The kinesin-5 tail and bipolar minifilament domains are the origin of its microtubule crosslinking and sliding activity

The kinesin-5 motor protein plays an essential role in mitotic spindle assembly through its interaction and sliding of microtubules. Kinesin-5 has several key domains including its c-terminal tail domains, N-terminal motor domains, and its bipolar assembly domain which is critical for its tetrameric bundling.  While the domains of kinesin-5 are known, the interplay that occurs between these domains to control MT crosslinking and sliding mechanisms is not well defined.  Recently, Nithianantham et al. performed structural and functional studies on kinesin-5 and determined that the bipolar assembly domain and c-terminal tail have important roles in MT interaction and regulation.  The group first performed x-ray crystallography structural studies and utilized the information to develop an active, bipolar mini-tetramer kinesin -5 motor (38 nm) that was roughly half the length of full-length kinesin-5 tetramer (80 nm); these versions were used in subsequent studies to define the role of the minifilament and tails on MT crosslinking and sliding.   The group utilized fluorescent TIRF microscopy to examine the motility of the kinesin-5 mini-tetramer and found that it had similar behavior to the full-length form where it moved towards the plus end of MTs and encountered frequent pauses due to the tail-motor domain interaction.  However, further comparison of the two tetramer versions revealed that the differing minifilament lengths affected its overall flexibility, and these differences in length and flexibility had a measurable effect on the tetramer's ability to crosslink MTs; specifically, their findings showed that mini-tetramers were less efficient than full-length kinesin-5 motors at forming anti-parallel MT bundles.  Once crosslinking occurred, the mini-tetramers were more efficient at sliding MTs relative to the full-length version.  Collectively, these results suggest that the minifilament can have profound effects on the kinesin-5 motor function and ability to regulate MTs, which may be important in the setting of mitotic spindle assembly.  Cytoskeleton Inc.’s fluorescent tubulins (Cat. # TL670M and Cat. # TL590M) in combination with biotin-labeled tubulin (Cat. # T333P) were critical reagents utilized in the MT crosslinking experiments. 


Figure Legend: Schematic showing kinesin-5 mini-tetramer vs full length tetramer, and its effect on its movement and interaction along microtubules.