Mechanism of transport of IFT particles in C. elegans cilia by the concerted action of kinesin-II and OSM-3 motors

Abstract

The assembly and function of cilia on Caenorhabditis elegans neurons depends on the action of two kinesin-2 motors, heterotrimeric kinesin-II and homodimeric OSM-3–kinesin, which cooperate to move the same intraflagellar transport (IFT) particles along microtubule (MT) doublets. Using competitive in vitro MT gliding assays, we show that purified kinesin-II and OSM-3 cooperate to generate movement similar to that seen along the cilium in the absence of any additional regulatory factors. Quantitative modeling suggests that this could reflect an alternating action mechanism, in which the motors take turns to move along MTs, or a mechanical competition, in which the motors function in a concerted fashion to move along MTs with the slow motor exerting drag on the fast motor and vice versa. In vivo transport assays performed in Bardet-Biedl syndrome (BBS) protein and IFT motor mutants favor a mechanical competition model for motor coordination in which the IFT motors exert a BBS protein–dependent tension on IFT particles, which controls the IFT pathway that builds the cilium foundation.

Regulation of a heterodimeric kinesin-2 through an unprocessive motor domain that is turned processive by its partner

“…Two KLP11 heads or two KLP20 heads along with their native neck linkers were therefore fused to the heterodimeric tail (Fig. 1), which ensures reliable dimerization (9). We designated these chimeric constructs as KLP11/11 or KLP20/20 to stress the presence of two identical heads.…”

“…In the Caenorhabditis elegans sensory cilium, the kinesin-like protein 11 (KLP11) and kinesin-like protein 20 (KLP20) heterodimerize to comprise a molecular motor with two distinct Nterminal catalytic motor domains (9)(10)(11). A third protein, termed kinesin-associated protein 1 (KAP1), is thought to provide the physical link between the C terminus of the heterodimeric motor protein and the IFT particle (12).…”

Assembly of primary cilia

“…The coordinated use of kinesin-II and OSM-3 in C. elegans amphid channel cilia is dependent on BBS proteins, which appear to be required for keeping the two motors together during anterograde IFT along the middle segment (Blacque et al, , 2008Ou et al, 2005;Pan et al, 2006). Interestingly, in mutant mice that lack BBS-1 or -4, the olfactory cilia are either greatly reduced in length or absent and the mice are anosmic (Kulaga et al, 2004).…”

Mutation of the MAP kinase DYF-5 affects docking and undocking of kinesin-2 motors and reduces their speed in the cilia of Caenorhabditis elegans

“…Live imaging of the movement of these kinesins suggests that kinesin II alone moves at 0.5 m/s, and OSM-3 alone moves at 1.3 m/s, whereas the two motor complexes together move at 0.7 m/s (8). Recently, Pan et al (9) have shown that these in vivo transport rates can be reconstituted in vitro by using purified kinesin II and OSM-3 motors. Thus far, two proteins have been identified that are required to stabilize IFT complexes transported by both kinesin II and OSM-3, BBS-7 and BBS-8 (10).…”