Mutations in the "dynein regulatory complex" alter the ATP-insensitive binding sites for inner arm dyneins in Chlamydomonas axonemes.

Abstract

Abstract. To understand mechanisms of regulation of dynein activity along and around the axoneme we further characterized the "dynein regulatory complex" (drc). The lack of some axonemal proteins, which together are referred to as drc, causes the suppression of flagellar paralysis of radial spoke and central pair mutants. The drc is also an adapter involved in the ATPinsensitive binding of 12 and 13 inner dynein arms to doublet microtubules. Evidence supporting these conclusions was obtained through analyses of five drc mutants: pf2, pf 3, suppi3, suppi4, and supp/5. Axonemes from drc mutants lack part of 12 and 13 inner dynein arms as well as subsets of seven drc components (apparent molecular weight from 29,000 to 192,000). In the absence of ATP-Mg, dynein-depleted axonemes from the same mutants bind 12 and 13 inner arms at both ATP-sensitive and -insensitive sites. At ATPinsensitive sites, they bind 12 and 13 inner arms to an extent that depends on the drc defect. This evidence suggested to us that the drc forms one binding site for the 12 and 13 inner arms on the A part of doublet microtubules.

Casein Kinase I Is Anchored on Axonemal Doublet Microtubules and Regulates Flagellar Dynein Phosphorylation and Activity

“…Reconstitution and functional assays demonstrate that the radial spokes are required for wild-type dynein activity, and the velocity of dyneindriven microtubule sliding is mediated by posttranslational modification of the inner dynein arms (20). These results are consistent with other evidence indicating that the central pair/ radial spoke apparatus along with the dynein regulatory complex controls dynein activity (14,17,(21)(22)(23)(24)(25)(26). The challenge has been to define the biochemical signaling pathway and to determine how this biochemistry can be altered by mechanical interaction between the central pair structures, radial spoke structures, and outer doublet microtubule components to control dynein activity.…”

“…Based on phenotypic analysis of motility in mutants lacking I1, it appears that I1 and phosphorylation of IC138 play a role in control of flagellar waveform (12,15). Moreover, genetic analysis has revealed a functional linkage between the central pair/radial spoke structures and control of waveform (14,(21)(22)(23)(24)(25)(26). Although we do not yet know how alteration in dynein activity can result in changes in waveform, part of the answer is that the central pair/radial spoke system alters dynein activity via changes in phosphorylation.…”

The radial spokes and central apparatus: Mechano‐chemical transducers that regulate flagellar motility

“…Importantly, the flagella from the suppressed mutant cells produce only symmetric waveforms [Brokaw et al, 1982], indicating that the radial spokes and central apparatus may be involved in converting simple symmetric bends into the asymmetric waveforms required for forward swimming. The suppressor mutations were later found to be mutations in outer Porter et al, 1994] or inner [Porter et al, 1992] dynein arm components, or in polypeptides comprising a dynein regulatory complex Piperno et al, 1992Piperno et al, , 1994. These observations provide strong evidence that the central apparatus ultimately regulates dynein-driven microtubule sliding in a pathway that includes the radial spokes and dynein regulatory complex.…”

Molecular architecture of axonemal microtubule doublets revealed by cryo-electron tomography

“…For example, there are distinctive densities inside the regions where the dynein heads are located. The largest feature, on PFs A10-A12, sits inside the attachment region for nexin and the dynein regulatory complex with which it is associated [23][24][25][26] . The density we observe between PFs 12 and 13 is at the point where the radial spokes meet the doublets and may be part of their attachment.…”