Direct visualization of microtubule flux during metaphase and anaphase in crane-fly spermatocytes

Microtubule flux in spindles of insect spermatocytes, long-used models for studies on chromosome behavior during meiosis, was revealed after iontophoretic microinjection of rhodamine-conjugated (rh)-tubulin and fluorescent speckle microscopy. In time-lapse movies of crane-fly spermtocytes, fluorescent speckles generated when rh-tubulin incorporated at microtubule plus ends moved poleward through each half-spindle and then were lost from microtubule minus ends at the spindle poles. The average poleward velocity of approximately 0.7 microm/min for speckles within kinetochore microtubules at metaphase increased during anaphase to approximately 0.9 microm/min. Segregating half-bivalents had an average poleward velocity of approximately 0.5 microm/min, about half that of speckles within shortening kinetochore fibers. When injected during anaphase, rhtubulin was incorporated at kinetochores, and kinetochore fiber fluorescence spread poleward as anaphase progressed. The results show that tubulin subunits are added to the plus end of kinetochore microtubules and are removed from their minus ends at the poles, all while attached chromosomes move poleward during anaphase A. The results cannot be explained by a Pac-man model, in which 1) kinetochore-based, minus end-directed motors generate poleward forces for anaphase A and 2) kinetochore microtubules shorten at their plus ends. Rather, in these cells, kinetochore fiber shortening during anaphase A occurs exclusively at the minus ends of kinetochore microtubules.     

Microtubule Flux Mediates Poleward Motion of Acentric Chromosome Fragments during Meiosis in Insect Spermatocytes

We applied a combination of laser microsurgery and quantitative polarization microscopy to study kinetochore-independent forces that act on chromosome arms during meiosis in crane fly spermatocytes. When chromosome arms located within one of the half-spindles during prometa- or metaphase were cut with the laser, the acentric fragments (lacking kinetochores) that were generated moved poleward with velocities similar to those of anaphase chromosomes (approximately 0.5 microm/min). To determine the mechanism underlying this poleward motion of detached arms, we treated spermatocytes with the microtubule-stabilizing drug taxol. Spindles in taxol-treated cells were noticeably short, yet with polarized light, the distribution and densities of microtubules in domains where fragment movement occurred were not different from those in control cells. When acentric fragments were generated in taxol-treated spermatocytes, 22 of 24 fragments failed to exhibit poleward motion, and the two that did move had velocities attenuated by 80% (to approximately 0.1 microm/min). In these cells, taxol did not inhibit the disjunction of chromosomes nor prevent their poleward segregation during anaphase, but the velocity of anaphase was also decreased 80% (approximately 0.1 microm/min) relative to untreated controls. Together, these data reveal that microtubule flux exerts pole-directed forces on chromosome arms during meiosis in crane fly spermatocytes and strongly suggest that the mechanism underlying microtubule flux also is used in the anaphase motion of kinetochores in these cells.     

Cell division and the microtubular cytoskeleton]

Kinetochore microtubules result from an interaction between astral microtubules and the kinetochore of the chromosomes after breakdown of the nuclear envelope at the end of prophase. In this process, the end of a microtubule projecting from one of the polar regions contacts the primary constriction of a chromosome. The latter then undergoes rapid poleward movement. Concerning the mechanism of anaphase chromosome movement, the motive force for the chromosome-to-pole movement appears to be generated at the kinetochore or in the region very close to it. It has not been determined whether chromosomes propel themselves along stationary kinetochore microtubules by a motor at the kinetochore, or they are pulled poleward by a traction fiber consisting of kinetochore microtubules and associated motors. As chromosomes move poleward coordinate disassembly of kinetochore microtubules might occur from their kinetochore ends. In diatom and yeast spindles, elongation of the spindle in anaphase (anaphase B) may be explained by microtubule assembly at polar microtubule ends in the spindle mid-zone and sliding of the antiparallel microtubules from the opposite poles. The sliding force appears to be generated through an ATP-dependent microtubule motor. In isolated sea urchin spindles, the microtubule assembly at the equator alone might provide the force for spindle elongation, although, in addition, involvement of microtubule sliding by a GTP-requiring mechanochemical enzyme cannot be excluded. Discussions were made on possible participation in anaphase chromosome movement of such microtubule motors as dynein, kinesin, dynamin and the claret segregation protein.     

