IDENTIFICATION OF A CA2-BINDING LIGHT-CHAIN WITHIN CHLAMYDOMONAS OUTER ARM DYNEIN()

We describe here the molecular cloning of the M(r) 18,000 dynein light chain from the outer arm of Chlamydomonas flagella, In vivo, this mol ecule is directly associated with the gamma dynein heavy chain. Sequen ce analysis indicates that this light chain is a novel member of the c almodulin superfamily of Ca2+ binding regulatory proteins; this molecu le is 42, 37 and 36% identical to calmodulin, centrin/caltractin and t roponin C, respectively, and also shows significant similarity to myos in light chains. Although four helix-loop-helix elements are evident, only two conform precisely to the EF hand consensus and are therefore predicted to bind Ca2+ in vivo, In vitro Ca2+ binding studies indicate that this dynein light chain (expressed as a C-terminal fusion with m altose binding protein) has at least one functional Ca2+ binding site with an apparent affinity for Ca2+ of similar to 3x10(-5) M, Within th e Chlamydomonas flagellum, the transition from an assymmetric to a sym metric waveform (which implies an alteration in dynein activity) is me diated by an increase in intraflagellar Ca2+ from 10(-6) to 10(-4) M; this transition is altered in mutants that lack the outer arm, The dat a presented here suggest that a Ca2+-dependent alteration in the inter action of this dynein light chain with the motor containing heavy chai n may affect outer arm function during flagellar reversal.