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Lodish H, Berk A, Zipursky SL, et al. Molecular Cell Biology. 4th edition. New York: W. H. Freeman; 2000.

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Molecular Cell Biology. 4th edition.

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Section 19.5Microtubule Dynamics and Motor Proteins during Mitosis

Mitosis is the process that partitions newly replicated chromosomes equally into separate parts of a cell. The last step in the cell cycle, mitosis takes about 1 hour in an actively dividing animal cell. During that period, the cell builds and then disassembles a specialized microtubule structure, the mitotic apparatus. Larger than the nucleus, the mitotic apparatus is designed to attach and capture chromosomes, align the chromosomes, and then separate them, so that the genetic material is evenly partitioned to each daughter cell. Fifteen hours later, the whole process is repeated by the two daughter cells. During interphase, the period between mitoses, chromosomes are decondensed and dispersed throughout the nucleus. Although the chromosomes are not visible by light microscopy during this period, DNA replication occurs during the S (synthesis) phase of interphase (see Figure 13-1).

Figure 19-34 depicts the characteristic series of events that can be observed by light microscopy during mitosis in a eukaryotic cell. Although the events unfold continuously, they are conventionally divided into four substages: prophase,metaphase,anaphase, and telophase. The beginning of mitosis is signaled by the appearance of visible condensed chromosomes, stainable as thin threads inside the nucleus. By late prophase, each chromosome is visible as two identical coiled filaments, the chromatids (often called sister chromatids), held together at a constricted region, the centromere. Each chromatid contains one of the two new daughter DNA molecules produced in the preceding S phase of the cell cycle; thus each cell that enters mitosis has four copies of each chromosomal DNA, designated 4n.

Figure 19-34. The stages of mitosis and cytokinesis in an animal cell.

Figure 19-34

The stages of mitosis and cytokinesis in an animal cell. (Morphological types of chromosomes are distinguished by color.) (a) Interphase: The G2 stage of inter-phase immediately precedes the beginning of mitosis and follows chromosomal DNA replication (more...)

The mitotic cycle of chromosome separation is linked to two checkpoints in the cell cycle: breakdown of the nuclear envelope during late prophase and attachment of the microtubules to the kinetochores of sister chromatids at the metaphase-anaphase transition. We saw in Chapter 13 that the cell cycle, and hence cell replication, is controlled primarily by regulating the initiation of DNA synthesis, entry into mitosis, and chromatid separation. In this section, we focus on the mechanism by which microtubules separate chromosomes during mitosis in a “typical” animal cell. Mistakes in mitosis can lead to missing or extra chromosomes, causing abnormal patterns of development when they occur during embryogenesis and pathologies when they occur after birth. To ensure that mitosis proceeds without errors during the billions of cell divisions that occur in the life span of an organism, a highly redundant mechanism has evolved in which each crucial step is carried out concurrently by microtubule motor proteins and microtubule assembly dynamics.

The Mitotic Apparatus Is a Microtubule Machine for Separating Chromosomes

The mitotic apparatus has no fixed structure: it is constantly changing during mitosis (Figure 19-35). For one brief moment at metaphase, however, when the chromosomes are aligned at the equator of the cell, the mitotic apparatus appears static. We will begin our discussion by examining the structure of the mitotic apparatus at metaphase and then discuss how it first organizes chromosomes during prophase, how it separates chromosomes during anaphase, and how it determines where cells are cleaved during telophase.

Figure 19-35. Fluorescence micrographs showing the organization of chromosomes and microtubules during four mitotic stages.

Figure 19-35

Fluorescence micrographs showing the organization of chromosomes and microtubules during four mitotic stages. Cultured PtK2 fibroblasts were stained with a fluorescent anti-tubulin antibody (green) and the DNA-binding dye ethidium homodimer (purple). (more...)

The mitotic apparatus at metaphase is organized into two parts (Figure 19-36): (1) a central mitotic spindle — a bilaterally symmetrical bundle of microtubules with the overall shape of a football, but divided into opposing halves at the equator of the cell by a plate of metaphase chromosomes — and (2) a pair of asters — a tuft of microtubules at each pole of the spindle.

Figure 19-36. (a) High-voltage electron micrograph of the mitotic apparatus in a metaphase mammalian cell.