Model of chromosome motility in Drosophila embryos: adaptation of a general mechanism for rapid mitosis

During mitosis, ensembles of dynamic MTs and motors exert forces that coordinate chromosome segregation. Typically, chromosomes align at the metaphase spindle equator where they oscillate along the pole-pole axis before disjoining and moving poleward during anaphase A, but spindles in different cell types display differences in MT dynamicity, in the amplitude of chromosome oscillations and in rates of chromatid-to-pole motion. Drosophila embryonic mitotic spindles, for example, display remarkably dynamic MTs, barely detectable metaphase chromosome oscillations, and a rapid rate of "flux-pacman-dependent" anaphase chromatid-to-pole motility. Here we develop a force-balance model that describes Drosophila embryo chromosome motility in terms of a balance of forces acting on kinetochores and kMTs that is generated by multiple polymer ratchets and mitotic motors coupled to tension-dependent kMT dynamics. The model shows that i), multiple MTs displaying high dynamic instability can drive steady and rapid chromosome motion; ii), chromosome motility during metaphase and anaphase A can be described by a single mechanism; iii), high kinetochore dynein activity is deployed to dampen metaphase oscillations, to augment the basic flux-pacman mechanism, and to drive rapid anaphase A; iv), modulation of the MT rescue frequency by the kinetochore-associated kinesin-13 depolymerase promotes metaphase chromosome oscillations; and v), this basic mechanism can be adapted to a broad range of spindles.     

Cytoplasmic dynein is required for poleward chromosome movement during mitosis in Drosophila embryos

The movement of chromosomes during mitosis occurs on a bipolar, microtubule-based protein machine, the mitotic spindle. It has long been proposed that poleward chromosome movements that occur during prometaphase and anaphase A are driven by the microtubule motor cytoplasmic dynein, which binds to kinetochores and transports them toward the minus ends of spindle microtubules. Here we evaluate this hypothesis using time-lapse confocal microscopy to visualize, in real time, kinetochore and chromatid movements in living Drosophila embryos in the presence and absence of specific inhibitors of cytoplasmic dynein. Our results show that dynein inhibitors disrupt the alignment of kinetochores on the metaphase spindle equator and also interfere with kinetochore- and chromatid-to-pole movements during anaphase A. Thus, dynein is essential for poleward chromosome motility throughout mitosis in Drosophila embryos.     

Microtubule dynamics determine chromosome lagging and transport of acentric fragments

The general direction of transport of spindle inclusions including acentric chromosome fragments during mitosis in endosperm of the higher plants Haemanthus is predictable and stage-dependent. Their segregation is random and they are usually eliminated from the spindle. This transport is superimposed on normal chromosome segregation. Thus, there are 2 superimposed mitotic transports: one which distributes kinetochores and the other which distributes spindle inclusions. The functional relation of these 2 transports to each other is not well understood. However, due to this 'non-kinetochore transport,' fragments may persist a few consecutive divisions before being permanently eliminated from the nucleus. Malfunction of kinetochores of any chromosome, resulting in the loss of their anchorage within the spindle, subjects them to 'non-kinetochore' transport and nearly certain, permanent elimination from the spindle. Additionally, experimental evidence presented here demonstrates that rapid polymerization (elongation) of microtubules may desynchronize anaphase and cause lagging of whole chromosomes. This may be one more, previously unconsidered, factor which may cause the malfunction of the kinetochore fiber and consequent elimination of one or a few chromosomes from the spindle.     

Functional autonomy of monopolar spindle and evidence for oscillatory movement in mitosis

The oscillations of chromosomes associated with a single spindle pole in monocentric and bipolar spindles were analysed by time-lapse cinematography in mitosis of primary cultures of lung epithelium from the newt Taricha granulosa. Chromosomes oscillate toward and away from the pole in all stages of mitosis including anaphase. The duration, velocity, and amplitude of such oscillations are the same in all stages of mitosis. The movement away from the pole in monocentric spindle is rapid enough to suggest the existence of a previously unrecognized active component in chromosome movement, presumably resulting from a pushing action of the kinetochore fiber. During prometaphase oscillations, chromosomes may approach the pole even more closely than at the end of anaphase. Together, these observations demonstrate that a monopolar spindle is sufficient to generate the forces for chromosome transport, both toward and away from the pole. The coordination of the aster/centrosome migration in prophase with the development of the kinetochore fibers determines the course of mitosis. After the breaking of the nuclear envelope in normal mitosis, aster/centrosome separation is normally followed by the rapid formation of bipolar chromosomal fibers. There are two aberrant extremes that may result from a failure in coordination between these processes: (a) A monocentric spindle will arise when aster separation does not occur, and (b) an anaphaselike prometaphase will result if the aster/centrosomal complexes are already well-separated and bipolar chromosomal fibers do not form. In the latter case, the two monopolar prometaphase half-spindles migrate apart, each containing a random number of two chromatid (metaphase) monopolar-oriented chromosomes. This random segregation of prometaphase chromosome displays many features of a standard anaphase and may be followed by a false cleavage. The process of polar separation during prometaphase occurs without any visible interzonal structures. Aster/centrosomes and monopolar spindles migrate autonomously by an unknown mechanism. There are, however, firm but transitory connections between the aster center and the kinetochores as demonstrated by the occasional synchrony of centrosome-kinetochore movement. The data suggest that aster motility is important in the progress of both prometaphase and anaphase in normal mitosis.     

