Dynein motors of the Chlamydomonas flagellum.

Chlamydomonas is a biflagellate unicellular green alga that has proven especially amenable for the analysis of microtubule (MT)-based molecular motors, notably dyneins. These enzymes form the inner and outer arms of the flagellum and are also required for intraflagellar transport. Dyneins have masses of approximately 1-2 MDa and consist of up to 15 different polypeptides. Nucleotide binding/hydrolysis and MT motor activity are associated with the heavy chains, and we detail here our current model for the substructural organization of these approximately 520-kDa proteins. The remaining polypeptides play a variety of roles in dynein function, including attachment of the motor to cargo, regulation of motor activity in response to specific inputs, and their necessity for the assembly and/or stability of the entire complex. The combination of genetic, physiological, structural, and biochemical approaches has made the Chlamydomonas flagellum a very powerful model system in which to dissect the function of these fascinating molecular motors.

The Tctex1/Tctex2 class of dynein light chains. Dimerization, differential expression, and interaction with the LC8 protein family.

The Tctex1/Tctex2 family of dynein light chains associates with the intermediate chains at the base of the soluble dynein particle. These components are essential for dynein assembly and participate in specific motor-cargo interactions. To further address the role of these light chains in dynein activity, the structural and biochemical properties of several members of this polypeptide class were examined. Gel filtration chromatography and native gel electrophoresis indicate that recombinant Chlamydomonas flagellar Tctex1 exists as a dimer in solution. Furthermore, yeast two-hybrid analysis suggests that this association also occurs in vivo. In contrast, both murine and Chlamydomonas Tctex2 are monomeric. To investigate protein-protein interactions involving these light chains, outer arm dynein from Chlamydomonas flagella was cross-linked using dimethylpimelimidate. Immunoblot analysis of the resulting products revealed the interaction of LC2 (Tctex2) with LC6, which is closely related to the highly conserved LC8 protein found in many enzyme systems, including dynein. Northern dot blot analysis demonstrated that Tctex1/Tctex2 family light chains are differentially expressed both in a tissue-specific and developmentally regulated manner in humans. These data provide further support for the existence of functionally distinct populations of cytoplasmic dynein with differing light chain content.