Figure 19-36

(a) High-voltage electron micrograph of the mitotic apparatus in a metaphase mammalian cell. To visualize the spindle microtubules more clearly, biotin-tagged anti-tubulin antibodies were added to make microtubules more massive. The large cylindrical (more...)

In each half of the spindle, a single centrosome at the pole organizes three distinct sets of microtubules, whose (−) ends all point toward the centrosome (Figure 19-36b). One set, the astral microtubules, forms the aster; they radiate outward from the centrosome toward the cortex of the cell, where they help position the mitotic apparatus and later help to determine the cleavage plane during cytokinesis. The other two sets of microtubules compose the spindle. The kinetochore microtubules attach to chromosomes at specialized attachment sites on the chromosomes called kinetochores. The third set, polar microtubules, do not interact with chromosomes but instead interdigitate with polar microtubules from the opposite pole.

In an overlap zone at the equator, two types of interactions hold the spindle halves together to form the bilaterally symmetric mitotic apparatus: (1) lateral interactions between the (+) ends of the interdigitating polar microtubules from each pole and (2) end-on interactions between the kinetochore microtubules from each pole and the kinetochores of the sister chromatids.

The spindle pole – aster organization of the mitotic apparatus is basic to mitosis in all organisms, but the appearance of the mitotic apparatus can vary widely. The number of microtubules in a spindle, the overall size of the mitotic apparatus, and the timing and duration of mitotic movements all vary among different organisms. In addition, organisms differ in the length or number of their astral microtubules.

In the common baker’s yeast, S. cerevisiae, for example, mitosis is carried out by a structurally simple mitotic apparatus that lacks centriole-based centrosomes and asters (Figure 19-37). Instead of a centrosome, the microtubules are organized around a spindle pole body, a trilaminated structure located in the nuclear membrane, which does not break down during mitosis. Furthermore, because a yeast cell is small, it does not require well-developed asters to assist in mitosis. Thus, the yeast mitotic apparatus comprises just a spindle, which itself is constructed from a minimal number of kinetochore and polar microtubules. In S. cerevisiae, which possesses 16 chromosomes, the spindle contains 32 kinetochore microtubules plus a few polar microtubules. (At metaphase, each kinetochore is attached to one microtubule; thus each chromosome is attached to two kinetochore microtubules.) Because the spindle pole body is functionally equivalent to, but structurally different from, an animal cell centrosome, the two structures must share centrosomal proteins such as γ-tubulin that act to organize the mitotic spindle.

Figure 19-37. Mitotic apparatus in S. cerevisiae.

Figure 19-37

Mitotic apparatus in S. cerevisiae. In yeast, the nucleus remains intact during mitosis; thus the chromosomes are isolated from direct interaction with the cytosol. (a) Spindle pole bodies, which are attached to the nuclear membrane, organize the spindle (more...)

Mitotic events in plant cells are generally similar to the events observed in animal cells. Although most higher plant cells do not contain visible centrioles, an analogous region of the plant cell acts as a microtubule-organizing center, from which the spindle microtubules radiate (Figure 19-38). Moreover, compared with an animal cell, the shape of a plant cell does not change greatly in mitosis because it is surrounded by a rigid cell wall. During telophase, the new cell membrane and cell wall are formed from membrane vesicles that fuse together in a plane perpendicular to a line separating the two nuclei.

Figure 19-38. Fluorescence micrograph showing arrangement of microtubules (red) and chromosomes (blue) during anaphase in a plant cell.

Figure 19-38

Fluorescence micrograph showing arrangement of microtubules (red) and chromosomes (blue) during anaphase in a plant cell. At this stage, the sister chromatids have separated and are moving toward opposite poles of the spindle. [Courtesy of Andrew Bajer.] (more...)

The Kinetochore Is a Specialized Attachment Site at the Chromosome Centromere

The sister chromatids of a metaphase chromosome are transported to each pole along the kinetochore microtubules. In terms of cargoes delivered on microtubules, a chromosome is a much different piece of baggage than the membrane vesicles and organelles we have discussed in previous sections. Figure 19-39 depicts the attachment of kinetochore microtubules to the kinetochore, a platelike structure lying within the centromere, a small, highly specialized region of the chromosome. (Do not confuse centromere with centrosome: the centromere is a region of the chromosome, while the centrosome is a microtubule-organizing center.) The centromere is recognized as a constriction in the condensed chromosome where the sister chromatids are most closely associated. The location of the centromere and hence that of the kinetochore is directly controlled by a specific sequence of chromosomal DNA termed centromeric DNA (Chapter 9).