Mitosis in Tilia americana endosperm

The endosperm cells of the American basswood Tilia americana are favorable experimental material for investigating the birefringence of living plant spindles and anaphase movement of chromosomes. The behavior of the chromosomes in anaphase and the formation of the phragmoplast are unique. The numerous (3 n equals 123), small chromosomes move in precise, parallel rows until midanaphase when they bow away from the poles. Such a pattern of anaphase chromosome distribution has been described once before, but was ascribed to fusion of the chromosomes. The bowing of chromosome rows in Tilia is explainable quantitatively by the constant poleward velocity of the chromosomes during anaphase. Peripheral chromosomes are moving both relative to the spindle axis and laterally closer to the axis, whereas chromosomes lying on the spindle axis possess no lateral component in their motion, and thus at uniform velocity progress more rapidly than peripheral chromosomes relative to the spindle axis. The chromosomes are moved poleward initially by pole-to-pole elongation of the spindle, then moved farther apart by shortening of the kinetochore fibers. In contrast to other plant cells where the phragmoplast forms in telophase, the phragmoplast in Tilia endosperm is formed before midanaphase and the cell during midanaphase, while the chromosomes are still in poleward transit.     

Directional instability of kinetochore motility during chromosome congression and segregation in mitotic newt lung cells: a push-pull mechanism

Most models of mitotic congression and segregation assume that only poleward pulling forces occur at kinetochores. However, there are reports for several different cell types that both mono-oriented and bi- oriented chromosomes oscillate toward and away from the pole throughout mitosis. We used new methods of high resolution video microscopy and computer-assisted tracking techniques to measure the positions over time of individual kinetochores with respect to their poles during mitosis in living newt lung cells. The results show that kinetochores oscillate throughout mitosis when they are tethered to spindle poles by attachment to the plus-ends of kinetochore microtubules (kMTs). Oscillations were not sinusoidal. Instead, kinetochores abruptly (as quick as 6 s or less) switched between persistent (approximately 1.5 min average duration) phases of poleward (P) and away from the pole (AP) movement. This kinetochore "directional instability" was a property of motility at the plus-ends of kMTs since fluorescent marks on the lattice of kMTs have previously been observed to exhibit only relatively slow P movement. Each P and AP phase consisted of one or a few constant velocity domains (approximately 1.7 microns/min average velocity). Velocities of P and AP phases were similar from prometaphase through mid-anaphase. Kinetochores occasionally switched to an indeterminant (N) phase of no or confused motion, which was usually brief compared to the durations of P and AP phases. Net chromosome displacements that occurred during congression to the equator or poleward movement during anaphase were primarily generated by differences in the durations and not the velocities of P and AP movements. Careful analysis of centromere deformation showed that kinetochore P movement produced pulling forces while kinetochore AP movement produced pushing forces. These data show that kinetochore directional instability is fundamental to the processes of chromosome congression and segregation. We argue that tension at the kinetochore attachment site is a key factor which controls the switching between P and AP phases of kinetochore motion.     

