RT Journal Article
SR Electronic
T1 Robust Sequence Determinants of α-Synuclein Toxicity in Yeast Implicate Membrane Binding
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2020.05.01.072884
DO 10.1101/2020.05.01.072884
A1 Robert W. Newberry
A1 Taylor Arhar
A1 Jean Costello
A1 George C. Hartoularos
A1 Alison M. Maxwell
A1 Zun Zar Chi Naing
A1 Maureen Pittman
A1 Nishith R. Reddy
A1 Daniel M. C. Schwarz
A1 Douglas R. Wassarman
A1 Taia S. Wu
A1 Daniel Barrero
A1 Christa Caggiano
A1 Adam Catching
A1 Taylor B. Cavazos
A1 Laurel S. Estes
A1 Bryan Faust
A1 Elissa A. Fink
A1 Miriam A. Goldman
A1 Yessica K. Gomez
A1 M. Grace Gordon
A1 Laura M. Gunsalus
A1 Nick Hoppe
A1 Maru Jaime-Garza
A1 Matthew C. Johnson
A1 Matthew G. Jones
A1 Andrew F. Kung
A1 Kyle E. Lopez
A1 Jared Lumpe
A1 Calla Martyn
A1 Elizabeth E. McCarthy
A1 Lakshmi E. Miller-Vedam
A1 Erik J. Navarro
A1 Aji Palar
A1 Jenna Pellegrino
A1 Wren Saylor
A1 Christina A. Stephens
A1 Jack Strickland
A1 Hayarpi Torosyan
A1 Stephanie A. Wankowicz
A1 Daniel R. Wong
A1 Garrett Wong
A1 Sy Redding
A1 Eric D. Chow
A1 William F. DeGrado
A1 Martin Kampmann
YR 2020
UL http://biorxiv.org/content/early/2020/07/11/2020.05.01.072884.abstract
AB Protein conformations are shaped by cellular environments, but how environmental changes alter the conformational landscapes of specific proteins in vivo remains largely uncharacterized, in part due to the challenge of probing protein structures in living cells. Here, we use deep mutational scanning to investigate how a toxic conformation of α-synuclein, a dynamic protein linked to Parkinson’s disease, responds to perturbations of cellular proteostasis. In the context of a course for graduate students in the UCSF Integrative Program in Quantitative Biology, we screened a comprehensive library of α-synuclein missense mutants in yeast cells treated with a variety of small molecules that perturb cellular processes linked to α-synuclein biology and pathobiology. We found that the conformation of α-synuclein previously shown to drive yeast toxicity—an extended, membrane-bound helix—is largely unaffected by these chemical perturbations, underscoring the importance of this conformational state as a driver of cellular toxicity. On the other hand, the chemical perturbations have a significant effect on the ability of mutations to suppress α-synuclein toxicity. Moreover, we find that sequence determinants of α-synuclein toxicity are well described by a simple structural model of the membrane-bound helix. This model predicts that α-synuclein penetrates the membrane to constant depth across its length but that membrane affinity decreases toward the C terminus, which is consistent with orthogonal biophysical measurements. Finally, we discuss how parallelized chemical genetics experiments can provide a robust framework for inquiry-based graduate coursework.Competing Interest StatementThe authors have declared no competing interest.