Figure 19-39. Centromeric attachment of microtubules.

Figure 19-39

Centromeric attachment of microtubules. (a) Schematic diagram of attachment of kinetochore microtubules to the sister chromatids of a metaphase chromosome. In animals and lower plants, the kinetochore is a three-layer, platelike structure lying within (more...)

The kinetochore is first recognizable during late prophase, after the chromosomes have condensed but well before the mitotic apparatus has assembled. The complex structure of the kinetochore is revealed in ultrathin sections of chromosomes as a stack of disklike plates (see Figure 19-39a, inset). Some of the protein components of kinetochores have been identified by their reaction with antibodies that specifically recognize kinetochores. For unknown reasons, these antibodies are frequently produced by patients suffering from scleroderma (an autoimmune disease of unknown origin that causes fibrosis of connective tissue). Using these scleroderma autoantibodies, researchers have identified four proteins that are localized to the inner layer of the mammalian kinetochore. Studies with other antibodies have detected cytosolic dynein and a kinesin-related protein, CENP-E, in the fibrous corona on the surface of the kinetochore.

Our understanding of how kinetochores link centromeres to microtubules is most advanced in yeast. In Chapter 9, we described how centromeric (CEN) DNA sequences from yeast chromosomes can be identified by their ability to make self-replicating plasmids into artificial chromosomes that can be passed from mother to daughter at mitosis (see Figure 9-40). Sequence analysis of cloned CEN DNAs from yeast chromosomes reveals that they are generally organized into three regions, denoted CDEs, or centromere DNA elements, I, II, and III (see Figure 9-41). Of the three regions, mutational analysis implicates CDE III as the most critical for centromere function. This region appears to interact with microtubules via centromere-binding factor (CBF3), a multiprotein subunit complex, and other identified proteins (Figure 19-39b).

Centrosome Duplication Precedes and Is Required for Mitosis

Since each half of the metaphase mitotic apparatus emanates from a polar centrosome, its assembly depends on duplication of the centrosome and their movement to opposite halves of the cell. This process, known as the centriole cycle (or centrosome cycle) begins during G1 when the centrioles and other centrosome components are duplicated (Figure 19-40). By G2 the two “daughter” centrioles have reached full length, but the duplicated centrioles are still present within a single centrosome. Early in mitosis, the two pairs of centrioles separate and migrate to opposite sides of the nucleus, establishing the bipolarity of the dividing cell (see Figure 19-40). In some respects, then, mitosis can be understood as the migration of duplicated centrosomes, which along their journey pick up chromosomes, pause in metaphase, and during anaphase continue their movement to new locations in the daughter cells, where they release the chromosomes and organize the cytosolic microtubule.

Figure 19-40. Relation of centrosome duplication to the cell cycle.

Figure 19-40

Relation of centrosome duplication to the cell cycle. After the pair of parent centrioles (red) within the centrosome matrix separate slightly, a daughter centriole (blue) buds from each and elongates. By G2 growth of the daughter centrioles is complete, (more...)

Dynamic Instability of Microtubules Increases during Mitosis

The observation of high tubulin turnover in microtubules during mitosis is strong evidence that microtubule dynamics is critical to the mitotic process. During mitosis, the first indication of a change in microtubule stability occurs at prophase, when long interphase microtubules disappear and are replaced by a spindle and astral microtubules (Figure 19-41). Mitotic microtubules that are nucleated from the newly replicated centrosomes are more numerous, shorter, and less stable than interphase microtubules. The average lifetime of a microtubule decreases from 10 minutes in interphase cells to 30 seconds in the mitotic spindle. This increase in the turnover enables microtubules to assemble and disassemble more quickly during mitosis.

Figure 19-41. Microtubule dynamics during mitosis.

Figure 19-41

Microtubule dynamics during mitosis. In both interphase and mitotic cells, most microtubules radiate from the microtubule-organizing centers (MTOCs), with the (−) ends of the microtubules closest to the MTOC and the (+) ends extending outward. (more...)

