Genetic control of mitosis. Mast(v40) protein--an element of checkpoints system?

The effect of the mastv40 mutation was studied using neural ganglion cells of third-instar larvae of Drosophila melanogaster. The distributions of the cells by the interphase nucleus diameter and by the distance between the sister chromosome sets in anaphase were analyzed. Three following types of defects induced by the mutation were described: (1) Monopolar mitosis or, in the case of bipolar mitosis, an abnormally short distance between the sister chromosome sets in anaphase and early telophase. We believe that these abnormalities are caused by damage of the start and (or) motor mechanisms of centrosome separation at the beginning and in the end of mitosis. (2) Lagging and bridging of chromosomes in anaphase and early telophase. These defects seem to be related to the disruption of functioning of mitotic spindle microtubules and (or) their defective attachment to the appropriate kinetochores. (3) Unlimited division of aneuploid and polyploid cells, which may be explained either by inactivation of the checkpoint system controlling the genome ploidy or by checkpoint adaptation. Taken collectively, our results and literature data suggest that the MAST protein is an element of the checkpoint system and that division of aneuploid and polyploid cells results from inactivation of the checkpoints.     

The Drosophila maternal-effect gene fs(1)Ya encodes a cell cycle-dependent nuclear envelope component required for embryonic mitosis

The maternal-effect gene fs(1)Ya is specifically required for embryonic mitosis in Drosophila. fs(1)Ya is involved in the initiation of the first embryonic mitosis and may also be necessary for subsequent embryonic mitotic divisions. fs(1)Ya encodes a 91.3 kd hydrophilic protein containing two putative MPF phosphorylation target sites and two potential nuclear localization signals. This protein is synthesized during postoogenic maturation from its maternal RNA and persists throughout embryogenesis. In early embryos, the fs(1)Ya protein is localized to the nuclear envelope from interphase to metaphase. During anaphase and telophase, it is dispersed in the nucleoplasm and cytoplasm, a behavior that is different from that of both the nuclear envelope and lamins. These results suggest that the fs(1)Ya protein is a cell cycle-dependent component of the nuclear envelope that specifically functions in embryonic mitosis.     

Timing of centrosome separation is important for accurate chromosome segregation

Spindle assembly, establishment of kinetochore attachment, and sister chromatid separation must occur during mitosis in a highly coordinated fashion to ensure accurate chromosome segregation. In most vertebrate cells, the nuclear envelope must break down to allow interaction between microtubules of the mitotic spindle and the kinetochores. It was previously shown that nuclear envelope breakdown (NEB) is not coordinated with centrosome separation and that centrosome separation can be either complete at the time of NEB or can be completed after NEB. In this study, we investigated whether the timing of centrosome separation affects subsequent mitotic events such as establishment of kinetochore attachment or chromosome segregation. We used a combination of experimental and computational approaches to investigate kinetochore attachment and chromosome segregation in cells with complete versus incomplete spindle pole separation at NEB. We found that cells with incomplete spindle pole separation exhibit higher rates of kinetochore misattachments and chromosome missegregation than cells that complete centrosome separation before NEB. Moreover, our mathematical model showed that two spindle poles in close proximity do not "search" the entire cellular space, leading to formation of large numbers of syntelic attachments, which can be an intermediate stage in the formation of merotelic kinetochores.     

Anaphase onset in vertebrate somatic cells is controlled by a checkpoint that monitors sister kinetochore attachment to the spindle

To test the popular but unproven assumption that the metaphase-anaphase transition in vertebrate somatic cells is subject to a checkpoint that monitors chromosome (i.e., kinetochore) attachment to the spindle, we filmed mitosis in 126 PtK1 cells. We found that the time from nuclear envelope breakdown to anaphase onset is linearly related (r2 = 0.85) to the duration the cell has unattached kinetochores, and that even a single unattached kinetochore delays anaphase onset. We also found that anaphase is initiated at a relatively constant 23-min average interval after the last kinetochore attaches, regardless of how long the cell possessed unattached kinetochores. From these results we conclude that vertebrate somatic cells possess a metaphase-anaphase checkpoint control that monitors sister kinetochore attachment to the spindle. We also found that some cells treated with 0.3-0.75 nM Taxol, after the last kinetochore attached to the spindle, entered anaphase and completed normal poleward chromosome motion (anaphase A) up to 3 h after the treatment--well beyond the 9-48-min range exhibited by untreated cells. The fact that spindle bipolarity and the metaphase alignment of kinetochores are maintained in these cells, and that the chromosomes move poleward during anaphase, suggests that the checkpoint monitors more than just the attachment of microtubules at sister kinetochores or the metaphase alignment of chromosomes. Our data are most consistent with the hypothesis that the checkpoint monitors an increase in tension between kinetochores and their associated microtubules as biorientation occurs.     

