SGER: Exploration of a Novel Nuclear Domain Tightly Associated with PML Bodies

这是一项探索性研究的小额资助。Lawrence博士将继续研究新的核“结构域”或“体”的最新初步证据，该结构域通常紧邻PML体，并与PML体形成“双重”结构。近年来，人们越来越认识到内部核结构的复杂性和重要性，越来越多的证据表明它们在各种核功能中发挥着根本作用。最前沿的是对PML体（也称为ND 10，POD或PML结构域）的强烈兴趣，因为这些结构富含重要的基因调控蛋白。 Lawrence博士的实验室一直在研究不同核区室的特征及其与基因表达的结构和功能关系。在她最近的工作过程中，她偶然发现了证据，表明存在一种基本上未知的和未表征的核“体”，它在结构上与PML体相关。这方面的证据还处于非常初步的状态，为了最终确定它是一个真正的核体，包含多个核因子，并具有明确的结构，还有一些重大问题需要解决。这项工作的潜在意义在于，它可能证明了细胞结构中一个以前未知的部分，相当于一个在基本核功能中具有未探索作用的核“细胞器”。此外，这些机构在结构上与一个机构，显然是在细胞周期调控牵连耦合。Lawrence博士和她的同事们最初发现了一种规则间隔的模式，约10-20个非常明亮的核“点”（直接邻接但不重叠PML体），使用的是一种针对FLAG表位的抗体，这些抗体被设计成在生理水平上表达标记有flag的c-myc癌蛋白。然而，最近的证据表明，检测到的抗原可能不是myc癌蛋白，而是一种内源性蛋白，可能是对MuLV病毒（鼠白血病病毒）的细胞反应的一部分。第一个目的是调查的假设，PML相关机构确实存在，并发现在各种细胞，使用超微结构分析和免疫荧光其他几个特定的蛋白质，以前的证据表明，可能共同定位于这些机构。其次，她将研究MuLV载体或MuLV表达与诱导相关蛋白和/或体之间的关系。 根据结果，还可以鉴定由抗flag抗体检测并定位于这些结构的未知细胞蛋白。

This is a Small Grant for Exploratory Research.Dr. Lawrence will pursue recent preliminary evidence of a novel nuclear "domain" or "body," which typically positions immediately adjacent to PML bodies and appears to form a "doublet" structure with the PML body. In recent years, there has been increasing appreciation for the complexity and importance of internal nuclear structures and growing evidence of their fundamental roles in diverse nuclear functions. At the forefront has been intense interest in PML bodies (also called ND10, PODs, or PML domains), because these structures are enriched for a collection of important gene regulatory proteins. Dr. Lawrence's laboratory has been studying the characterization of distinct nuclear compartments and their structural and functional relationship to gene expression. In the course of her very recent work she fortuitously uncovered evidence that there exists an essentially unknown and uncharacterized nuclear "body" that is structurally linked to PML bodies. The evidence for this is at a very preliminary state, and there are major questions to be sorted out in order to establish conclusively the existence of this as a bona fide nuclear body containing multiple nuclear factors and having defined structure. The potential significance of the work is that it may demonstrate a previously unknown part of cell structure, the equivalent of a nuclear "organelle" with an unexplored role in basic nuclear function. In addition, these bodies are structurally coupled with a body that is clearly implicated in cell cycle regulation. Dr. Lawrence and her colleagues initially discovered a regularly spaced pattern of ~10-20 very bright nuclear "dots" (immediately abutting but not overlapping PML bodies) using an antibody to the FLAG-epitope in cells engineered to express flag-tagged c-myc oncoprotein at physiological levels. However, recent evidence suggests that the antigen detected is probably not the myc oncoprotein but rather an endogenous protein, possibly induced as part of a cellular reaction to MuLV virus (murine leukemia virus). The first aim is to investigate the hypothesis that PML-associated bodies indeed exist and are found in a variety of cells, using both ultrastructural analysis and immunofluorescence to several other specific proteins that prior evidence suggests may co-localize to these bodies. Second, she will investigate the relationship between MuLV vectors or MuLV expression and the induction of the relevant protein and/or body. Depending on results, it would also be feasible to identify an unknown cellular protein detected by the anti-flag antibodies and localized to these structures.