Mitosis, the process of cell division, ensures the accurate distribution of genetic material into two new daughter cells. This precision relies on the kinetochore, a complex molecular machine. This protein structure acts as the central regulatory hub on each chromosome, bridging the duplicated DNA to the spindle apparatus that pulls the chromosomes apart. The kinetochore’s primary function is to serve as the attachment site for spindle microtubules and to monitor attachment correctness before allowing cell division to proceed. Errors in chromosome separation can lead to aneuploidy, a condition linked to developmental disorders and cancer.
Kinetochore Structure and Assembly
The kinetochore is a massive, multi-layered protein assembly built upon the centromere, a specialized region of the chromosome. It is a dynamic structure whose composition changes throughout the cell cycle. The vertebrate kinetochore has a characteristic trilaminar appearance.
The deepest layer is the inner kinetochore, which is tightly associated with the centromeric DNA. This region contains specialized chromatin, specifically nucleosomes with the histone variant CENP-A. The inner kinetochore acts as a permanent scaffold that persists throughout the cell cycle, ensuring the kinetochore is built at the correct location. It is composed of the Constitutive Centromere-Associated Network (CCAN), a 16-protein complex that interacts with the CENP-A chromatin.
External to this foundational layer is the outer kinetochore, the dynamic interface responsible for interacting with spindle microtubules. This outer plate is assembled only during cell division and is composed primarily of the KMN network (Knl1, Mis12, and Ndc80). The structure also features a fibrous corona, a mesh-like network of proteins visible when microtubules are not yet attached. This temporary corona aids in the initial capture of microtubules.
Microtubule Capture and Anchoring
The kinetochore’s initial mechanical function is to establish a stable connection with the dynamic, searching tips of the spindle microtubules. This “search-and-capture” mechanism involves the outer kinetochore acting as a high-affinity receptor for the microtubule plus-ends. The primary complex forming this robust, load-bearing linkage is the Ndc80 complex, a component of the KMN network that projects outward.
The Ndc80 complex forms a stable “end-on” attachment, coupling the microtubule plus-end directly to the kinetochore. This allows the chromosome to move with the microtubule’s growth and shrinkage. In humans, a single kinetochore can anchor approximately 20 microtubules. This robust connection withstands the pulling forces generated by the mitotic spindle.
Motor proteins associated with the kinetochore assist in the capture process and subsequent movement. The minus-end-directed motor protein dynein plays a role in initial microtubule capture, transporting the attached chromosome toward the spindle pole. Dynein and the Ndc80 complex coordinate during prometaphase to efficiently capture the microtubule plus-end, ensuring the chromosome is integrated into the spindle.
The motor protein CENP-E, a plus-end-directed kinesin, contributes to alignment by generating movement that positions the chromosome at the center of the cell, known as the metaphase plate. CENP-E works with Ndc80 to transition the attachment from a less stable lateral interaction to the final, stable end-on attachment. The combined activities of these motors and the Ndc80 complex create the strong linkage required for force transduction and chromosome alignment.
Monitoring Chromosome Alignment
The kinetochore acts as the cell’s quality control center, ensuring chromosome attachment is correct before division. This monitoring involves two interconnected mechanisms: sensing mechanical tension and activating the Spindle Assembly Checkpoint (SAC). This dual surveillance confirms that every sister chromatid pair is correctly bioriented, meaning each chromatid is attached to microtubules from opposite spindle poles.
Correct biorientation generates mechanical strain, or tension, across the centromere region linking the two sister kinetochores. The kinetochore is designed to sense this tension, and the presence of strain stabilizes the microtubule attachment while suppressing error-correction mechanisms. If attachment is incorrect (e.g., syntelic attachment), the lack of opposing tension signals a defect. The Aurora B kinase acts as a sensor, promoting the detachment of incorrect, low-tension attachments until stable biorientation is achieved.
Simultaneously, the kinetochore activates the SAC, a signaling pathway that halts mitosis until all chromosomes are properly attached. Unattached or improperly attached kinetochores are the primary SAC activators. These defective kinetochores recruit a cascade of checkpoint proteins, including Mad1 and Mad2, which generate the inhibitory signal.
The recruitment of Mad1-Mad2 catalyzes the formation of the Mitotic Checkpoint Complex (MCC). The MCC is a potent, diffusible inhibitor composed of Mad2, BubR1, and Bub3. It binds to and inactivates the anaphase-promoting complex/cyclosome (APC/C). By inhibiting the APC/C, the SAC prevents the cell from prematurely entering anaphase, pausing the cell cycle until the error is resolved. Once stable biorientation is established, checkpoint proteins are stripped from the kinetochore, silencing the SAC signal and allowing the cell to move forward.
Initiating Chromosome Segregation
The kinetochore’s final act is signaling the transition from metaphase to anaphase, which involves sister chromatid separation. This transition is regulated by the silencing of the SAC, which occurs only after every kinetochore achieves a high-tension, bioriented attachment. Once the SAC is satisfied and the inhibitory MCC signal stops, the Anaphase Promoting Complex/Cyclosome (APC/C) is activated by its co-activator Cdc20.
The activated APC/C is an E3 ubiquitin ligase that targets specific proteins for destruction. Its primary target is securin, an inhibitory protein that keeps the protease separase inactive. The rapid degradation of securin releases the active separase enzyme.
Separase performs the decisive molecular action of anaphase onset: it cleaves a subunit of the cohesin protein complex. Cohesin forms ring-like structures that hold the sister chromatids together. Cleaving the cohesin rings removes the final physical link between the sisters, allowing the spindle microtubules to draw the now-individual chromosomes to opposite poles.