Determining gene targets and pathways for primate-specific zinc finger proteins

描述（由申请人提供）：Kruppel型锌指（ZNF）基因座包括所有人类基因家族中最大的基因之一。在哺乳动物中，大多数ZNF基因是单一亚型，编码的蛋白质中，DMA结合锌指阵列连接到染色质相互作用域，称为KRAB，赋予一个强大的转录抑制活性。虽然某些KRAB-ZNF基因座是高度保守的，但正在进行的片段重复事件在每个哺乳动物谱系中产生了很大程度上独特的基因组。在404个人类KRAB-ZNF基因座中，超过三分之一是灵长类动物特异性的，即使是最近的重复基因也以指示选择具有不同DNA识别位点的新蛋白质的方式出现分歧。这个单一的基因家族包括所有预测的人类转录因子基因座的五分之一，现有的数据表明，在调节过程中发挥着广泛的作用，对人类健康至关重要。然而，由于调控目标和途径是已知的，只有少数KRAB-ZNF蛋白的功能，大多数家庭成员仍然是一个猜测的问题。我们假设这个动态基因家族在塑造人类健康相关生物学方面发挥了重要作用，包括高度保守和灵长类动物特异性特征。拟议的研究计划旨在通过对25个KRAB-ZNF基因的功能分析来解决这一假设。作为一个主要的重点，我们将分析蛋白质编码的25个基因参与最近的灵长类动物特异性复制，重点是旁系同源蛋白质，最适合实验分析。具体来说，我们将（1）通过操纵人类细胞中的基因表达来确定由每种ZNF蛋白调控的基因和途径;（2）使用染色质免疫沉淀技术来定义ZNF蛋白在人类染色质中结合的基因组区域;（3）使用生物信息学和实验方法相结合来识别和验证定义每种蛋白的有利识别位点的共有DNA基序。这项研究将首次深入研究这个人类转录抑制因子大家族的功能，使我们能够研究它们在调节人类免疫，生殖，发育，癌症易感性和其他涉及KRAB-ZNF基因的过程中的作用。我们产生的数据将提供新的指导方针，以预测其他家庭成员的功能，并评估KRAB-ZNF基因的获得，丢失，突变和失调对人类健康的潜在影响。

DESCRIPTION (provided by applicant): Kruppel-type zinc finger (ZNF) loci comprise one of the largest of all human gene families. In mammals, the majority of ZNF genes are of a single subtype, encoding proteins in which DMA-binding zinc finger arrays are attached to a chromatin-interacting domain, called KRAB, that confers a potent transcriptional repressor activity. Although certain KRAB-ZNF loci are highly conserved, ongoing segmental duplication events have created largely unique gene sets in each mammalian lineage. More than one-third of the 404 human KRAB- ZNF loci are primate-specific, and even the most recent duplicate genes have diverged in ways that indicate a selection for novel proteins with distinct DNA recognition sites. This single gene family comprises one-fifth of all predicted human transcription factor loci, and available data suggest broad roles in regulating processes that are critical to human health. However, since regulatory targets and pathways are known for only a handful of KRAB-ZNF proteins the functions of most family members remain a matter of conjecture. We hypothesize that this dynamic gene family has played a significant role in shaping human health-related biology, including both deeply conserved and primate-specific traits. The proposed research program is designed to address this hypothesis through functional analysis of 25 KRAB-ZNF genes. As a primary focus we will analyze proteins encoded by 25 genes involved in the most recent primate-specific duplications with emphasis on paralogous proteins that are most amenable to experimental analysis. Specifically, we will (1) determine genes and pathways that are regulated by each ZNF protein by manipulating gene expression in human cells; (2) define genomic regions to which the ZNF proteins bind in human chromatin using chromatin immunoprecipitation techniques; and (3) identify and validate consensus DNA motifs defining favored recognition sites for each protein using combined bioinformatics and experimental approaches. This study will provide a first in-depth look at functions for this large family of human transcriptional repressors, permitting us to examine their roles in regulating in human immunity, reproduction, development, cancer susceptibility and other processes in which KRAB-ZNF genes have been implicated. The data we generate will provide new guidelines to predict the functions of additional family members and to assess the potential impact of the gain, loss, mutation and dsyregulation of KRAB-ZNF genes on human health.