Epigenetics and Proteostasis in Trafficking and Misfolding Disease

William E. Balch

W.E. Balch, *S. Becker, M. Bouchecareilh , **J. Hulleman, D. Hutt, V. Gupta, K. Routledge,
J. Matteson, A. Murray, A. Nauli, L. Page, ***S. Pankow, H. Plutner,
A. Pottekat, A. Razvi, D. Roth, L. Ryno, K. J. Singh, K. Subramanian

* Joint Manning/Balch Labs
** Joint Kelly/Balch Labs
*** Joint Yates/Balch Labs

A major challenge in human health is to understand and treat the many conformational diseases that affect protein homeostasis (referred to as proteostasis) (Balch et al. (2008) Science 319:916; Powers et al. (2009) Ann. Rev. Biochem. Epub) during development and aging. These diseases arise as a consequence of an imbalance between the energetics of the protein fold and the properties of the local folding environment that has both genetic and epigenetic foundations. The folding machinery involves numerous chaperone systems such as Hsp70 and Hsp90 that both direct folding and protect the fold from stressors that assault human physiology. Loss of proteostasis leads to major diseases including, among others, type 2 diabetes, emphysema, multiple amyloidoses including Alzheimers and systemic (light chain) myeloma disease, and cystic fibrosis. These diseases are broadly classified as membrane trafficking disease because a defect in folding during transit through the mammalian exocytic and/or endocytic trafficking pathways leads to loss of normal function and/or a gain-of-toxic function. Key goals of the laboratory are to (1) define the operation of these trafficking pathways, (2) determine the cause of the underlying folding disorders and, (3) learn how these events can be altered pharmacologically to restore the ability of the protein to function in the cell.

Structural basis for misfolding and membrane traffic

Eukaryotic cells are highly compartmentalized. Each compartment of the exocytic and endocytic pathways provides a unique chemical and biological environment in which protein folding and function can be modulated to maintain cellular proteostasis. During export from the first compartment of the exocytic pathway, the endoplasmic reticulum (ER) where folding is initiated, nearly one-third of the protein cargo encoded by the human genome is mobilized to the rest of the cell by the activity vesicle budding machines that utilize tethering/fusion and coat components to direct membrane traffic. In collaboration with C. Potter and B. Carragher (Cell Biology), we have used cryo-electron microscopy (cryoEM) to solve the structure of the COPII coat- a self-assembling scaffold that contains an outer cage polymeric lattice with an unprecedented iscosidecahedron geometry and an inner, asymmetrically disposed tetrameric adaptor complexes responsible for collecting newly synthesized and folded protein cargo into the emerging budding vesicle. To understand the mechanism of tethering and fusion to downstream compartments, we have solved the x-ray structure of the COPII tether complex in collaboration with I.A. Wilson (Molecular Biology). The structure reveals a superhelical platform based on a tripod motif that directs functional interaction with regulatory GTPases (Fig. 1- p115/tether complex). Surprisingly, we have found that the assembly and disassembly of such tethering systems is likely biologically regulated by the activity of the Hsp90 family of chaperone/co-chaperone components. These results suggest that extensive rearrangements to the protein fold are necessary to accomplish docking and fusion. Using bioinformatics and systems biology approaches, we are beginning to define how networks likely integrate the folding and trafficking networks to achieve the overall structure and function of membrane compartments.

Epigenetics and proteostasis in misfolding disease

Many mutations disrupt cargo traffic from the ER by preventing proper protein folding during synthesis in the ER resulting in loss of recognition by the COPII machinery. In collaboration with J. Kelly (Chemistry) and E. Powers (Chemistry) we are using modeling approaches to develop a rigorous quantitative framework to describe in a global way the adaptable role of proteostasis in health and disease. In collaboration with J. Yates (Chemical Physiology), we have developed and used novel mass spectrometry techniques to analyze the proteome involving Hsp70 and Hsp90 chaperone system that regulates the trafficking and function of the cystic fibrosis transmembrane conductance regulator (CFTR), a chloride channel that when defective is responsible for the disease cystic fibrosis (CF). Working with the Gottesfeld laboratory, we have discovered that manipulation of the chromatin environment (the epigenome) through modulation of histone deacetylases (HDACs) using HDAC inhibitors (HDACi) or siRNA silencing of HDAC enzymes modulates a global network of interacting factors that contribute to restoration of the function of the CFTR in disease. These results suggest that transcriptional regulatory circuits controlled by gene silencing/unsilencing pathways may allow us to reprogram the disease state and are consistent with the effects of caloric intake and epigenetic modifiers that also influence the onset of other conformational diseases including type 2 diabetes and neurodegenerative amyloid diseases of aging that we are studying with our colleagues J. Kelly (Chemistry), J. Buxbaum (Experimental Medicine) and A. Dillin (Salk).

Through such a multidisciplinary approach, we hope to gain critical insight into the fundamental principles of protein folding and trafficking, and a new understanding of the role of the proteostasis and epigenetic environments in controlling human health and aging. We anticipate that knowledge of these pathways will enable the development of small-molecule proteostasis regulators that adjust the folding environment that restore function.