Pac-Man does not resolve the enduring problem of anaphase chromosome movement

Summary The Pac-Man hypothesis suggests that poleward movement of chromosomes during anaphase A is brought about by: disassembly of kinetochore microtubules (MTs) at the kinetochore; generation of the poleward force exclusively at or very close to the kinetochore; and the required energy coming from coupled disassembly of these MTs. This model has become widely accepted and cited as the sole or major mechanism of anaphase A. Rarely acknowledged are several significant phenomena that refute some or all of these postulates. We summarise these anomalies as follows: poleward movement of chromosomes occurring without insertion of any MTs at the kinetochore; anaphase shortening of kinetochore fibres in spindles entirely devoid of chromosomes and, presumably, kinetochores; continued movement of chromosomes while their severed kinetochore stub elongated poleward after treatment with UV microbeams; and fluxing of tubulin subunits through kinetochore MTs during anaphase A, indicating that during anaphase, kinetochore MTs disassemble partly or solely at the poles.Dedicated to Professor Brian E. S. Gunning on the occasion of his 65th birthday     

Motile kinetochores and polar ejection forces dictate chromosome position on the vertebrate mitotic spindle

We argue that hypotheses for how chromosomes achieve a metaphase alignment, that are based solely on a tug-of-war between poleward pulling forces produced along the length of opposing kinetochore fibers, are no longer tenable for vertebrates. Instead, kinetochores move themselves and their attached chromosomes, poleward and away from the pole, on the ends of relatively stationary but shortening/elongating kinetochore fiber microtubules. Kinetochores are also "smart" in that they switch between persistent constant-velocity phases of poleward and away from the pole motion, both autonomously and in response to information within the spindle. Several molecular mechanisms may contribute to this directional instability including kinetochore-associated microtubule motors and kinetochore microtubule dynamic instability. The control of kinetochore directional instability, to allow for congression and anaphase, is likely mediated by a vectorial mechanism whose magnitude and orientation depend on the density and orientation or growth of polar microtubules. Polar microtubule arrays have been shown to resist chromosome poleward motion and to push chromosomes away from the pole. These "polar ejection forces" appear to play a key role in regulating kinetochore directional instability, and hence, positions achieved by chromosomes on the spindle.     

Polar ejection forces are operative in crane-fly spermatocytes, but their action is limited to the spindle periphery.

Laser microsurgery was employed to reveal kinetochore-independent forces acting on chromosome arms in crane-fly spermatocytes. When a portion of an arm situated along the interpolar axis between the equator and a pole was cut off, the resultant acentric fragment was transported poleward and outward into the peripheral domain of the spindle. If the fragment was generated well in advance of the onset of anaphase, then at the spindle periphery, it changed direction and moved away from the pole and back toward the equator. That domain-specific movement-poleward in the central spindle and away from the pole at the spindle periphery-not only provides the first evidence for polar ejection forces acting on acentric fragments in a meiotic system, but it is the first example of kinetochore-independent forces in both directions at the same stage of division. Sniglets generated by laser pulses directed at specific sites in the spindle revealed that the mechanism underlying ejection forces was specific to chromosomes. At anaphase onset, polar ejection forces ceased, and pole-directed forces took over. At that time, chromosome fragments that had been ejected to the equator moved poleward again, providing clear evidence for kinetochore-independent forces on chromosome arms during anaphase.     

Chromosome micromanipulation

Two types of unusual motion within the spindle have heen studied in a grasshopper (Melanoplus differentialis) spermatocyte. The first is the motion of granules placed by micromanipulation within the normally granule-free spindle. The most specific motions are poleward, approximate the speed of the chromosomes in anaphase, and occur in the area between the kinetochores and the nearer pole during both metaphase and anaphase. Exactly the same transport properties were earlier observed by Bajer inHaemanthus endosperm spindles. The absence of significant motion in the interzone between the separating chromosomes at anaphase has been unequivocally demonstrated inMelanoplus spermatocytes. Thus very specific motion of non-kinetochoric materials is probably a general spindle capability which would much restrict admissible models of mitotic force production,if the same forces move both granules and chromosomes. The second unusual motion is seen following chromosome detachment from the spindle by micromanipulation during anaphase. These tend to move toNearer pole rather than to the pole the chromosome's kinetochoresFace. The latter preference was earlier demonstrated after detachment during prometaphase or metaphase and has been confirmed without exception in the present studies. The apparent preference for motion to the nearer pole in anaphase provides the first evidence for poleward forces within each half-spindle which cannot be entirely specified by the chromosomal spindle fibers. Almost certainly these would be the usual forces responsible for chromosome motion since they act specifically at the kinetochores of detached chromosomes. This evidence requires interpretation, however because additional factors influence chromosome motion following detachment at anaphase. On thesimplest interpretation, certain current models of mitosis clearly are not satisfactory and others are favored.     

