Structural biology of chromatin in vitro and in cells

University Of California, San Diego, La Jolla CA

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Abstract

Project Summary The laboratory led by PI Galia Debelouchina has the following long-term objectives: 1) The development of structural biology methodology to study complex and dynamic biological assemblies in vitro, and 2) The extension of these methodologies for structural biology investigations in the cellular environment. Our methodology development combines solid-state nuclear magnetic resonance (NMR) spectroscopy with state- of-the-art chemical biology tools for the comprehensive description of biological systems both in vitro and in cells. Over the next five years, we plan to accomplish the following goals: 1) Elucidate the molecular basis of heterochromatin formation and regulation. Heterochromatin compartments are associated with gene silencing and repetitive DNA sequences, and their formation is a vital step in cell differentiation and development. Recent hypotheses suggest that they are formed through a process called liquid-liquid phase separation and that a central player in this process is the heterochromatin protein 1 (HP1) family. Our goal is to understand how HP1 proteins orchestrate a complex network of interactions with each other, with chromatin, and with other protein binding partners to regulate the material properties of heterochromatin environments and their implications for gene silencing. We will tackle this goal using NMR spectroscopy, chemical biology, cell biology and computational tools to obtain a comprehensive molecular view of heterochromatin interactions. In the process, we also hope to develop new NMR-based tools to characterize dynamic and heterogenous biological systems such as those formed by HP1 proteins. 2) Development of NMR-based tools for structural biology in cells. Here, we plan to focus on the development and implementation of a technique called targeted dynamic nuclear polarization (DNP), which allows us to zoom in on a specific protein in the cellular environment and to increase its NMR signals selectively over the cellular background. We plan to apply this technique to study protein-membrane interactions in whole bacterial and mammalian cells. Ultimately, we aim to develop targeted DNP as a tool to build an atomic resolution picture of the cellular milieu and to investigate changes in protein structure and dynamics in health and disease.