Combinatorial regulation of the balance between dynein microtubule end accumulation and initiation of directed motility

Cytoplasmic dynein is involved in a multitude of essential cellular functions. Dynein's activity is controlled by the combinatorial action of several regulatory proteins. The molecular mechanism of this regulation is still poorly understood. Using purified proteins, we reconstitute the regulation of the human dynein complex by three prominent regulators on dynamic microtubules in the presence of end binding proteins (EBs). We find that dynein can be in biochemically and functionally distinct pools: either tracking dynamic microtubule plus‐ends in an EB‐dependent manner or moving processively towards minus ends in an adaptor protein‐dependent manner. Whereas both dynein pools share the dynactin complex, they have opposite preferences for binding other regulators, either the adaptor protein Bicaudal‐D2 (BicD2) or the multifunctional regulator Lissencephaly‐1 (Lis1). BicD2 and Lis1 together control the overall efficiency of motility initiation. Remarkably, dynactin can bias motility initiation locally from microtubule plus ends by autonomous plus‐end recognition. This bias is further enhanced by EBs and Lis1. Our study provides insight into the mechanism of dynein regulation by dissecting the distinct functional contributions of the individual members of a dynein regulatory network. Synopsis The balance between processive minus‐end directed motility and microtubule plus‐end localisation of dynein is controlled by various regulatory proteins. In vitro reconstitutions of the human dynein complex in the presence of end binding proteins reveal the differential control of dynein activity on dynamic microtubules by a combination of its key regulators. Processively moving and plus end tracking dynein pools both contain dynactin, but show otherwise distinct preferences for binding regulators. Dynactin recruits dynein to microtubule plus ends in the presence, and—to a lesser extent—also in the absence of EB1. BicD2 is necessary for initiating processive minus‐end directed motility of the dynein/dynactin complex. Dynactin causes a bias of dynein motility initiation from microtubule plus ends. Lis1 has a dual role in the initiation of dynein/dynactin processive motility and microtubule plus end recruitment. Graphical Abstract In vitro reconstitution of the human dynein complex reveals the differential control of dynein activity on dynamic microtubules by a combination of its key regulators.