Functional redundancy in the maize meiotic kinetochore

Kinetochores can be thought of as having three major functions in chromosome segregation: (a) moving plateward at prometaphase; (b) participating in spindle checkpoint control; and (c) moving poleward at anaphase. Normally, kinetochores cooperate with opposed sister kinetochores (mitosis, meiosis II) or paired homologous kinetochores (meiosis I) to carry out these functions. Here we exploit three- and four-dimensional light microscopy and the maize meiotic mutant absence of first division 1 (afd1) to investigate the properties of single kinetochores. As an outcome of premature sister kinetochore separation in afd1 meiocytes, all of the chromosomes at meiosis II carry single kinetochores. Approximately 60% of the single kinetochore chromosomes align at the spindle equator during prometaphase/metaphase II, whereas acentric fragments, also generated by afd1, fail to align at the equator. Immunocytochemistry suggests that the plateward movement occurs in part because the single kinetochores separate into half kinetochore units. Single kinetochores stain positive for spindle checkpoint proteins during prometaphase, but lose their staining as tension is applied to the half kinetochores. At anaphase, approximately 6% of the kinetochores develop stable interactions with microtubules (kinetochore fibers) from both spindle poles. Our data indicate that maize meiotic kinetochores are plastic, redundant structures that can carry out each of their major functions in duplicate.     

Efficient mitosis in human cells lacking poleward microtubule flux

Chromosome segregation relies on the dynamic properties of spindle microtubules (MTs). Poleward MT flux contributes to spindle dynamics through the disassembly of MT minus ends at spindle poles coupled to the continuous poleward transport of spindle MTs. Despite being conserved in metazoan cells, the function of flux remains controversial because flux rates differ widely in different cell types. In meiotic systems, the rate of flux nearly matches that of chromosome movement, but in mitotic systems, flux is significantly slower than chromosome movement. Here, we show that spindles in human mitotic cells depleted of the kinesin-13 proteins Kif2a and MCAK lack detectable flux and that such cells frequently fail to segregate all chromosomes appropriately at anaphase. Elimination of flux reduces poleward chromosome velocity approximately 20%, but does not hinder bipolar spindle assembly, chromosome alignment, or mitotic progression. Thus, mitosis proceeds efficiently in human cells lacking detectable poleward MT flux. These data demonstrate that in human cultured cells, kinetochores are sufficient to effectively power chromosome movement, leading us to speculate that flux is maintained in these cells to fulfill other functional roles such as error correction or kinetochore regulation.     

The chromokinesin Kid is necessary for chromosome arm orientation and oscillation, but not congression, on mitotic spindles

Chromokinesins have been postulated to provide the polar ejection force needed for chromosome congression during mitosis. We have evaluated that possibility by monitoring chromosome movement in vertebrate-cultured cells using time-lapse differential interference contrast microscopy after microinjection with antibodies specific for the chromokinesin Kid. 17.5% of cells injected with Kid-specific antibodies have one or more chromosomes that remain closely opposed to a spindle pole and fail to enter anaphase. In contrast, 82.5% of injected cells align chromosomes in metaphase, progress to anaphase, and display chromosome velocities not significantly different from control cells. However, injected cells lack chromosome oscillations, and chromosome orientation is atypical because chromosome arms extend toward spindle poles during both congression and metaphase. Furthermore, chromosomes cluster into a mass and fail to oscillate when Kid is perturbed in cells containing monopolar spindles. These data indicate that Kid generates the polar ejection force that pushes chromosome arms away from spindle poles in vertebrate-cultured cells. This force increases the efficiency with which chromosomes make bipolar spindle attachments and regulates kinetochore activities necessary for chromosome oscillation, but is not essential for chromosome congression.     

Anaphase A Chromosome Movement and Poleward Spindle Microtubule Flux Occur At Similar Rates in Xenopus Extract Spindles

We have used local fluorescence photoactivation to mark the lattice of spindle microtubules during anaphase A in Xenopus extract spindles. We find that both poleward spindle microtubule flux and anaphase A chromosome movement occur at similar rates (~2 μm/min). This result suggests that poleward microtubule flux, coupled to microtubule depolymerization near the spindle poles, is the predominant mechanism for anaphase A in Xenopus egg extracts. In contrast, in vertebrate somatic cells a “Pacman” kinetochore mechanism, coupled to microtubule depolymerization near the kinetochore, predominates during anaphase A. Consistent with the conclusion from fluorescence photoactivation analysis, both anaphase A chromosome movement and poleward spindle microtubule flux respond similarly to pharmacological perturbations in Xenopus extracts. Furthermore, the pharmacological profile of anaphase A in Xenopus extracts differs from the previously established profile for anaphase A in vertebrate somatic cells. The difference between these profiles is consistent with poleward microtubule flux playing the predominant role in anaphase chromosome movement in Xenopus extracts, but not in vertebrate somatic cells. We discuss the possible biological implications of the existence of two distinct anaphase A mechanisms and their differential contributions to poleward chromosome movement in different cell types.     

