Lmo2-Lyl1 and the bHLH factor network in pro-T cells

pro-T 细胞中的 Lmo2-Lyl1 和 bHLH 因子网络

• 批准号: 10624261
• 负责人: ELLEN V. ROTHENBERG
• 金额: $ 61.34万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-15 至 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10624261
• 关键词: Acute; Acute Lymphocytic Leukemia; Affect; B-Lymphocytes; BHLH Protein; Benign; Binding; Binding Sites; Biochemical; Cell Cycle; Cell Differentiation process; Cell Lineage; Cells; Chromatin; Classification; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Data; Development; Dimerization; Dose; Down-Regulation; E protein; Enzymes; Event; GATA3 gene; Gene Expression; Gene Rearrangement; Genes; Genome; Genomics; Helix-Turn-Helix Motifs; Heterodimerization; Human; Immune; Immune system; In Vitro; Inhibitor of Differentiation Proteins; Kinetics; Lymphoid Cell; Maps; Molecular; Monitor; Mutate; Normal Cell; Oncogenes; Phase; Phenotype; Population Dynamics; Probability; Process; Proliferating; Property; Proto-Oncogenes; Rag1 Mouse; Regulation; Regulator Genes; Regulatory Element; Role; Site; T cell differentiation; T-Cell Development; T-Cell Receptor; T-Cell Receptor Genes; T-Lymphocyte; TCF3 gene; Testing; Thymus Gland; Work; dimer; gene network; gene product; gene regulatory network; genetic signature; genome-wide; in vivo; leukemia; network architecture; preservation; prevent; programs; protein complex; response; stem cells; transcription factor

PROJECT SUMMARY The progression of T cell precursors from multipotency to commitment occurs after multiple cell cycles in the thymus. These early, pre-commitment cell cycles are important for expansion of the precursors to generate enough pro-T cells to survive later selection events. However, the early stages are poorly understood, and in particular it has not been clear what controls the precise trajectory of the cells' differentiation nor the timing or irreversibility of commitment when it occurs. Genomic regulatory elements that are active after commitment tend to be characterized by motifs for basic helix-loop-helix (bHLH) E proteins, E2A or HEB, whereas those that are active before commitment have other signatures, suggesting that there is a major increase in E protein activity across this transition. In fact, the expression of E proteins is almost equally high before commitment, but our recent data show that they are occupied in substantially different roles before commitment, in complexes with other heterodimerization partners. This proposal is based on recent evidence that the alternative complex, containing Lmo2 and Lyl1 dimerized with E2A or HEB, may actually be a major controller of the commitment transition in early T cells. Our recent evidence shows that expression of Lmo2 and Lyl1 is sufficient to make committed pro-T cells reverse their differentiation in terms of gene expression. Not only does the Lmo2/Lyl1/E2A complex bind to different genomic sites than E protein dimers, but also the addition of Lmo2 and Lyl1 to committed pro-T cells is sufficient to remove E proteins from sites that they occupy after commitment and shift them to sites that are normally active only before commitment. The implication is that the kinetics of downregulation of Lmo2 and Lyl1 in normal T-cell differentiation could be vital for determining the timing of commitment and of the maturation of the pro-T cells. Lmo2 and Lyl1 have been considered as T- lineage proto-oncogenes, but our evidence suggests a potent role in normal development. This proposal is to determine the mechanism of how this works and to test its significance for the actual developmental dynamics of normal early T cells. Our preliminary work identifies the signature target genes affected by Lmo2+Lyl1 and their overlap with genes expressed in normal pro-T cells. We now propose to define: (1) the distinct molecular mechanisms that control different subsets of these signature genes, based on genome-wide mapping of the chromatin state changes caused by Lmo2+Lyl1/E protein complex binding as compared to pure E protein dimer binding; (2) whether endogenous Lmo2/Lyl1/E protein complexes indeed control differentiation kinetics and commitment of normal pro-T cells, based on acute CRISPR and monitoring in vitro and in vivo; and (3) the gene regulatory network architecture, involving factors regulated by Lmo2+Lyl1, through which Lmo2+Lyl1 exert their surprisingly broad impacts on T cell development. The results should determine whether and how a biochemical mechanism of transcription factor heterodimerization partner switching may explain a central unsolved problem in the dynamics of T cell development.

项目摘要 T细胞前体从多能性到定型的进展发生在淋巴细胞中的多个细胞周期之后。 胸腺这些早期的预定型细胞周期对于前体细胞的扩增是重要的， 足够的pro-T细胞在随后的选择事件中存活。然而，对早期阶段了解甚少， 特别地，尚不清楚是什么控制了细胞分化的精确轨迹，也不清楚细胞分化的时间或机制。 承诺发生时的不可逆转性。承诺后活跃的基因组调控元件 倾向于以碱性螺旋-环-螺旋（bHLH）E蛋白、E2 A或HEB的基序为特征，而那些 在承诺之前活跃的有其他的特征，这表明E蛋白有一个主要的增加， 在这一过渡过程中。事实上，E蛋白的表达在承诺之前几乎同样高， 但我们最近的数据显示，在承诺之前，他们扮演着截然不同的角色， 与其他异源二聚化伴侣的复合物。这一建议是基于最近的证据， 另一种复合物，含有Lmo 2和Lyl 1与E2 A或HEB二聚，实际上可能是一个主要的控制器 早期T细胞的定型转变我们最近的证据表明，Lmo 2和Lyl 1的表达是 足以使定向的pro-T细胞逆转其在基因表达方面的分化。不仅 Lmo 2/Lyl 1/E2 A复合物与E蛋白二聚体结合不同的基因组位点，而且Lmo 2/Lyl 1/E2 A复合物与E蛋白二聚体结合不同， Lmo 2和Lyl 1对定型pro-T细胞的作用足以将E蛋白从它们在活化后占据的位点移除。 承诺，并将其转移到通常只在承诺之前活跃的网站。言下之意是 正常T细胞分化中Lmo 2和Lyl 1下调的动力学对于确定 pro-T细胞的定型和成熟的时机。Lmo 2和Lyl 1被认为是T- 谱系原癌基因，但我们的证据表明，在正常发育中发挥着重要作用。 这个建议是为了确定这是如何工作的机制，并测试其实际意义 正常早期T细胞的发育动力学。我们的初步工作确定了标志性的靶基因 受Lmo 2 + Lyl 1的影响以及它们与正常pro-T细胞中表达的基因的重叠。我们现建议 定义：（1）控制这些特征基因的不同子集的不同分子机制，基于 Lmo 2 + Lyl 1/E蛋白复合物结合引起的染色质状态变化的全基因组作图， 与纯E蛋白二聚体结合相比;（2）内源性Lmo 2/Lyl 1/E蛋白复合物是否确实 基于急性CRISPR和监测，控制正常pro-T细胞的分化动力学和定型 体外和体内;和（3）基因调控网络结构，涉及由Lmo 2 + Lyl 1调控的因子， Lmo 2 + Lyl 1通过其对T细胞发育发挥其令人惊讶的广泛影响。结果应 确定是否以及如何一个转录因子异源二聚化伴侣的生化机制 转换可以解释T细胞发育动力学中一个未解决的核心问题。