Genetic Control of Mitosis: Is Protein MASTν40 an Element of the Checkpoint System?

The effect of the mast ^v40 mutation was studied using neural ganglion cells of third-instar larvae of Drosophila melanogaster. The distributions of the cells by the interphase nucleus diameter and by the distance between the sister chromosome sets in anaphase were analyzed. Three following types of defects induced by the mutation were described: (1) Monopolar mitosis or, in the case of bipolar mitosis, an abnormally short distance between the sister chromosome sets in anaphase and early telophase. We suppose that these abnormalities are caused by damage of the start and (or) motor mechanisms of centrosome separation at the beginning and in the end of mitosis. (2) Lagging and bridging of chromosomes in anaphase and early telophase. These defects seem to be related to the disruption of functioning of mitotic spindle microtubules and (or) their defective attachment to the appropriate kinetochores. (3) Unlimited division of aneuploid and polyploid cells, which may be explained either by inactivation of the checkpoint system controlling the genome ploidy or by checkpoint adaptation. Taken collectively, our results and literature data suggest that the MAST protein is an element of the checkpoint system and that division of aneuploid and polyploid cells results from inactivation of the checkpoints.     

Dynamic distribution of TTK in HeLa cells： insights from an ultrastructural study

Entry into mitosis is driven by signaling cascades of mitotic kinases. Our recent studies show that TTK, a kinetochore-associated protein kinase, interacts with CENP-E, a mitotic kinesin located to corona fiber of kinetochore. Using immunoelectron microscopy, here we show that TTK is present at the nuclear pore adjacent complex of interphase HeLa cells. Upon nuclear envelope fragmentation, TTK targets to the outermost region of the developing kinetochores of monoorient chromosome as well as to spindle poles. After stable attachment, throughout chromosome congression, TTK is a constituent of the corona fibers, extending up to 90 nm away from the kinetochore outer plate. Upon metaphase alignment, TTK departs from the kinetochore and migrates toward the centrosomes. Taken together, this evidence strongly supports a model in which TTK functions in spindle checkpoint signaling cascades at both kinetochore and centrosome.     

Origin of the mitotic spindle in onion root cells

Summary Immunofluorescence studies on microtubule arrangement during the transition from prophase to metaphase in onion root cells are presented. The prophase spindle observed at late preprophase and prophase is composed of microtubules converged at two poles near the nuclear envelope; thin bundles of microtubules are tracable along the nuclear envelope. Prior to nuclear envelope breakdown diffuse tubulin staining occurs within the prophase nuclei. During nuclear envelope breakdown the prophase spindle is no longer identifiable and prominent tubulin staining occurs among the prometaphase chromosomes. Patches of condensed tubulin staining are observed in the vicinity of kinetochores. At advanced prometaphase kinetochore bundles of microtubules are present in some kinetochore regions. At metaphase the mitotic spindle is mainly composed of kinetochore bundles of microtubules; pole-to-pole bundles are scarce. Our observations suggest that the prophase spindle is decomposed at the time of nuclear envelope breakdown and that the metaphase spindle is assembled at prometaphase, with the help of kinetochore nucleating action.     

The GTPase Ran regulates chromosome positioning and nuclear envelope assembly in vivo

The GTPase Ran is known to regulate transport of proteins across the nuclear envelope. Recently, Ran has been shown to promote microtubule polymerization and spindle assembly around chromatin in Xenopus mitotic extracts and to stimulate nuclear envelope assembly in Xenopus or HeLa cell extracts. However, these in vitro findings have not been tested in living cells and do not necessarily describe the generalized model of Ran functions. Here we present several lines of evidence that Ran is indispensable for correct chromosome positioning and nuclear envelope assembly in C. elegans. Embryos deprived of Ran by RNAi showed metaphase chromosome misalignment and aberrant chromosome segregation, while astral microtubules seemed unaffected. Depletion of RCC1 or RanGAP by RNAi resulted in essentially the same defects. The immunofluorescent staining showed that Ran localizes to kinetochore regions of metaphase and anaphase chromosomes, suggesting the role of Ran in linking chromosomes to kinetochore microtubules. Ran was shown to localize to the nuclear envelope at telophase and during interphase in early embryos, and the depletion of Ran resulted in failure of nuclear envelope assembly. Thus, Ran is crucially involved in chromosome positioning and nuclear envelope assembly in C. elegans.     

