Using Total Internal Reflection Fluorescence (TIRF) microscopy technique and with the help of simulations, we are investigating how Kinesin2 motors work together.

Motor-DNA assembly.PNG

Once attached, processive motors take multiple steps on cytoskeletal filaments before falling off. This remarkable, but not yet fully understood property is considered to be the mechanistic basis of long-range transport in vivo. Transport in vivo, however, can also require different types of processive motors to team up leading to more complicated transport mechanisms. One such transport occurs in the cilium of C. elegans where the slow heterotrimeric KLP11/20/KAP and the fast homodimeric Osm-3 coordinate to facilitate anterograde intraflagellar transport (IFT). IFT is an excellent model to scrutinize motor coupling because the two-dimensional transport geometry with motors lined up on IFT trains can be closely mimicked in vitro. In our assays, dsDNA served as an IFT train mimic to specifically couple distinct types of IFT motors (kinesin-2). We engineered constitutively active fast and slow kinesin-2 motors to investigate the resulting transport behavior under defined conditions. This study sheds light on how distinct kinesin-2 motors may cooperate to move long IFT trains.