As we will see in the following sections, however, microtubule motor proteins, as well as dynamic microtubules, participate in the reorganization of the spindle microtubules, separation of centrosomes, capture and alignment of chromosomes, and subsequent movement of chromosomes poleward. We have already discussed how some microtubule-dependent movements in the cytosol are generated by microtubule dynamics or by microtubule motor proteins. The involvement of both mechanisms in mitosis perhaps ensures the fail-safe apportionment of chromosomes equally to each daughter cell.

Organization of the Spindle Poles Orients the Assembly of the Mitotic Apparatus

Genetic and cell biology studies, primarily in yeast and flies, have implicated several kinesin-related proteins (KRPs), or spindle kinesins, in the separation and migration of centrosomes, thereby orienting assembly of the spindle and spindle asters. For instance, antibodies against either a (+) or (−) end – directed KRP will inhibit the formation of a bipolar spindle when they are microinjected into a cell before but not after prophase. Such experiments suggest that a (−) end – directed motor protein in the nascent overlap region of the spindle aligns the oppositely oriented microtubules extending from each centrosome, and then a (+) end – directed motor protein pushes the centrosomes farther apart as the polar microtubules elongate. Cytosolic dynein, which is localized by antibodies to the centrosome and cortex of animal cells, also probably is involved in centrosome movement and spindle orientation. Genetic studies in yeast suggest that dynein at the cortex simultaneously helps tether the astral microtubules and orient one pole of the spindle. The combination of microtubule growth, polar microtubule cross-linking, and astral microtubule – cortex interactions provides a reasonable explanation of how the spindle poles separate and orient during prophase, as depicted in Figure 19-42.

Figure 19-42. Model for participation of microtubule motor proteins in centrosome movements during mitosis.

Figure 19-42

Model for participation of microtubule motor proteins in centrosome movements during mitosis. (a) During late prophase centrosomes are aligned by (−) end – directed motors (dark green), which pull on polar microtubules (more...)

Formation of Poles and Capture of Chromosomes Are Key Events in Spindle Assembly

Assembly of the metaphase spindle requires two types of events: attachment of spindle microtubules to the poles and capture of chromosomes by kinetochore microtubules. Experiments with Xenopus egg extracts suggest that cytosolic dynein may play a critical role in organizing spindle microtubules into a pole. In the presence of sperm nuclei, centrosomes, and microtubules, bipolar spindles form in egg extracts. However, addition of antibodies against cytosolic dynein releases and splays the spindle microtubules but leaves the centrosomal astral microtubules intact (Figure 19-43a). The same results are obtained when antibodies to dynein, dynactin, or NuMA are microinjected into a cell. According to a recently proposed model, cytosolic dynein and NuMA cross-link the (−) ends of the spindle microtubules forming a spindle pole, which is linked to the centrosome through other interactions perhaps involving KRPs (Figure 19-43b). These findings also suggest that formation of the spindle pole is not tightly linked with assembly of the aster. Consistent with this is the fact that cells lacking a centrosome (e.g., plant cells) form a spindle pole even though they lack astral microtubules.

Figure 19-43. Spindle pole formation.

Figure 19-43

Spindle pole formation. (a) Immunofluorescence micrographs showing in vitro reconstituted spindles stained with fluorescent-labeled antibodies to tubulin (green) and dynein (red). The control spindle (left) has cytosolic dynein at each tapered end, as (more...)

At the opposite end of the spindle microtubules, rapid fluctuation in their length is used to capture chromosomes during prophase as the nuclear membrane begins to break down. By quickly lengthening and shortening at its (+) end, a dynamic microtubule acts like a poker that can probe into a chromosome-rich environment (Figure 19-44). Sometimes the end of a microtubule contacts a kinetochore, scoring a “bull’s-eye.” More commonly, a kinetochore contacts the side of a microtubule and then slides along the microtubule to the (+) end in a process that may involve kinesins on the kinetochore. Whether a chromosome attaches to the (+) end of a spindle microtubule by a direct hit or by the side capture/sliding process, the kinetochore “caps” the (+) end of the microtubule. Eventually, each sister chromatid in a chromosome is captured by microtubules arising from the nearest spindle poles. Each kinetochore also becomes attached to additional microtubules as mitosis progresses toward metaphase.