HURP controls spindle dynamics to promote proper interkinetochore tension and efficient kinetochore capture

Through a functional genomic screen for mitotic regulators, we identified hepatoma up-regulated protein (HURP) as a protein that is required for chromosome congression and alignment. In HURP-depleted cells, the persistence of unaligned chromosomes and the reduction of tension across sister kinetochores on aligned chromosomes resulted in the activation of the spindle checkpoint. Although these defects transiently delayed mitotic progression, HeLa cells initiated anaphase without resolution of these deficiencies. This bypass of the checkpoint arrest provides a tumor-specific mechanism for chromosome missegregation and genomic instability. Mechanistically, HURP colocalized with the mitotic spindle in a concentration gradient increasing toward the chromosomes. HURP binds directly to microtubules in vitro and enhances their polymerization. In vivo, HURP stabilizes mitotic microtubules, promotes microtubule polymerization and bipolar spindle formation, and decreases the turnover rate of the mitotic spindle. Thus, HURP controls spindle stability and dynamics to achieve efficient kinetochore capture at prometaphase, timely chromosome congression to the metaphase plate, and proper interkinetochore tension for anaphase initiation.     

Mitosis of Micronuclei During Division and Regeneration in the Ciliate Stentor coeruleus1

Stages of mitosis of the micronuclei of Stentor coeruleus were described as seen by transmission electron microscopy. Cells in division and those regenerating new oral membranelles were studied. Microtubules were found in early prophase in the karyoplasm and interspersed between the condensing chromatin. A monaxial intranuclear spindle is formed by early metaphase, with kinetochore microtubule attachment sites on the chromosomes. The spindle elongates, separating the daughter nuclei at anaphase. A new nuclear envelope, consisting of two unit membranes, begins to form at late anaphase. Small segments of membrane found in the space between the newly forming and the old micronuclear envelopes appear to fuse to form the new nuclear envelope. No ultrastructural differences were found in the mitotic nuclei of cells in division or regeneration.     

Nuclear and microtubular cycles in heterophasic multinuclearTriticum root-tip cells induced by caffeine

Summary Nuclear and microtubular cycles were studied in large heterophasic multinuclear cells induced in root tips ofTriticum turgidum by caffeine treatment. Multinuclear cells and cells with polyploid nuclei exhibited various configurations of multiple and complex preprophase microtubule (Mt) bands (PPBs), including helical ones. The developmental stages of PPBs in some heterophasic cells did not comply with the cell cycle stages of the associated nuclei, a fact indicating that these events are not directly controlled by the associated nuclei. The heterophasic cells exhibited asynchronous nuclei at different stages of mitosis. In cells displaying prophase and interphase nuclei, the prophase spindle was either absent or developed around both of them or developed around the prophase nuclei earlier than around the interphase ones. During prometaphase-metaphase of the advanced nuclei the lagging interphase nuclei were induced to form prematurely condensed chromosomes (PCCs) along with spindle formation around them. These observations suggest that the mitotic transition in heterophasic cells is delayed but is ultimately achieved due to the effect of the advanced nuclei, which induces a premature mitotic entry of the lagging nuclei. Although kinetochore Mt bundles were found associated with PCCs, their metaphase and anaphase spindles were abnormal resulting in abnormal or abortive anaphases. In some heterophasic cells, metaphase-anaphase transition did not take place simultaneously in different chromosome groups, signifying that the cells do not exit from the mitotic state after anaphase initiation of the advanced nuclei. Asynchronous pace of mitosis of different chromosome groups was also observed during anaphase and telophase. Implications of these observations in understanding plant cell cycle regulation are discussed.Abbreviations cdk cyclin dependent kinase - Mt microtubule - PCC prematurely condensed chromosome - PPB preprophase band     

Cell cycle-dependent dynamics and regulation of mitotic kinesins in Drosophila S2 cells

