Zinc finger methyl-CpG binding proteins: Bimodal regulators of cellular transcription

本研究的主要目的是了解细胞内基因表达调控的机制。DNA是遗传信息的长期存储单元，为细胞和生命功能提供指令。进入不同遗传区域的途径部分受DNA局部区域可逆化学变化的调控，这被定义为表观遗传修饰。这个项目评估了一个小家族的蛋白质ZBTB家族的作用，它在这些表观遗传修饰中很重要。该项目的广泛影响包括两个主要方面：首先，在一个跨学科研究项目中培训研究生，该项目利用广泛的技术来识别和表征三种ZBTB。其次，该项目包括在盐湖城市中心的莱昂纳多博物馆开展以生物化学和dna为重点的实践和探究活动。除了更广泛的公众，莱昂纳多容纳了大量来自盐湖城山谷学校中代表性不足和服务不足的群体的学生。众所周知，儿童对科学的兴趣最好在早期和易受影响的年龄灌输，并且预计动手活动的开发和实施将对该州在STEM领域的多样性增长产生长期的重大影响。DNA甲基化是表观遗传修饰的一种形式。虽然已经深入了解了DNA甲基化标记是如何建立和去除的，但对于甲基化cpg结合蛋白（MBPs）读取DNA甲基化标记并将该信息转化为特定转录结果的机制，特别是对于ZBTB MBPs，知之甚少。对ZBTB MBPs如何调节转录的理解是复杂的，因为它们表现出双峰DNA识别，靶向甲基化和序列特异性非甲基化DNA位点，这使它们能够同时作为转录抑制因子和激活因子。此外，每个ZBTB MBPs都可以与靶基因位点的多个协同抑制因子/激活因子蛋白复合物相互作用。因此，了解每一种蛋白质如何引起对其结合伙伴的选择性是辨别它们在转录调节中的作用的核心。为了研究这一点，将利用体外生物物理和细胞内结合的方法来描述MBPs家族识别其DNA和蛋白质目标的机制。总体目标是在结构上表征ZBTB蛋白：DNA相互作用以获得每种蛋白如何优先选择DNA靶标的机制见解；利用功能特异性突变探索双峰DNA识别的分子基础；并定义每个ZBTB MBP的蛋白质相互作用组。这项研究的结果将为识别ZBTB MBPs家族的每个成员引发选择性基因靶向，将特定蛋白质复合物招募到目标位点并改变转录以提供特定细胞功能的完整机制提供必要的基础。

The primary goal of this research is to understand the mechanisms by which gene expression is regulated within the cell. DNA is the long-term storage unit for genetic information, providing instructions for cellular and life functions. Access to different genetic regions is partially regulated by reversible chemical changes to localized regions of the DNA, defined as epigenetic modifications. This project evaluates the role of a small family of proteins, the ZBTB family, that are important in making these epigenetic modifications. The broader impacts for this project include two major aspects: First, the training of graduate students in an interdisciplinary research program that utilizes a broad spectrum of techniques to identify and characterize the three ZBTB. Second, this project includes the development of biochemical and DNA-focused hands-on and inquiry-based activities at The Leonardo, a museum in downtown Salt Lake City. In addition to the broader public, The Leonardo accommodates a significant number of students from underrepresented and underserved groups from Salt Lake Valley schools. It is well recognized that the interest of a child in science is best instilled at an early and impressionable age, and it is expected that development and implementation of hands-on activities will have a long-term significant impact on the state's diversity growth in STEM areas.DNA methylation is one form of epigenetic modification. While insight into how the DNA methylation mark is established and removed has been gained, there is only minimal knowledge for the mechanisms by which methyl-CpG binding proteins (MBPs),read DNA methylation marks and translate this information into a specific transcriptional outcome, particularly for the ZBTB MBPs. An understanding for how the ZBTB MBPs modulate transcription is complicated by the fact that they exhibit bimodal DNA recognition, targeting both methylated and sequence-specific non-methylated DNA sites, which affords them the capability of functioning as both transcriptional repressors and activators. Additionally, each of the ZBTB MBPs can interact with multiple co-repressor/-activator protein complexes at target gene sites. Thus, an understanding for how each of these proteins elicits selectivity for their binding partners is central to discerning their role in transcriptional regulation. To investigate this, a combined in vitro biophysical and in cell approach will be utilized to delineate the mechanisms by which this family of MBPs recognize their DNA and proteins targets. The overall goals are to structurally characterize ZBTB protein:DNA interactions to gain mechanistic insight into how each protein preferentially selects DNA targets; to utilize function-specific mutations to probe the molecular basis for bimodal DNA recognition; and to define protein interactomes for each ZBTB MBP. The outcomes of this research will provide an essential foundation for the larger goal of discerning the full mechanisms by which each member of this under studied family of ZBTB MBPs elicits selective gene targeting, recruits specific protein complexes to the target site and alters transcription to provide a particular cellular function.