RT Journal Article
SR Electronic
T1 Improved mammalian retromer cryo-EM structures reveal a new assembly interface
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2022.03.04.482375
DO 10.1101/2022.03.04.482375
A1 Amy K. Kendall
A1 Mintu Chandra
A1 Boyang Xie
A1 William Wan
A1 Lauren P. Jackson
YR 2022
UL http://biorxiv.org/content/early/2022/03/04/2022.03.04.482375.abstract
AB Retromer (VPS26/VPS35/VPS29 subunits) assembles with multiple sorting nexin (SNX) proteins on membranes to mediate endosomal recycling of transmembrane protein cargoes. Retromer has been implicated in other cellular events, including mitochondrial homeostasis, nutrient sensing, autophagy, and fission events. Mechanisms for mammalian retromer assembly remain undefined, and retromer engages multiple sorting nexin proteins to sort cargoes to different destinations. Published structures demonstrate mammalian retromer forms oligomers in vitro, but several structures were poorly resolved. We report here improved retromer oligomer structures using single particle cryo-electron microscopy (cryo-EM) by combining data collected from tilted specimens with multiple improvements in data processing, including using a three-dimensional (3D) starting model for improved automated particle picking in RELION. A retromer mutant (3KE retromer) that breaks VPS35-mediated interfaces was used to determine a structure of a new assembly interface formed by the VPS26A and VPS35 N-termini. The interface reveals how an N-terminal VPS26A arrestin saddle can link retromer chains by engaging a neighboring VPS35 N-terminus, on the opposite side from the well-characterized C-VPS26/N-VPS35 interaction observed within heterotrimers. The new interaction interface exhibits substantial buried surface area (∼7,000 Å2) and further suggests metazoan retromer may serve as an adaptable scaffold.Competing Interest StatementThe authors have declared no competing interest.