Constructing a mitotic spindle requires the coordinated actions of several kinesin motor proteins. Here, we have visualized the dynamics of five green fluorescent protein (GFP)-tagged mitotic kinesins (class 5, 6, 8, 13, and 14) in live Drosophila Schneider cell line (S2), after first demonstrating that the GFP-tag does not interfere with the mitotic functions of these kinesins using an RNA interference (RNAi)-based rescue strategy. Class 8 (Klp67A) and class 14 (Ncd) kinesin are sequestered in an active form in the nucleus during interphase and engage their microtubule targets upon nuclear envelope breakdown (NEB). Relocalization of Klp67A to the cytoplasm using a nuclear export signal resulted in the disassembly of the interphase microtubule array, providing support for the hypothesis that this kinesin class possesses microtubule-destabilizing activity. The interactions of Kinesin-5 (Klp61F) and -6 (Pavarotti) with microtubules, on the other hand, are activated and inactivated by Cdc2 phosphorylation, respectively, as shown by examining localization after mutating Cdc2 consensus sites. The actions of microtubule-destabilizing kinesins (class 8 and 13 [Klp10A]) seem to be controlled by cell cycle-dependent changes in their localizations. Klp10A, concentrated on microtubule plus ends in interphase and prophase, relocalizes to centromeres and spindle poles upon NEB and remains at these sites throughout anaphase. Consistent with this localization, RNAi analysis showed that this kinesin contributes to chromosome-to-pole movement during anaphase A. Klp67A also becomes kinetochore associated upon NEB, but the majority of the population relocalizes to the central spindle by the timing of anaphase A onset, consistent with our RNAi result showing no effect of depleting this motor on anaphase A. These results reveal a diverse spectrum of regulatory mechanisms for controlling the localization and function of five mitotic kinesins at different stages of the cell cycle.     

The astrin-kinastrin/SKAP complex localizes to microtubule plus ends and facilitates chromosome alignment

Astrin is a mitotic spindle-associated protein required for the correct alignment of all chromosomes at the metaphase plate. Astrin depletion delays chromosome alignment and causes the loss of normal spindle architecture and sister chromatid cohesion before anaphase onset. Here we describe an astrin complex containing kinastrin/SKAP, a novel kinetochore and mitotic spindle protein, and three minor interaction partners: dynein light chain, Plk1, and Sgo2. Kinastrin is the major astrin-interacting protein in mitotic cells, and is required for astrin targeting to microtubule plus ends proximal to the plus tip tracking protein EB1. Cells overexpressing or depleted of kinastrin mislocalize astrin and show the same mitotic defects as astrin-depleted cells. Importantly, astrin fails to localize to and track microtubule plus ends in cells depleted of or overexpressing kinastrin. These findings suggest that microtubule plus end targeting of astrin is required for normal spindle architecture and chromosome alignment, and that perturbations of this pathway result in delayed mitosis and nonphysiological separase activation.     

New insights into the regulation of anaphase by mitotic cyclins in budding yeast

Mitotic cyclins drive initiation and progression through mitosis. However, their role during progression remains poorly understood due to their essential function in initiation of mitosis and redundant activities. The function of the principal mitotic cyclin, Clb2, in S. cerevisiae, was investigated during progression through anaphase in diploid cells after DNA damage and during normal growth using fixed and live cell fluorescence techniques. I find that during anaphase, absence of Clb2 affects chromosome movement and plays an important role in inhibiting kinetochore microtubules regrowth. In addition, absence of Clb2 leads to defects and the collapse of spindle pole body separation. Most unexpectedly, new bipolar spindle forms and spindle re-forms. The intensity of the defects appears to correlate with strength of checkpoint activation, and during adaptation to DNA damage, these defects lead to important chromosome missegregation, during normal growth, defects are resolved rapidly. During recovery, intermediate phenotypes are observed. Altogether, data reveal new and unexpected roles for mitotic cyclins during progression through mitosis; results indicate that mitotic cyclins play key role in growth suppression of kinetochore microtubules and suggest that new bipolar spindle formation might be actively inhibited by mitotic cyclins during anaphase.     

