Gene Expression in Cancer by Microarray Hybridization

描述(由申请人提供)：从一开始，该计划的目标就是开发和应用全基因组方法来系统和定量分析癌症中的基因表达模式。在我们开始这个项目的八年里，在转录本丰度水平上系统研究全球基因表达模式的实验和分析工具取得了巨大的发展，它们已经成为广泛可用的工具，并被广泛用于癌症研究。由此产生的研究产生了对癌症的丰富的新见解，甚至开始影响患者的护理。然而，分析mRNA转录水平只提供了全球基因表达计划的部分图景。虽然有大量证据表明，对翻译、亚细胞定位和mRNA衰退的调控是生物调控的关键因素，但研究这些调控水平的实用和强大的全基因组方法仍有待开发。因此，我们对这些调控机制的系统架构、分子机制和生物学作用的了解，包括它们在人类癌症中所起的作用，几乎还处于起步阶段。因此，我们建议开发实用的、健壮的、高通量的方法，使用DNA微阵列在转录后水平描述全球调控的三个关键方面：翻译调控、mRNA降解的调控以及RNA结合蛋白与其靶标的特定相互作用。这些研究将建立在我们已经在酿酒酵母模型系统中开始的初步研究基础上，但将重点放在人类细胞上。随着基本方法论的发展，我们将应用它们来建立一个基本的知识框架，调查这些调节机制在基本生理和发育程序中使用的模式，它们如何在一种细胞类型到另一种细胞类型以及不同个体之间变化，并开始研究潜在的分子机制。我们的目标是发展实验方法和解释框架，以使这些转录后基因表达水平可以在全基因组范围内被系统地描述和研究，几乎就像今天的转录水平一样。

DESCRIPTION (provided by applicant): From the start, the goal of this program has been the development and application of genome-wide approaches to systematic and quantitative analysis of gene expression patterns in cancer. In the eight years since we began this project, the experimental and analytical tools for systematic studies of global gene expression patterns at the level of transcript abundance have developed dramatically and they have become widely available and widely used for the study of cancer. The resulting studies have produced a wealth of new insight into cancer and they are even beginning to influence patient care. Profiling mRNA transcript levels, however, provides only a partial picture of the global gene expression program. While there is abundant evidence that regulation of the translation, subcellular localization and decay of mRNA are critical elements of biological regulation, practical and robust genome-wide approaches to studying these levels of regulation remain to be developed. As a result, our knowledge of the systems architecture, molecular mechanisms and biological roles of these regulatory mechanisms, including the roles they play in human cancer, is barely in its infancy. We therefore propose to develop practical, robust, high-throughput methods using DNA microarrays to profile three critical aspects of global regulation at the post-transcriptional level: translational regulation, regulation of mRNA degradation, and the specific interactions of RNA binding proteins with their targets. These studies will build on preliminary studies we've already begun in the Saccharomyces cerevisiae model system, but will focus on human cells. As the essential methodologies are developed, we will apply them to developing a foundational framework of knowledge, investigating the patterns in which these regulatory mechanisms are used in basic physiological and developmental programs, how they vary from one cell type to another and between individuals, and beginning to investigate the underlying molecular mechanisms. The goal is to develop both the experimental methodology and an interpretive framework to the point that these post-transcriptional levels of gene expression can be systematically profiled and studied on a genome-wide scale almost as routinely as transcript levels are today.