Figure 19-44. Dynamic instability and the capture of chromosomes.

Figure 19-44

Dynamic instability and the capture of chromosomes. (a) During mitotic prophase, some spindle microtubules are growing at their distal (+) end, while others are shrinking rapidly. (b) In late prophase, the ends of some microtubules interact with kinetochores (more...)

Kinetochores Generate the Force for Poleward Chromosome Movement

During late prophase (prometaphase), the newly condensed chromosomes attached to their kinetochore microtubules congress, or move, to the equator of the spindle (see Figure 19-34). Along the way, the chromosomes exhibit saltatory behavior, oscillating between movements toward and then away from the pole or equator. Until the kinetochore microtubules are attached to all kinetochores, the cell cycle is held in check. A single unattached kinetochore is sufficient to prevent entry into anaphase. A combination of microtubule motor proteins at the kinetochore and microtubule dynamics at the (+) end of kinetochore microtubules is thought to position the chromosomes equally between the two spindle poles. Figure 19-45 depicts several mechanisms that could position and hold chromosomes at the equatorial plate.

Figure 19-45. Proposed alternative mechanisms for chromosome congression.

Figure 19-45

Proposed alternative mechanisms for chromosome congression. A coupling of microtubule dynamics and microtubule motors may keep a chromosome positioned at the equator. (a) The flow of tubulin subunits through kinetochore microtubules can be used to pull (more...)

Experimental micromanipulation of chromosomes provides the best evidence of a force that pulls the two kinetochores on sister chromatids toward opposite poles. These studies suggest that the strength of the force is proportional to the distance from the chromosome to the pole. Thus, if a metaphase chromosome is displaced toward one pole by micromanipulation, then the force exerted from the opposite pole momentarily increases and quickly pulls the displaced chromosome back to the equator. An alternative explanation is that the pole closest to the chromosome exerts a pushing force that restores the chromosome to the equator. Whatever mechanism is in operation, these opposing forces are balanced when a chromosome is at the equator of the spindle, so the chromosome remains stationary there.

By metaphase, then, the chromosomes are aligned at the equator, their position fixed midway between each pole of the spindle. Although the lengths of kinetochore and polar microtubules have stabilized, there continues to be a flow, or treadmilling, of subunits through the microtubules toward the poles, but the loss of subunits at the (−) end is balanced by the addition of subunits at the (+) end.

During Anaphase Chromosomes Separate and the Spindle Elongates

The same forces that form the spindle during prophase and metaphase also direct the separation of chromosomes toward opposite poles at anaphase. Anaphase is divided into two distinct stages, anaphase A and anaphase B (or early and late anaphase). Anaphase A is characterized by the shortening of kinetochore microtubules, which pulls the chromosomes toward the poles. During anaphase B, the two poles move farther apart, bringing the chromosomes with them into what will become the two daughter cells.

In part because methods for studying anaphase A and anaphase B in certain cell-free extracts have been developed, we understand a good deal about their molecular mechanisms. Similar mechanisms may operate during other mitotic phases.

Microtubule Shortening during Anaphase A

In vitro studies have indicated that the depolymerization of microtubules can generate sufficient force to move chromosomes. In one such study, purified microtubules were mixed with purified anaphase chromosomes, and as expected, the kinetochores bound preferentially to the (+) ends of the microtubules. To induce depolymerization of the microtubules, the reaction mixture was diluted, thus lowering the concentration of free tubulin dimers. Video microscopy analysis then showed that the chromosome moved toward the (−) end, at a rate similar to that of chromosome movement in intact cells. Since no ATP (or any other energy source) was present in these experiments, chromosome movement toward the (−) end must be powered, in some way, by microtubule disassembly and must not be powered by microtubule motor proteins.

The in vivo fluorescence tagging experiment depicted in Figure 19-46 provides additional evidence that disassembly of kinetochore microtubules at their (+) ends coincides with the poleward movement of chromosomes. The results of these two experiments suggest that the proteins tethering the kinetochore to the microtubule progressively interact with increasingly distal portions of the microtubule as the (+) end disassembles. In this way, the kinetochores move poleward by a passive process that does not require a motor protein.

Figure 19-46. Experimental demonstration that during anaphase A chromosomes move poleward along stationary kinetochore microtubules, which coordinately disassemble from their kinetochore ends.