Merotelic kinetochore orientation is a major mechanism of aneuploidy in mitotic mammalian tissue cells

In mitotic cells, an error in chromosome segregation occurs when a chromosome is left near the spindle equator after anaphase onset (lagging chromosome). In PtK1 cells, we found 1.16% of untreated anaphase cells exhibiting lagging chromosomes at the spindle equator, and this percentage was enhanced to 17.55% after a mitotic block with 2 microM nocodazole. A lagging chromosome seen during anaphase in control or nocodazole-treated cells was found by confocal immunofluorescence microscopy to be a single chromatid with its kinetochore attached to kinetochore microtubule bundles extending toward opposite poles. This merotelic orientation was verified by electron microscopy. The single kinetochores of lagging chromosomes in anaphase were stretched laterally (1.2--5.6-fold) in the directions of their kinetochore microtubules, indicating that they were not able to achieve anaphase poleward movement because of pulling forces toward opposite poles. They also had inactivated mitotic spindle checkpoint activities since they did not label with either Mad2 or 3F3/2 antibodies. Thus, for mammalian cultured cells, kinetochore merotelic orientation is a major mechanism of aneuploidy not detected by the mitotic spindle checkpoint. The expanded and curved crescent morphology exhibited by kinetochores during nocodazole treatment may promote the high incidence of kinetochore merotelic orientation that occurs after nocodazole washout.     

THE ULTRASTRUCTURE OF A MAMMALIAN CELL DURING THE MITOTIC CYCLE

With a technique of preselecting the mitotic cell in the living state for subsequent electron microscopy, it has been possible to examine the ultrastructure of the various stages of mitosis with greater precision than has been reported previously. The early dissolution of the nuclear envelope has been found to be preceded by a marked undulation of this structure within the nuclear "hof." This undulation appears to be intimately related to the spindle-forming activity of the centriole at this time. Marked pericentriolar osmiophilia and extensive arrays of vesicles are also prominent at this stage, the former continuing into anaphase. Progression of the cell through prophase is accompanied by a disappearance of these vesicles. A complex that first makes its appearance in prophase but becomes most prominent in metaphase is a partially membrane-bounded cluster of dense osmiophilic bodies. These clusters which have a circumferential distribution in the mitotic cell are shown to be derived from multivesicular bodies and are acid phosphatase-positive. The precise selection of cells during the various stages of anaphase has made it possible to follow chronologically the morphological features of the initiation of nuclear membrane reformation. The nuclear membrane appears to be derived from polar aggregates of endoplasmic reticulum, and the process begins less than 2 minutes after the onset of karyokinesis. While formation of the nuclear envelope is initiated on the polar aspects of the chromatin mass, envelope elements appear on the equatorial aspect long before the polar elements fuse. Apparently interfering with this fusion are continuous spindle tubules which traverse the chromatin mass in striking density at characteristic points. Several cortical changes, also most pronounced in anaphase, have been described, as has the kinetochore which is seen to good advantage only in this stage. The Golgi complex has been found to disappear both morphologically and histochemically during mitosis and to reappear rapidly in telophase. Evidence is presented which implicates the continuous spindle tubules in certain phases of chromosome movement.     

1,2-Dioctanoylglycerol accelerates or retards mitotic progression in Tradescantia stamen hair cells as a function of the time of its addition

We have treated living, intact stamen hair cells from the spiderwort plant, Tradescantia virginiana, with 0.5 microgram/ml or 60 micrograms/ml 1,2-dioctanoylglycerol, a potent and permeant activator of protein kinase C, and have observed the rates of progression of mitosis from prophase through anaphase. We have found that in addition to the concentration used, the time of initial treatment with 1,2-dioctanoylglycerol defines the response by the cells. The cells rapidly undergo nuclear envelope breakdown when this diglyceride is added in very late prophase, 0 to approximately 8 min prior to the time of normal nuclear envelope breakdown. Anaphase onset occurs 28 min after nuclear envelope breakdown, rather than after the 33 min interval observed in untreated cells. Rapid progression through metaphase is also observed if cells are treated with 0.5 microgram/ml 1,2-dioctanoylglycerol during prometaphase, up to 15 min after nuclear envelope breakdown. The addition of 0.5 microgram/ml 1,2-dioctanoylglycerol in late metaphase, approximately 26 min after nuclear envelope breakdown, results in sister chromatid separation slightly ahead of its normal time, 33 min after nuclear envelope breakdown, and in precocious cell plate vesicle aggregation, 3-5 min earlier than that observed in untreated cells. Treatment of cells with 60 micrograms/ml of 1,2-dioctanoylglycerol at any point during the interval from 0 to approximately 5 min prior to nuclear envelope breakdown results in precocious entry into anaphase. If cells are treated with either 0.5 microgram/ml or 60 micrograms/ml 1,2-dioctanoylglycerol earlier than 20 min before nuclear envelope breakdown, they do not enter mitosis, but instead revert to interphase without dividing. When 1,2-dioctanoylglycerol is added at other times during mitosis, the rate of subsequent mitotic progression is dramatically slowed; the cells require greater than 55 min to progress from nuclear envelope breakdown to anaphase onset, though once in anaphase, the cells progress onward to cytokinesis at normal rates. Treatments o of cells with 1,3-dioctanoylglycerol at any point during prophase, prometaphase, or metaphase are without effect on the rate of subsequent mitotic progression. The shifts in response by cells treated at specific times with 1,2-dioctanoylglycerol during mid- and late metaphase may be indicative of the existence of one or more regulatory switch points (i.e., checkpoints) just prior to anaphase onset.     

