STRUCTURAL AND MOLECULAR CHARACTERIZATION OF DYNEIN IN A GALL-MIDGE INSECT HAVING MOTILE SPERM WITH ONLY THE OUTER ARM

The dipteran Monarthropalpus flavus possesses a peculiar sperm axoneme , characterized by multiple rows of microtubular doublets linked by th e outer dynein arms only, lacking any equivalent of the central pair/r adial spoke complex. The structure of these dynein molecules was studi ed by electron microscopy (EM). Using the quick-freeze, deep-etch meth od of EM, they were found to be similar to outer dynein arms described previously. Two globular ''heads,'' each subdivided by a cleft, are c learly discernible. ''Stalks'' extend from proximal head to contact th e B-tubule of the adjacent doublet. Unlike the situation in vertebrate sperm, the stalks sometimes branch into two thinner strands that cont act the B-tubule at different sites. Treatment of demembranated sperm cells with ATP and vanadate induces conformational changes in the dyne in outer arms. These are interpreted as the result of rotation of the dynein head with respect to what is observed in axonemes in rigor cond ition (after ATP depletion). SDS-PAGE indicates that the high-molecula r-weight complement of this molecule comprises a single heavy chain. S pecific dynein heavy chain-related DNA sequences corresponding to the catalytic-phosphate binding region were amplified by RT-PCR. Only one axonemal dynein sequence was identified among all amplified fragments. Southern blot analysis performed on genomic DNA using this sequence a s a probe identified two hybridizing genes, only one of which is able to encode a functional product. Thus, genetic analysis indicates that this axonemal outer arm dynein is a homodymer of a single heavy chain subunit. In vivo, spermatozoa of this species are stored in a rolled c onfiguration in female spermatheca, where they move rapidly with a wav e-like motion. This movement could not be reproduced in vitro, except when spermatozoa were constrained in a bent configuration by some mech anical impediment. We propose that, in the absence of both the central pair/radial spoke complex and the inner arms, a curvature-dependent a ctivation acts to trigger motility in these spermatozoa. (C) 1998 Wile y-Liss, Inc.