Cells all have intricate cytoskeletal networks, comprised of microtubules, actin, and intermediate filaments. This network provides structural integrity to the cell, allows motility and cellular migration, and also provides a scaffold of structure on which to transport organelles and proteins within the cell. During mitosis, there is a great amount of protein trafficking along the microtubules within the spindle. To accomplish the task of moving proteins along microtubules, the cell employs microtubule-based motors. These motor proteins all have ATPase activity, and use the energy from the hydrolysis of ATP to physically walk along the length of microtubules. There are two classes of microtubule motors: the kinesin superfamily that moves along microtubules toward the plus-end; and the dyneins, which move towards the minus end.
During mitosis, dynein walks toward the spindle poles. Dynein is required for many proteins to localize to the spindle poles, and for removing checkpoint proteins from kinetochores after microtubule attachment, and is also for focusing of microtubules at the poles.
The kinesin superfamily of motors provides very diverse cellular functions. At one time, kinesins were classified by the location of their motor domains: kinesins with the motor domain at the N-terminus are usually plus-end directed, those with the motor domain at the C-terminus are minus-end directed. Many kinesins are localized throughout the spindle, and are required for many events during mitosis. Eg5 is a bipolar plus-end directed kinesin that binds antiparallel microtubules in the middle of the spindle, and pushes the spindle poles in opposite directions. Inhibition of Eg5 in the spindle results in monopolar spindles.
A special subgroup of kinesins with the motor domain on the interior of the protein (kinesin-13 family) does not have motile properties, but instead use the energy from ATP hydrolysis to depolymerize microtubules. These kinesins bind to the ends of microtubules, and promote the curling out of microtubule protofilaments, thereby destabilizing them. ATP hydrolysis by these enzymes is required for release of tubulin subunits from these kinesins, and is thus required for the processivity of these enzymes. The members of this family in mammals are Kif2a, Kif2b, and Kif2c (MCAK). These proteins are global regulators of microtubule dynamics during mitosis, and are essential for assembling a functional spindle.
Mitotic Centromere-Associated Kinesin (MCAK)
Mitotic Centromere-Associated Kinesin (MCAK, or XKCM1 in Xenopus) is a member of the kinesin-13 protein family, and possesses microtubule depolymerase activity. MCAK is located on the spindle poles and also colocalizes with Aurora B kinase at the inner centromere during prometaphase and metaphase. MCAK is a key microtubule catastrophe factor during mitosis. Inhibition of MCAK in Xenopus extracts generates large microtubule structures. Addition of the purified N-terminus of MCAK to mitotic cells blocks the native protein from localizing to the centromere, and generates chromosome misalignment and anaphase lagging chromosomes.
MCAK is phosphorylated and regulated by Aurora B kinase during mitosis. Phosphorylation of MCAK on SI 96 by Aurora B inhibits its depolymerase activity, and this modification is seen at the inner centromere and at the spindle poles. Phosphorylation of T95 (pT95) on MCAK by Aurora B targets it to chromatin arms, pT95 excludes MCAK from the inner centromere. These results suggest that MCAK is differentially regulated by phosphorylation on different residues and in different locations by the same kinase. MCAK is also phosphorylated on SI96 by Aurora A at the centrosome, and is involved in the formation of Ran-mediated asters.
The protein phosphatase 2A (PP2A) is located on the kinetochore, and has been hypothesized to play a role in MCAK activation, although no proof for this model exists. Although there are no data that suggest that MCAK is deposphorylated to stimulate its activity, there is one protein that stimulates MCAK, most likely via direct binding. MCAK is stimulated by Inner Centromere Kin-I Stimulator (ICIS), which is another microtubule associated protein (MAP).
Microtubule Motor Proteins reference
1. Sharp D J, Rogers G C, Scholey J M. Microtubule motors in mitosis[J]. Nature, 2000, 407(6800): 41-47.
2. Gaetz J, Kapoor T M. Dynein/dynactin regulate metaphase spindle length by targeting depolymerizing activities to spindle poles[J]. The Journal of cell biology, 2004, 166(4): 465-471.
3. Hirokawa N, Takemura R. Kinesin superfamily proteins and their various functions and dynamics[J]. Experimental cell research, 2004, 301(1): 50-59.
4. Langford G M. Actin-and microtubule-dependent organelle motors: interrelationships between the two motility systems[J]. Current opinion in cell biology, 1995, 7(1): 82-88.
5. Hunter A W, Caplow M, Coy D L, et al. The kinesin-related protein MCAK is a microtubule depolymerase that forms an ATP-hydrolyzing complex at microtubule ends[J]. Molecular cell, 2003, 11(2): 445-457.
6. Lan W, Zhang X, Kline-Smith S L, et al. Aurora B phosphorylates centromeric MCAK and regulates its localization and microtubule depolymerization activity[J]. Current biology, 2004, 14(4): 273-286.
7. Andrews P D, Ovechkina Y, Morrice N, et al. Aurora B regulates MCAK at the mitotic centromere[J]. Developmental cell, 2004, 6(2): 253-268.