Crm1 is a mitotic effector of Ran-GTP in somatic cells

The Ran GTPase controls multiple cellular processes, including nuclear transport, mitotic checkpoints, spindle assembly and post-mitotic nuclear envelope reassembly. Here we examine the mitotic function of Crm1, the Ran-GTP-binding nuclear export receptor for leucine-rich cargo (bearing nuclear export sequence) and Snurportin-1 (ref. 3). We find that Crm1 localizes to kinetochores, and that Crm1 ternary complex assembly is essential for Ran-GTP-dependent recruitment of Ran GTPase-activating protein 1 (Ran-GAP1) and Ran-binding protein 2 (Ran-BP2) to kinetochores. We further show that Crm1 inhibition by leptomycin B disrupts mitotic progression and chromosome segregation. Analysis of spindles within leptomycin B-treated cells shows that their centromeres were under increased tension. In leptomycin B-treated cells, centromeres frequently associated with continuous microtubule bundles that spanned the centromeres, indicating that their kinetochores do not maintain discrete end-on attachments to single kinetochore fibres. Similar spindle defects were observed in temperature-sensitive Ran pathway mutants (tsBN2 cells). Taken together, our findings demonstrate that Crm1 and Ran-GTP are essential for Ran-BP2/Ran-GAP1 recruitment to kinetochores, for definition of kinetochore fibres and for chromosome segregation at anaphase. Thus, Crm1 is a critical Ran-GTP effector for mitotic spindle assembly and function in somatic cells.     

Kinetochores on chromosomes enclosed within the nuclear envelope

The kinetochore plate which develops after nuclear envelope breakdown in normal cells can be seen to be formed on condensed chromosomes still enclosed in the nuclear envelope in fused multinucleate cells where some nuclei show delayed envelope breakdown caused by nuclear interaction. This suggests that neither nuclear envelope breakdown nor assembly of microtubules is directly related to the formation of the kinetochore plate. Furthermore, it can be clearly observed in these cells that the kinetochores do not have any special association with the nuclear envelope.     

Merotelic kinetochores in mammalian tissue cells

Merotelic kinetochore attachment is a major source of aneuploidy in mammalian tissue cells in culture. Mammalian kinetochores typically have binding sites for about 20-25 kinetochore microtubules. In prometaphase, kinetochores become merotelic if they attach to microtubules from opposite poles rather than to just one pole as normally occurs. Merotelic attachments support chromosome bi-orientation and alignment near the metaphase plate and they are not detected by the mitotic spindle checkpoint. At anaphase onset, sister chromatids separate, but a chromatid with a merotelic kinetochore may not be segregated correctly, and may lag near the spindle equator because of pulling forces toward opposite poles, or move in the direction of the wrong pole. Correction mechanisms are important for preventing segregation errors. There are probably more than 100 times as many PtK1 tissue cells with merotelic kinetochores in early mitosis, and about 16 times as many entering anaphase as the 1% of cells with lagging chromosomes seen in late anaphase. The role of spindle mechanics and potential functions of the Ndc80/Nuf2 protein complex at the kinetochore/microtubule interface is discussed for two correction mechanisms: one that functions before anaphase to reduce the number of kinetochore microtubules to the wrong pole, and one that functions after anaphase onset to move merotelic kinetochores based on the ratio of kinetochore microtubules to the correct versus incorrect pole.