRM1; C. Regulation of transcriptional activity; D. Regulation of chromatin structure; E. Post-transcriptional regulation of mRNA stability and translation. 3. Test the effects of HOXA cluster intergenic ncRNAs on myeloid development and function. A. Effects of HOTAIRM1 knockdown or overexpression on myeloid gene expression, differentiation and function. Hypothesis: HOTAIRM1 regulates myeloid lineage commitment, differentiation, and function through control of myeloid genes and downstream effectors. B. Effects of knockdown or overexpression of additional HOXA intergenic ncRNAs. Hypothesis: Multiple HOX region intergenic transcripts cooperatively regulate myeloid gene expression and function. Characterization of other HOXA intergenic ncRNAs expressed in myeloid cells; knockdown and overexpression of additional HOXA intergenic transcripts The proposed studies will increase our fundamental understanding of myeloid gene regulation, differentiation, and function. The results could reveal potential targets for intervention in disorders of myeloid maturation, such as myelodysplasia and leukemia, as well as neutrophil-mediated inflammation disorders. PUBLIC HEALTH RELEVANCE: The proposed studies should determine the roles of a novel non-coding RNA transcripts in myeloid differentiation and mature neutrophil function. The results will increase our fundamental understanding of these processes and could reveal potential targets for intervention in disorders of myeloid maturation, such as myelodysplasia and leukemia, and in neutrophil-mediated inflammatory disorders, such as arthritis, inflammatory bowel disease, and post-ischemic tissue injury.