One of the most astounding examples of molecular machinery in the cell is the mitotic spindle that governs the segregation of chromosomes during cell division. You can watch the astonishing process of mitotic cell division unfold in this animation:

A crucial aspect of mitosis is the movement of chromosomes — initially the alignment of as paired sister chromatids during prometaphase and metaphase, and then the separation of these sister chromatids during anaphase. But how does the spindle exert force on the chromosomes? And how do the chromosomes attach to the spindle such that they can be moved to the proper place at the proper time? Here, I will give an introduction to this astounding process.

Microtubules

Two structural components make up the mitotic spindle — two large structures, found at the poles, called centrosomes, and protein filaments known as microtubules. From each of the centrosomes radiate the spindle microtubules, which bind to a chromosome (kinetochore microtubules), or a microtubule emanating from the other centrosome (polar microtubules), or the cell’s plasma membrane (astral microtubules).

The subunit of microtubules are heterodimers of tubulin, comprised of α-tubulin and β-tubulin. These heterodimers fit together to create linear protofilaments, 13 of which are aligned side-by-side to form a microtubule. Due to the asymmetry of tubulin heterodimers, the microtubule has intrinsic polarity. The plus end (which extends towards the chromosomes) is comprised of β-tubulin subunits, whereas the minus end (which is anchored at the centrosome) is comprised of α-tubulin subunits.

In the early mitotic stages, microtubules assemble and disassemble rapidly (mediated by the addition and removal of tubulin heterodimers), a phenomenon known as dynamic instability (the processes of depolymerization and polymerization are referred to as catastrophe and rescue respectively). By this process, microtubules randomly probe the cell, until they by chance encounter their target. The regulation of microtubule catastrophe and rescue is linked to the hydrolysis of guanosine triphosphate (GTP), a nucleotide bound to tubulin within the microtubule lattice. Each tubulin subunit is able to bind to one molecule of GTP. During polymerization, tubulin subunits bound to GTP are added to the growing end of the microtubule, forming a GTP cap which stabilizes the microtubule and promotes growth.

As the microtubule continues to polymerize, the GTP-bound tubulin subunits undergo hydrolysis, converting GTP to guanosine diphosphate (GDP). Once hydrolyzed, the tubulin subunits become less stable, and the microtubule is more prone to depolymerization. Upon reaching a critical concentration of GDP-bound tubulin at the microtubule end, catastrophe occurs, leading to rapid depolymerization. Microtubule rescue involves the exchange of GDP-bound tubulin for GTP-bound tubulin, thereby promoting microtubule growth. Certain cellular factors, such as microtubule-associated proteins (MAPs) and motor proteins, can facilitate this process by promoting the incorporation of GTP-bound tubulin or by protecting the GTP cap from hydrolysis. For example, kinesin-8 proteins are plus-end directed motor proteins that destabilize microtubules.¹ Another motor protein, kinesin-13, is bidirectional (being able to move in the direction of either the plus or minus ends of microtubules), in contrast to most motors that are unidirectional. They are thereby able to promote the depolymerization of a microtubule from both ends.² The activity of kinesin-13 proteins is “regulated by distinct targeting to regions of the spindle, by regulatory phosphorylation events, and by interactions with different binding partners.”³ These remarkable kinesin-13 motors bind the end of the microtubule and trigger a conformational change that results in microtubule depolymerization.⁴

Motor Proteins

In 2014, Discovery Institute published an animation of the molecular motor kinesin, which carries cellular cargo along microtubule tracks and is driven by ATP hydrolysis:

Kinesin typically moves in the direction of the plus end of the microtubule, towards the periphery of the cell. Another form of motor protein, called dynein, generally moves towards the minus end of the microtubule, i.e., towards the cell center. Whereas kinesin motor proteins walk step-by-step by placing one foot in front of the other, dynein moves by a swinging crossbridge mechanism.

One of the most incredible functions of motor proteins in the cell is their role in the assembly and function of the mitotic spindle during eukaryotic cell division.⁵ Envision a robotic factory that assembles and organizes the cell in preparation for undergoing division, helping to facilitate the controlled segregation of the genetic material into the daughter cells. This is represented in the (highly simplified) figure below:

As shown in the figure, the motor proteins involved in the organization of the mitotic spindle can be divided into four major classes: kinesin-5, kinesin-14, kinesins-4 and 10, and dynein. Kinesin-5 is directed towards the plus end and slides apart those microtubules that have their polarities oriented in opposite directions (i.e., antiparallel microtubules). Thus, kinesin-5 contributes to spindle elongation and bipolarity by pushing apart the spindle poles.⁶ On the other hand, kinesin-14 motors are directed towards the minus-end of microtubules. They each possess one motor domain in addition to other domains that associate with a different microtubule.⁷ They cross-link antiparallel interpolar microtubules at the spindle midzone, and thereby pull the poles towards one another.

Another group of kinesins, kinesin-4 and kinesin-10 (collectively known as chromokinesins), play a critical role in the positioning and segregation of chromosomes during mitosis.⁸ The job of chromokinesins is to move the chromosomes to their proper positions such that each daughter cell will receive the right number of chromosomes.

The last group of motor proteins involved in mitosis are dyneins that have their motor domains associated with the microtubules, which emanate from the centrosomes, and their cargo-binding domains bound to proteins embedded in the cell cortex. The movement of dynein exerts a pulling force on the microtubule minus ends and pulls the centrosomes in the direction of the cell cortex.⁹ This movement ensures that each daughter cell receives a complete set of chromosomes. Note that, although I have, for simplicity, only shown one microtubule emanating from each centrosome towards the cell cortex, in real life there would be many more than this.

Incredible Engineering

In any other realm of experience, we would, upon observing a robotic factory like this that operates at a nanoscale, immediately ascribe it to design. The engineering of the eukaryotic cell division cycle is astounding at every level. Moreover, these molecular motors are absolutely indispensable to successful cell division in eukaryotes and thus constitute an irreducibly complex system. It therefore points forcefully and unequivocally to conscious intent.

Notes

1. Shrestha S, Hazelbaker M, Yount AL, Walczak CE. Emerging Insights into the Function of Kinesin-8 Proteins in Microtubule Length Regulation. Biomolecules. 2018 Dec 20;9(1):1.
2. Ems-McClung SC, Walczak CE. Kinesin-13s in mitosis: Key players in the spatial and temporal organization of spindle microtubules. Semin Cell Dev Biol. 2010 May;21(3):276-82.
3. Ibid.
4. Desai A, Verma S, Mitchison TJ, Walczak CE. Kin I kinesins are microtubule-destabilizing enzymes. Cell. 1999 Jan 8;96(1):69-78.
5. Goshima G, Vale RD. The roles of microtubule-based motor proteins in mitosis: comprehensive RNAi analysis in the Drosophila S2 cell line. J Cell Biol. 2003 Sep 15;162(6): 1003-16.
6. Ferenz NP, Gable A, Wadsworth P. Mitotic functions of kinesin-5. Semin Cell Dev Biol. 2010 May;21(3): 255-9. 
7. Mitra A, Meißner L, Gandhimathi R, Renger R, Ruhnow F, Diez S. Kinesin-14 motors drive a right-handed helical motion of antiparallel microtubules around each other. Nat Commun. 2020 May 22;11(1): 2565.
8. Mazumdar M and Misteli T. Chromokinesins: multitalented players in mitosis. Trends Cell Biol. 2005 Jul;15(7): 349-55.
9. Raaijmakers JA and Medema RH (2014). Function and regulation of dynein in mitotic chromosome segregation. Chromosoma. 123(5): 407-22.