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Dieter Klopfenstein and Ron Vale
Kinesin-family member Unc104/KIF1A is conserved from Dictyostelium
to humans and is involved in microtubule-dependent vesicle transport. While
little is known about the molecular mechanism how this motor interacts with
its vesicular cargo, a candidate membrane interaction region is a pleckstrin
homology (PH) domain located at the C-terminus of the motor protein. We show
here that the PH domain of Dictyostelium Unc104 is sufficient for binding to
and transporting protein-free liposomes composed of phophatidylinositol-4,5-bisphosphate
(PtdIns-4,5-P2). The PH domain binds to and transports native and trypsinized
Dictyostelium vesicles along microtubules in an in vitro assay. This suggests
that lipid binding may be important for binding native vesicles although membrane
proteins are likely to play a role as well. To further demonstrate that the
PH domain is the critical cargo binding domain, we show that a mini-Unc104 consisting
of the motor domain fused directly to the minimal PH domain is sufficient to
reconstitute Dictyostelium vesicle and liposome transport. Moreover, the PH
domain can be substituted for the tail domain of mitotic minus-end directed
motor ncd and the chimeric motor will transport vesicles towards the microtubule-minus
end. Taken together, our data suggest that the PH domain of Unc104 functions
as a cargo binding domain for vesicle transport.
ASCB 2001
last updated 1/14/02
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