RECONSTITUTION OF CHLAMYDOMONAS OUTER DYNEIN ARMS FROM ALPHA-BETA-SUBUNIT AND GAMMA-SUBUNIT - REQUIREMENT OF A 3RD FACTOR

The outer dynein arm of Chlamydomonas flagella, when isolated under Mg 2+-free conditions, tends to dissociate into an 11 to 12S particle (12 S dynein) containing the gamma heavy chain and a 21S particle (called 18S dynein) containing the alpha and beta heavy chains. We show here t hat functional outer arms can be reconstituted by the addition of 12S and 18S dyneins to the axonemes of the outer armless mutants oda1-oda6 . A third factor that sediments at similar to 7S is required for effic ient reconstitution of the outer arms on the axonemes of oda1 and oda3 . However, this factor is not necessary for reconstitution on the axon emes of oda2, oda4, oda5, and oda6 SDS-PAGE analysis indicates that th e axonemes of the former two mutants lack a similar to 70-kD polypepti de that is present in those of the other mutants as well as in the 7S fraction from the wild-type extract. Furthermore, electron micrographs of axonemal cross sections revealed that the latter four mutants, but not oda1 or oda3, have small pointed structures on the outer doublets , at a position in cross section where outer arms normally occur. We s uggest that the 7S factor constitutes the pointed structure on the out er doublets and facilitates attachment of the outer arm. The discovery of this structure raises a new question as to how the attachment site for the outer arm dynein is determined within the axoneme.