Kinetochore dynein generates a poleward pulling force to facilitate congression and full chromosome alignment

For proper chromosome segregation, all kinetochores must achieve bipolar microtubule （MT） attachment and subsequently align at the spindle equator before anaphase onset. The MT minus end-directed motor dynein/dynactin binds kinetoehores in prometaphase and has long been implicated in chromosome congression. Unfortunately, inactivation of dynein usually disturbs spindle organization, thus hampering evaluation of its kinetochore roles. Here we specifically eliminated kinetochore dynein/dynactin by RNAi-mediated depletion of ZW10, a protein essential for kinetochore localization of the motor. Time-lapse microscopy indicated markedly-reduced congression efficiency, though congressing chromosomes displayed similar velocities as in control cells. Moreover, cells frequently failed to achieve full chromosome alignment, despite their normal spindles. Confocal microcopy revealed that the misaligned kinetochores were monooriented or unattached and mostly lying outside the spindle, suggesting a difficulty to capture MTs from the opposite pole. Kinetoehores on monoastral spindles were dispersed farther away from the pole and exhibited only mild oscillation. Furthermore, inactivating dynein by other means generated similar phenotypes. Therefore, kinetochore dynein produces on monooriented kinetochores a poleward pulling force, which may contribute to efficient bipolar attachment by facilitating their proper microtubule captures to promote congression as well as full chromosome alignment.     

Quartet: a Drosophila developmental mutation affecting chromosome separation in mitosis

The Drosophila mutation, quartet, affects development at points in the life cycle that require intense mitotic activity. Examination of embryos affected by the maternal effect of quartet has revealed defects that can be attributed to incomplete chromosome separation at mitosis. These defects include uneven spacing of nuclei, strands of DNA creating bridges between nuclei, and abnormal amounts of DNA per nucleus. Nuclei in quartet-affected embryos also have a greater-than-normal number of centrosomes. Immunofluorescent examination of the spindles in quartet-affected embryos has revealed tripolar spindles and adjacent spindles that share a common spindle pole. Finally, chromosome separation distance was measured in anaphase and telophase spindles in quartet-affected embryos and found to be blocked in anaphase. Examination of mitotic figures in quartet larvae revealed a reduced mitotic index and an elevated frequency of abnormal mitotic figures. quartet could encode a function necessary for the disengagement of chromosomes in mitosis, for kinetochore function or for function of a spindle motor. Mutations in quartet prevent the post-translational modification of three abundant proteins. These proteins may be involved in chromosome separation in mitosis.     

CLASPs prevent irreversible multipolarity by ensuring spindle-pole resistance to traction forces during chromosome alignment

Loss of spindle-pole integrity during mitosis leads to multipolarity independent of centrosome amplification. Multipolar-spindle conformation favours incorrect kinetochore-microtubule attachments, compromising faithful chromosome segregation and daughter-cell viability. Spindle-pole organization influences and is influenced by kinetochore activity, but the molecular nature behind this critical force balance is unknown. CLASPs are microtubule-, kinetochore- and centrosome-associated proteins whose functional perturbation leads to three main spindle abnormalities: monopolarity, short spindles and multipolarity. The first two reflect a role at the kinetochore-microtubule interface through interaction with specific kinetochore partners, but how CLASPs prevent spindle multipolarity remains unclear. Here we found that human CLASPs ensure spindle-pole integrity after bipolarization in response to CENP-E- and Kid-mediated forces from misaligned chromosomes. This function is independent of end-on kinetochore-microtubule attachments and involves the recruitment of ninein to residual pericentriolar satellites. Distinctively, multipolarity arising through this mechanism often persists through anaphase. We propose that CLASPs and ninein confer spindle-pole resistance to traction forces exerted during chromosome congression, thereby preventing irreversible spindle multipolarity and aneuploidy.