Testing the 'Occlusis' Model of Cell Fate Restriction

Type of Award: Catalyst
Date Awarded: August 2008
Award End Date: July 2010
Amount Awarded: $ 200,000.00
PI(s): Bruce T. Lahn, PhD, UChicago; John A. Kessler, MD, NU;

Abstract: A gene's transcriptional output is the combined product of two inputs: diffusible factors in the cellular milieu acting in trans, and the biochemical state of chromatin acting in cis. Although many studies in the field of epigenetics have pointed to the possibility that genes could be silenced by cis-acting, chromatin-based mechanisms, there is as yet no ready experimental system for ascertaining whether the silent state of a gene is indeed the result of cis rather than trans regulation. To address this problem, we have recently developed a cell fusion strategy with which it is possible to dissect out the relative contribution of cis versus trans mechanisms to gene silencing. The strategy entails fusing two disparate cell types and searching for genes differentially expressed between the two genomes of fused cells. Any differential expression can be causally attributed to cis mechanisms because the two genomes of fused cells share a single homogenized milieu. Using this strategy, we uncovered the presence of many 'occluded' genes in the genome -- defined as genes existing in a state of transcriptional competency whereby they are silenced by cis-acting mechanisms in a manner that blocks them from responding to the trans-acting milieu of the cell. Here, we propose to further develop the cell-fusion-based approach for identifying and analyzing occluded genes. Specifically, we intend to pursue the following aims: (1) to test whether the occluded state of a gene, once acquired during development, is essentially irreversible; (2) to explore the biochemical mechanisms underlying gene occlusion.

The identification and analysis of occluded genes is likely to have important implications for many fields of biology. The following are some examples: (1) knowing the competent/occluded status of genes could be central to studies in systems biology aimed at producing comprehensive circuit diagrams of the regulatory networks within cells; (2) the occlusis model could establish an important theoretical framework for studies of developmental biology and stem cell biology, especially the mechanisms underlying cell fate restriction; (3) genome-wide maps of occluded genes might offer a more fundamental molecular definition of cell type identity; (4) the description of competent/occluded status of genes provides a novel functional readout of the genome with which the biological significance of chromatin signatures such as DNA methylation and histone modifications could be better interpreted; and (5) disruption of the competent/occluded status of genes by environmental, genetic or stochastic factors might contribute to aging and disease processes such as cancer. For these reasons, we believe that the proposed work has the potential to make a significant impact on a broad swath of biomedical research.