Mechanisms of Gene Silencing

点击翻译按钮获取中文摘要

DESCRIPTION (provided by applicant): Complex processes such as development and disease progression are regulated by gene expression. The majority of gene expression is controlled through chromatin packaging. Heterochromatin Protein 1 (HP1) is an evolutionary conserved non-histone chromosomal protein required for chromatin packaging and gene expression. HP1 shows a non-uniform distribution on chromosomes, being enriched within centric heterochromatin and also localizing to telomeres and euchromatic sites. The long-term goal of this proposal is to determine the mechanisms of gene regulation by HP1 in a variety of chromatin contexts. HP1 exhibits the property of "spreading" along the chromosome and silencing genes. In Specific Aim 1, we propose HP1 tethering experiments and genetic screens in Drosophila to determine the mechanism of silent chromatin spreading. Recently, HP1 has been linked to the activation of both heterochromatin and euchromatic genes. In Specific Aim 2, we propose to identify the sequences required for HP1 targeting. In addition, we will determine the role of HP1 in the expression of heterochromatic genes that normally reside in a repressive chromatin environment. Knowledge of HP1 function obtained from studies in Drosophila has important implications for breast cancer progression. We have shown that HP1Hsa, 1 of 3 HP1 family members in humans, is down-regulated in metastatic breast cancer cell lines and metastatic clinical samples. Modulation of HP1Hsa levels alters the invasive potential of breast cancer cells. Therefore, we hypothesize that HP1Hsa functions to regulate genes required for invasion and metastasis. In Specific Aim 3, we will determine which genes are mis-regulated following knock-down of HP1Hsa levels and identify direct target genes by chromatin immunoprecipitation. The mechanism of HP1Hsa localization and function at such genes will be determined by a knock-down and replacement strategy where endogenous HP1Hsa is exchanged for mutant forms lacking specific functions. Collectively, these studies will expand our knowledge about the mechanisms of gene regulation by HP1 proteins and identify the molecular pathways that regulate metastasis. Relevance to Public Heath: Studies described here will identify the genes and molecular pathways involved in regulating breast cancer invasion/metastasis. Protein products encoded by such genes could represent new markers for metastatic progression and targets for therapy.