Nde1 promotes Lis1-mediated activation of dynein.

Cytoplasmic dynein drives the motility and force generation functions towards the microtubule minus end. The assembly of dynein with dynactin and a cargo adaptor in an active transport complex is facilitated by Lis1 and Nde1/Ndel1. Recent studies proposed that Lis1 relieves dynein from its autoinhibited conformation, but the physiological function of Nde1/Ndel1 remains elusive. Here, we investigate how human Nde1 and Lis1 regulate the assembly and subsequent motility of mammalian dynein using in vitro reconstitution and single molecule imaging. We find that Nde1 recruits Lis1 to autoinhibited dynein and promotes Lis1-mediated assembly of dynein-dynactin adaptor complexes. Nde1 can compete with the α2 subunit of platelet activator factor acetylhydrolase 1B (PAF-AH1B) for the binding of Lis1, which suggests that Nde1 may disrupt PAF-AH1B recruitment of Lis1 as a noncatalytic subunit, thus promoting Lis1 binding to dynein. Before the initiation of motility, the association of dynactin with dynein triggers the dissociation of Nde1 from dynein by competing against Nde1 binding to the dynein intermediate chain. Our results provide a mechanistic explanation for how Nde1 and Lis1 synergistically activate the dynein transport machinery.

Nde1 Promotes Lis1-Mediated Activation of Dynein.

Cytoplasmic dynein is the primary motor that drives the motility and force generation functions towards the microtubule minus end. The activation of dynein motility requires its assembly with dynactin and a cargo adaptor. This process is facilitated by two dynein-associated factors, Lis1 and Nde1/Ndel1. Recent studies proposed that Lis1 rescues dynein from its autoinhibited conformation, but the physiological function of Nde1/Ndel1 remains elusive. Here, we investigated how human Nde1 and Lis1 regulate the assembly and subsequent motility of the mammalian dynein/dynactin complex using in vitro reconstitution and single molecule imaging. We found that Nde1 promotes the assembly of active dynein complexes in two distinct ways. Nde1 competes with the α2 subunit of platelet activator factor acetylhydrolase (PAF-AH) 1B, which recruits Lis1 as a noncatalytic subunit and prevents its binding to dynein. Second, Nde1 recruits Lis1 to autoinhibited dynein and promotes Lis1-mediated assembly of dynein-dynactin-adaptor complexes. However, excess Nde1 inhibits dynein, presumably by competing against dynactin to bind the dynein intermediate chain. The association of dynactin with dynein triggers Nde1 dissociation before the initiation of dynein motility. Our results provide a mechanistic explanation for how Nde1 and Lis1 synergistically activate the dynein transport machinery.