CAREER: Discovering Rhomboid Dynamics and Function in Membrane Protein Homeostasis

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The complex biochemical processes occurring in biological membranes are fascinating and essential for life. Most of these processes are carried out by proteins embedded in the membranes surrounding the cells and organelles inside; these proteins comprise one third of total proteins in plants and animals. Unfortunately, misfolding of membrane proteins is a common occurrence, either due to chemical and UV damage, aging, or genetic mutations. To survive the constant, ongoing threat of protein misfolding, organisms are equipped with quality-control systems that detect and degrade these molecules. Despite intense study on these pathways, the quality-control mechanisms for removing misfolded membrane proteins have remained surprisingly elusive for over two decades. This project seeks to advance our knowledge of the critical and widely conserved process of the quality-control system(s) that maintains functional and folded membrane proteins. To attain this scientific goal, this project includes a significant component of URM education, outreach, and training to community college students. Efforts of this project focuses on: 1) providing an iBiology Spotlight lecture to expose community college students to hands-on research, role models and careers in science; 2) incorporating and developing Course-based Research Experience modules at Miramar Community College in coordination with instructors; and 3) direct involvement of research projects in the Neal laboratory at UCSD. Fulfillment of this research and educational plan will build and sustain interest in science, in particular for persistently underrepresented STEM students currently learning at community colleges.Endoplasmic Reticulum-Associated Degradation (ERAD) describes a set of conserved degradation pathways that remove misfolded proteins from the ER. Retrotranslocation is a universal feature of all ERAD pathways, entailing complete removal of misfolded substrates to the cytosol for degradation. For integral membrane substrates, this involves complete extraction of the protein, including all transmembrane spans, from the ER membrane. Despite its commonality in ERAD, retrotranslocation of integral membrane substrates has remained a mystery. This project builds on Dr. Neal’s discovery of a yeast rhomboid pseudoprotease Dfm1, which is a dedicated protein export factor for removing deleterious membrane proteins from the ER. The underlying hypothesis of this project is that Dfm1 and related rhomboid superfamily proteins are central mediators for removing aberrant membrane substrates, and loss of their function leads to profound cellular stress. Accordingly, the primary objectives to investigate this hypothesis are: 1) To understand the mechanism associated with Dfm1-mediated retrotranslocation function. This objective will employ genetic, biochemical and biophysical approaches using a tractable model system, S. cerevisiae, for deep characterization of Dfm1’s retrotranslocation function. 2) Explore a newly identified stress response pathway associated with loss of Dfm1 function. This objective will use functional genomics, biochemical, and genetic approaches to define an entirely new sensing system for membrane proteins. 3) Extend acquired understanding of yeast Dfm1 function to higher eukaryotes. This objective will determine the extent by which Dfm1’s molecular actions and functions are conserved in the related rhomboid proteins in human cells. Overall, results emerging from this work will reveal novel mechanisms for safeguarding the membrane proteome.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.