ABSTRACT
Dynein was first isolated from Tetrahymena cilia nearly four decades ago (1,2). This discovery of a microtubule motor together with the realization that dynein produces directed movements along microtubules revolutionized the thinking about how cells achieve microtubule-based movements (3-5).Dynein is the high molecular weight microtubule molecular motor that powers ciliary beating and nonaxonemal movements (reviewed in Refs. 6-8). A dynein particle transduces the free energy of ATP hydrolysis into mechanical force that is applied to the surface of the microtubule, enabling the dynein to translocate toward the proximal or minus-end of the microtubule. The dynein cross-bridge cycle includes the tight attachment of the dynein to the microtubule, the dissociation of the dynein from the microtubule upon dynein binding to ATP, the conformational change in the dynein structure that depends on ATP hydrolysis and subsequent release of the hydrolysis products, and the reattachment of the dynein to the microtubule. Under the appropriate conditions, a molecular cargo tethered to the dynein will be carried from one part of the cell to another along a microtubule track. The cargo may be a membrane-bounded organelle or vesicle, the centrosome, a kinetochore, or another microtubule. 5
In situ, dynein is a complex oligomer comprised of up to one dozen polypeptides of different sizes. These subunits include several light chains (Mr < 25 kDa), up to four intermediate chains (Mr 60-140 kDa), and one, two, or three heavy chains (Mr > 500 kDa). The light and intermediate chains, together with nondy-nein proteins, including the dynactin complex, contribute to the regulation of dynein activity and mediate the tethering of the dynein to its molecular cargo (e.g., Ref. 9). The actual motor activity of dynein resides in each heavy chain. An isolated heavy chain, separated from the other dynein constituents, is able to produce microtubule translocation in vitro at a velocity similar to that produced by intact dynein (10). Depending on the source of the dynein, the dynein particle comprises a single heavy chain, a homodimer of two identical heavy chains, a heterodimer, or a heterotrimer.Favorable electron micrographs reveal that each heavy chain forms a tail, a globular head, and a short “ antenna” that extends from the head (11). The tail interacts with other proteins to tether the dynein to its cargo, and in dyneins comprising multiple heavy chains, the heavy chains are gathered together by their tail domains to form a “bouquet” structure. Controlled proteolysis can separate the head from the tail and has revealed that the globular head domain contains the motor activity and the tail tethers the dynein to its cargo (12,13). The “ antenna” is the B-link that is thought to be the ATP-sensitive microtubule-binding site required for movement.Over the last decade, the complete sequences of nearly 20 dynein heavy chains have been reported. These include dyneins of all recognized functional classes-axonemal (ciliary) outer and inner arms, and nonaxonemal types 1 and 2-and from a wide variety of organisms. The examination of these sequences reveals several features found in all dyneins: 1. The dynein heavy chain is very large, approximately 4600 residues in length.2. The N-terminal —1300 residues form the tail domain. The sequence of this domain is the most divergent portion of the heavy chain, which is consistent with its role in specifying the cargo to which the dynein is tethered.3. The central catalytic domain contains four evenly spaced phosphatebinding P-loops. The sequence of the first P-loop (P-1) is invariant-GPAGTGKT-and the first P-loop is the site of MgATP2" hydrolysis, which is required for movement. The function(s) of the other P-loops is not understood, but one dynein heavy chain can bind up to four molecules of ATP and the dynein activity may be modulated by the binding of nucleotides to these other sites (14-16).
