Structural basis for the binding of SNAREs to the multisubunit tethering complex Dsl1

Abstract

Multisubunit tethering complexes (MTCs) are large (250 to >750 kDa), conserved macromolecular machines that are essential for SNARE-mediated membrane fusion in all eukaryotes. MTCs are thought to function as organizers of membrane trafficking, mediating the initial, long-range interaction between a vesicle and its target membrane and promoting the formation of membrane-bridging SNARE complexes. Previously, we reported the structure of the Dsl1 complex, the simplest known MTC, which is essential for COPI-mediated transport from the Golgi to the endoplasmic reticulum (ER). This structure suggested how the Dsl1 complex might function to tether a vesicle to its target membrane by binding at one end to the COPI coat and at the other end to ER SNAREs. Here, we use x-ray crystallography to investigate these Dsl1-SNARE interactions in greater detail. The Dsl1 complex comprises three subunits that together form a twolegged structure with a central hinge. Our results show that distal regions of each leg bind N-terminal Habc domains of the ER SNAREs Sec20 (a Qb-SNARE) and Use1 (a Qc-SNARE). The observed binding modes appear to anchor the Dsl1 complex to the ER target membrane while simultaneously ensuring that both SNAREs are in open conformations with their SNARE motifs available for assembly. The proximity of the two SNARE motifs, and therefore their ability to enter the same SNARE complex, depends on the relative orientation of the two Dsl1 legs.

Biochemical Characterization of the Num1-Mdm36 Complex at the Mitochondria-Plasma Membrane Contact Site

“…The binding affinity between Num1CC and Mdm36 is very similar to other tethering factors, and the affinity might be strong enough not to be fallen apart during mitochondrial fission. It should be noted that the binding affinity of the multisubunit tethering complex Dsl1 and soluble N-ethylmaleimide-sensitive factor attachment protein receptor was similar to that of the Num1CC-Mdm36 complex (Travis et al, 2020). Furthermore, the ITC experiment also indicated that the stoichiometry of Num1CC and Mdm36 was 1:1, which was consistent with the results previously anticipated by the SEC-MALS experiment (Fig.…”