Figure 19-46

Experimental demonstration that during anaphase A chromosomes move poleward along stationary kinetochore microtubules, which coordinately disassemble from their kinetochore ends. Fibroblasts are injected with fluorescent tubulin and then allowed to enter (more...)

Spindle Elongation during Anaphase B

In the second stage of anaphase, polar microtubules slide past one another and elongate, and pulling forces are exerted by the cellular cortex on astral microtubules. Detergent treatment of mitotic cells, which causes ATP to leak out, does not affect poleward chromosome movement (anaphase A), but prevents the separation of spindle poles that occurs in anaphase B. Thus microtubule motor proteins clearly are involved in separating the spindle poles, as they are in centrosome movement during prometaphase (see Figure 19-42).

Anaphase A and B movements can be reconstituted in vitro by activating artificial spindles that assemble from frog egg extracts. In the presence of calcium, the spindles elongate, simulating anaphase B, and the zone of overlap between the two half-spindles decreases in length by a distance similar to the original length of the overlap. If we were to analyze the direction of microtubule movement during anaphase B, we would find that adjacent microtubules migrate in the direction of their pole-facing (−) ends. This polarity of movement suggests that a (+) end – directed KRP is responsible for generating the force for spindle pole separation during anaphase B. In one model, a KRP attached to a microtubule in the overlap region walks toward the (+) end of a neighboring but antiparallel microtubule, thus pushing the adjacent microtubule in the direction of its (−) end (Figure 19-47). This model is supported by experiments in which antibodies raised against a conserved region of the kinesin superfamily inhibit ATP-induced elongation of diatom spindles in vitro. The involvement of a kinesin-like protein and the requirement for ATP hydrolysis are two strong pieces of evidence that a KRP could be responsible for anaphase B movements.

Figure 19-47. Model of spindle elongation and movement during anaphase B. Tubulin (light purple) adds to the (+) ends of all polar microtubules, lengthening these fibers.

Figure 19-47

Model of spindle elongation and movement during anaphase B. Tubulin (light purple) adds to the (+) ends of all polar microtubules, lengthening these fibers. Simultaneously, (+) end – directed KRPs (pink) bind to the polar microtubules (more...)

In addition to the sliding forces between polar microtubules, spindle elongation involves microtubule dynamics and interactions at the cortex. In the presence of αβ-tubulin, reactivated frog spindles and isolated diatom spindles add tubulin subunits to the (+) end of polar microtubules (see Figure 19-47). A third component of anaphase B is the interaction between astral microtubules and the cortex, which generates a pulling force on the asters. This force can be demonstrated by cutting the spindle in half with a microneedle during anaphase; the resulting half-spindles move quickly to the poles, at a rate faster than usual during anaphase. This observation suggests that a (−) end – directed motor associated with the cortex, maybe cytosolic dynein or a KRP, pulls the asters farther apart toward the poles of the daughter cells (see Figure 19-47).

Astral Microtubules Determine Where Cytokinesis Takes Place

Once the chromosomes have been separated to the poles of the cell during anaphase, the nuclear envelope re-forms around each complete set of chromosomes during telophase and the cytoplasm divides (cytokinesis) (see Figure 19-34). With completion of cytokinesis, the last event in mitosis, the life of a daughter cell begins. In Chapter 18, we discussed how the contractile ring of actin and myosin constricts the cell during cytokinesis, but not the mechanism that determines the plane of cleavage through the cell (see Figure 18-37). It is clear that the contractile ring, and hence the cleavage furrow, always develops where the chromosomes lined up during metaphase, but it is not obvious which component of the mitotic apparatus dictates where the contractile ring will assemble.

Ingenious micromanipulation experiments with dividing sea urchin embryos have shown that the presence of two asters, not the spindle itself, is necessary to determine the cleavage plane. In one key experiment, illustrated in Figure 19-48, a hole is poked into a one-cell-stage sea urchin embryo before the mitotic apparatus has started to assemble, transforming it into a doughnut-shaped cell. During the first division, a spindle assembles on one side, and cytokinesis between the asters forms a single, binucleated, C-shaped cell. The two nuclei of this cell undergo another round of mitosis with assembly of a pair of mitotic apparatuses. If the spindle determines the cleavage plane, then we would expect the C-shaped cell to form two cleavage furrows and divide into three cells. In fact, four cells are generated in this experiment. The interpretation of this result is that an “extra” cell is generated by cleavage in the region between asters and lacking a spindle.

