PTIP在转录调控与DNA损伤修复中的功能研究

Epigenetic control relies in part on Histone modifications, also termed as Histone code, which is apparently regulated in tissue and gene specific manners. However, the underlying mechanisms remain poorly understood. We and others have previously shown that PTIP is among essential elements for the proper Histone code formation. In addition, we have shown that PTIP mediates specifically the anterior/posterior patterning processes in flies. We have also demonstrated that PTIP may be involved in the regulation of up to a hundred of genes in Drosophila cells by means of microarray analysis (Fang et al., 2009). In this grant application, we propose to carry out genome-wide profiling of PTIP, as well as its known associated proteins, such as UTX, in staged fly embryos. Upon gain of information on the dynamics of PTIP in embryo development, we will make use of fly genetics to further study in detail one or two direct target genes of PTIP. On the other hand, since PTIP also associates with DNA damage response pathway, we plan to sample PTIP binding data by ChIP analysis in irradiated embryos. In combination with Histone modification analysis, we aim to understand how PTIP may discriminate in mediating different biological processes, such as transcription regulation and DNA damage response. Our hope is to present an example that Histone code dynamics is tightly regulated by epigenetic guider proteins, such as PTIP.

组蛋白编码（Histone Code）是表观遗传控制的分子基础之一，在发育过程中其形成具有组织和基因特异性的特点。然而，其分子机制并不十分清楚。我们前期研究发现PTIP对在特定发育过程的组蛋白编码形成中起重要作用，细胞中的Microarray分析表明，PTIP可能直接参与了百余Polycomb基因家族靶基因的调控（Fang et al., 2009）。本研究将以果蝇为模式动物，通过胚胎中PTIP与DNA相互作用的全基因组分析，并进一步用果蝇遗传学的方法深入研究PTIP在组蛋白修饰中的在体作用机制。另一方面，鉴于PTIP在DNA损伤修复中的必要作用，我们将观察辐射后胚胎中PTIP与DNA结合的动态变化，以阐明PTIP参与转录调控与DNA损伤修复这两个生物学过程的相互联系和功能区分。我们希望以此为例阐明组蛋白编码通过像PTIP这样的蛋白因子（我们称为向导蛋白）来实现精密调控。

本研究围绕PTIP基因功能研究，一方面特别阐明了果蝇PTIP在表观遗传中的在体作用和机制。我们利用Pc杂合体果蝇的表型，直接观察PTIP基因在Pc功能中的作用。我们发现PTIP基因剂量的减少显著增强Pc杂合表型，包括Sex comb transformation和成虫翅表型。表明在表观遗传情景，PTIP的功能作用方向与Pc一致，与我们之前研究发现PTIP拮抗trxG表型的结果相一致。我们进一步阐明了这种加强作用的基因基础是PTIP似乎与Pc共同阻遏Pc靶基因，特别是HOX家族的Scr和Ubx。这些发现首次表明了PTIP可能的基因阻遏作用。我们的研究还表明PTIP基因的缺失导致DNA损伤修复的障碍，表现为γH2A信号明显增强，更多的细胞凋亡和细胞增殖的减弱。我们针对果蝇PTIP蛋白的超长PolyQ区域对DNA损伤修复的作用进行了初步探索，我们发现以小鼠ptip（mPtip）替代果蝇ptip，显著延长果蝇的寿命,虽然程度有限。我们还进一步利用果蝇复眼的形态学改变，发现ptip和78Q之间存在遗传相互作用。综上，我们对PTIP在表观遗传和DNA修复两个方面的功能进行深入和广泛的探索研究，取得了大量令人鼓舞的研究结果。对于全面理解PTIP的基因功能具有重要意义。