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  • Cryo-EM Reveals How Human Cytoplasmic Dynein Is Auto-inhibited and Activated.

Cryo-EM Reveals How Human Cytoplasmic Dynein Is Auto-inhibited and Activated.

Cell (2017-06-13)
Kai Zhang, Helen E Foster, Arnaud Rondelet, Samuel E Lacey, Nadia Bahi-Buisson, Alexander W Bird, Andrew P Carter
ABSTRACT

Cytoplasmic dynein-1 binds dynactin and cargo adaptor proteins to form a transport machine capable of long-distance processive movement along microtubules. However, it is unclear why dynein-1 moves poorly on its own or how it is activated by dynactin. Here, we present a cryoelectron microscopy structure of the complete 1.4-megadalton human dynein-1 complex in an inhibited state known as the phi-particle. We reveal the 3D structure of the cargo binding dynein tail and show how self-dimerization of the motor domains locks them in a conformation with low microtubule affinity. Disrupting motor dimerization with structure-based mutagenesis drives dynein-1 into an open form with higher affinity for both microtubules and dynactin. We find the open form is also inhibited for movement and that dynactin relieves this by reorienting the motor domains to interact correctly with microtubules. Our model explains how dynactin binding to the dynein-1 tail directly stimulates its motor activity.

MATERIALS
Product Number
Brand
Product Description

Millipore
CellASIC ONIX switching plate mammalian cells (4 chamber), The M04 switching plates enable continuous perfusion culture for live cell analysis. Laminar flow provides rapid & uniform solution exchange for maximum cell health. The glass coverslide bottom surface ensures the highest resolution optical viewing.
Sigma-Aldrich
Anti-α-Tubulin antibody, Mouse monoclonal, clone DM1A, purified from hybridoma cell culture
Sigma-Aldrich
Anti-Fas Antibody (human, neutralizing), clone ZB4, clone ZB4, Upstate®, from mouse