Figure 19-48. Experimental demonstration that asters alone determine the cleavage plane during cytokinesis.

Figure 19-48

Experimental demonstration that asters alone determine the cleavage plane during cytokinesis. A small glass ball is pressed against a fertilized sea urchin egg until membranes from opposite sides of the cell touch and fuse. This changes the spherical (more...)

One hypothesis is that astral microtubules send a signal to the region of the cortex midway between asters. This signal activates the assembly of actin and myosin, resulting in the formation of the contractile ring followed by the development of the cleavage furrow. The signal is unidentified as yet, but an alluring candidate is cdc2 kinase. Not only is this protein kinase activated in the cell cycle (Chapter 13), but caldesmon and myosin are among its known substrates. As discussed in the previous chapter, phosphorylation of these two proteins will activate the assembly of myosin filaments and cause caldesmon to relieve the inhibition of actin-myosin interactions.

Plant Cells Reorganize Their Microtubules and Build a New Cell Wall during Mitosis

Image plant.jpgMitotic events in plants are generally similar to those in animal cells, although a rigid cell wall must be constructed during cytokinesis (Figure 19-49). Interphase cells are girdled by a bundle of cortical microtubules that influence the pattern of deposition of cellulose in the cell wall. Despite the absence of a centrosome, plant cells reorganize their microtubules into a spindle during mitosis. At cytokinesis, in place of a contractile ring, a membrane structure, the phragmoplast, is assembled from vesicles in the interzone between the daughter nuclei. The vesicles originate from the Golgi complex and are first observed during metaphase, when they extend into the mitotic apparatus and in some cases even appear to contact the kinetochores. Later in anaphase, by traveling along microtubules that radiate from each daughter nucleus, the vesicles line up near the center of the dividing cell, where they fuse to form the phragmoplast in telophase. The membranes of the vesicles become the plasma membranes of the daughter cells, and their contents form the cell plate. The vesicles also contain material for the future cell wall, such as polysaccharide precursors of cellulose and pectin.

Figure 19-49. Mitosis in a higher plant cell.

Figure 19-49

Mitosis in a higher plant cell. Immunofluorescence micrographs (top) and corresponding diagrams (bottom) showing arrangement of microtubules in a plant cell. A cortical array of microtubules girdle the cell during interphase. As the cell enters mitosis, (more...)


  •  During mitosis, the replicated chromosomes are separated and evenly partitioned to two daughter chromosomes. This portion of the cell cycle is commonly divided into four substages: prophase, metaphase, anaphase, and telophase (see Figure 19-34).
  •  Early in mitosis, the chromosomes begin to condense, the nuclear membrane breaks down, and the mitotic spindle is assembled. These events culminate in the metaphase cell in which the fully condensed chromosomes, each composed of sister chromatids connected at the centromere, are aligned along the equatorial plate.
  •  The major components of the mitotic apparatus are the astral microtubules forming the asters; the polar and kinetochore microtubules forming the football-shaped spindle; the spindle poles, and chromosomes attached to the kinetochore microtubules (see Figure 19-36).
  •  A simple set of microtubule motor proteins — BimC, CENP-E, and cytosolic dynein — are conserved in all spindles. Additional motors are present in more complex organisms.
  •  The two centrioles in the centrosome, the MTOC in most animal cells, are replicated during interphase. As mitosis begins, the centriole pairs migrate to the two poles, where they organize the astral microtubules.
  •  Interactions between asters and the cell cortex and between opposing spindle microtubules separate and align the centrosomes during prophase, establishing the bipolar orientation of the spindle (see Figure 19-42).
  •  Spindle microtubules capture the kinetochores of chromosomes and center the chromosomes at metaphase.
  •  The combined action of microtubule dynamics (polymerization and depolymerization) and microtubule motors positions and holds chromosomes at the metaphase plate and translocates chromosomes to the opposite poles at anaphase (see Figures 19-45 and 19-47).
  •  Cleavage during cytokinesis occurs between the asters and does not depend on the presence of a spindle or chromosomes. The signal for cleavage is unknown.
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