Structural comparison of purified dynein proteins with in situ dynein arms.
Using the quick-freeze deep-etch technique, we describe the structure of outerarm dynein proteins from Chlamydomonas and Tetrahymena after adsorption to a mica surface, after high-salt dissociation, and after glutaraldehyde fixation, and compare these images to the configuration of outer arms bound to microtubules. After adsorption to mica, the extracted dyneins from both organisms look like three-headed "bouquets", as reported for Tetrahymena by Johnson & Wall (1983b). High magnification images demonstrate that each head carries a slender "stalk" and a long "stem", and that small subunits decorate the stems and create a "flowerpot" domain at the base of the bouquet. Exposure to high salt induces this trimer to dissociate into a two-headed species and a single-headed species; it also stimulates the decorative elements to dissociate from the stems. Dynein is thus constructed on the same general plan as myosin, with large globular heads, narrow stems and additional small subunits that associate with the stems. The splayed-out image of the bouquet appears to be a distortion arising during adsorption to mica since, after brief glutaraldehyde fixation, the three heads remain closely associated as vertices of a triangular unit. In situ, the three heads also adopt this trigonal configuration. Two of the three are visible from the exterior of the axoneme and constitute the bilobed rigor head we described previously (Goodenough & Heuser, 1982). The third head faces the interior of the axoneme where, we propose, it forms the "hook" of the outer arm as seen in thin section. We further propose that the decorative elements associated with the stem coalesce to form the two outer-arm "feet" seen in situ, and that at least one of the in vitro stalks is equivalent to the in situ stalk, which extends from the head to the B microtubule. Deep-etch images of stretched axonemes, partially extracted axonemes, and dynein-decorated brain microtubules indicate that each outer arm, as traditionally viewed, is a hybrid of two dynein molecules: its two feet derive from one molecule, whereas its trigonal head derives from the molecule located distally. The resultant overlapping configuration creates the diagonal "linkers" seen in situ, which correspond to the in vitro stems. Thus, a row of dynein arms is essentially a dynein polymer that extends from the tip to the base of a doublet microtubule, each head riding on its neighbor's feet like a row of circus elephants.(ABSTRACT TRUNCATED AT 400 WORDS)
D000251:Adenosine Triphosphatases / Q000302:isolation & purification*; D000255:Adenosine Triphosphate; D000818:Animals; D002696:Chlamydomonas / Q000032:analysis* / Q000648:ultrastructure; D004398:Dyneins / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D005613:Freeze Etching; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D008870:Microtubules / Q000648:ultrastructure; D013768:Tetrahymena / Q000032:analysis* / Q000648:ultrastructure
D046911:Macromolecular Substances; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D004398:Dyneins