Unraveling the Mechanisms of Protein Sorting and Localization to Organelles

PROJECT SUMMARY Protein import to subcellular compartments is fundamental to maintaining cellular and organelle proteostasis. The sorting of nascent proteins begins during their synthesis on ribosomes and is mediated by molecular “zip codes” encoded within their primary sequence. In the subsequent steps, a network of chaperones and receptors mediate protein processing and folding during import on the respective organelle membrane. Disruptions in these pathways can lead to protein misfolding and aggregation, contributing to the onset of neurodegenerative diseases such as Alzheimer’s and Parkinson’s. Furthermore, defects in protein import are linked to Congenital Disorders of Glycosylation (CDG), highlighting the critical health implications of our research focus. My lab aims to uncover the molecular mechanisms for protein biogenesis on mitochondrial and endoplasmic reticulum (ER) membranes. The two ambitious projects in my research program will explore: 1) how nascent polypeptide chains are processed and folded by chaperones during transport across the ER membrane, and 2) the molecular basis of mitochondrial protein import. Here, we will employ a set of innovative and integrative approaches such as cell-free protein synthesis and cell-based assays to trap ribosome-nascent chain complexes at different stages during protein import. We will then employ high-resolution cryo-electron microscopy and cellular cryo-electron tomography to determine the structures of these ribosome complexes with associated chaperones and receptors on the ER and mitochondrial membranes to delineate their molecular interactions at atomic detail. Finally, we will conduct functional studies, both in-vitro and using cell-based assays, to complement and validate our structural observations. Our research program is uniquely positioned to fill critical gaps in the understanding of co-translational protein sorting and processing on organelle membranes. The anticipated outcomes will provide molecular snapshots of these processes with novel insights and implications for therapeutic strategies targeting protein misfolding and aggregation in disease.