- Email: [email protected]
Mitosis: Disorderly Conduct at the Kinetochore
Current Biology Vol 16 No 13 R494
Dispatches
Mitosis: Disorderly Conduct at the Kinetochore Conventional models posit that microtubules bound to kinetochores act coordinately during chromosome movement. Such models need to be revised in the light of new data demonstrating uncoordinated behavior among kinetochore-associated microtubules. Duane A. Compton Chromosome alignment and segregation are performed by the microtubule-based structure called the spindle. The primary attachment site for spindle microtubules on chromosomes is the kinetochore, a discrete structure associated with the centromeric heterochromatin of each sister chromatid. Kinetochores can interact with microtubule sidewalls, but the insertion of microtubule ends into the kinetochore marks a mature interaction. Kinetochores in animal cells bind multiple microtubules (w25 in human cells), and microtubule marking experiments in the late 1980s demonstrated that chromosomes in cultured cells move poleward as their associated spindle microtubules shorten through loss of tubulin subunits from their attached kinetochore ends [1]. Those data were interpreted to suggest that microtubules bound to a kinetochore behaved in a cohesive and coordinated fashion. When chromosomes move poleward all microtubules bound to the leading kinetochore lost tubulin subunits. Surprising new data, reported recently in Current Biology [2], call that interpretation into question by showing that the conduct of microtubules attached to kinetochores is uncoordinated. Microtubules are dynamic polymers that elongate and shrink by addition and loss of tubulin subunits (ab heterodimers) from their ends. Tubulin subunits associate in a head-to-tail fashion to build protofilaments that associate laterally to form the cylindrical microtubule. When studied under controlled in vitro
conditions, disassembling microtubules display curved or curled ends as tubulin subunits peel back from the microtubule end before dissociating. Assembling microtubules display blunt ends or ends that are transiently flat sheets prior to rolling into a cylinder. Thus, morphological differences can be used to distinguish between microtubules that are assembling or disassembling [3]. It was against that backdrop that McEwen and colleagues [2] used computerized electron tomography to examine the morphology of microtubule ends bound to kinetochores during mitosis. Tomography compiles multiple images of the same object to construct a threedimensional view of the object [4]. The increased contrast that tomography provides, compared to conventional electron microscopy, made it possible to visualize microtubule end morphology in kinetochores in cells. When the microtubule ends observed with this technique in metaphase cells were categorized according to morphological criteria (blunt versus curled), the startling result emerged that roughly two-thirds of kinetochore-bound microtubules had a curled conformation at the time the cells were fixed. To rule out technical artifacts, McEwen and colleagues [2] used two different methods of fixation and observed similar distributions in microtubule end morphology. Also, they observed a predictable shift in microtubule end morphology when mitotic cells were treated with microtubule targeting drugs. Kinetochore-bound microtubule ends became predominantly curled in nocodazole-treated cells and
predominantly blunt in taxol-treated cells. Thus, assuming that the microtubule end morphology observed in kinetochores in vivo reflects the properties of microtubule ends previously documented in vitro, these data indicate that at any given time, the ends of most kinetochore microtubules are in a state of disassembly. McEwen and colleagues [2] then focused their attention on the morphology of microtubule ends within individual kinetochores. They observed only one kinetochore that was occupied by microtubules with exclusively blunt morphology and one kinetochore occupied by microtubules with exclusively curled morphology. All other kinetochores (eight) contained a mixture of microtubule end morphologies. Thus, individual kinetochores contain both assembling and disassembling microtubules, although a slight bimodal distribution in the percentage of microtubule ends with curled morphology suggests that kinetochores prefer either most or just a few microtubules disassembling at any time. Also, in a few cases, sister kinetochores could be examined and there was little coordination in the microtubule end morphology between sister kinetochores. Importantly, these trends were observed in both mammalian PtK1 cells and Drosophila S2 cells, demonstrating the generality of this phenomenon. These observations run counter to conventional models that assumed all kinetochore microtubules act coordinately during chromosome movement (Figure 1A), and indicate that kinetochore microtubules are not well coordinated (Figure 1B). To explain these observations McEwen and colleagues [2] suggest that microtubule ends can continuously switch between
Dispatch R495
states of assembly and disassembly while remaining within the confines of the kinetochore. Thus, at any given time, both assembling and disassembling microtubules are observed in kinetochores. In this scenario, kinetochores do not act deterministically on microtubule ends: instead, they influence the probability with which microtubules switch from assembly to disassembly and vice versa. For example, during poleward chromosome movement, the leading kinetochore is in a state that increases the probability that microtubules will switch into disassembly. In that situation, as was observed in anaphase kinetochores, most kinetochoreassociated microtubules are disassembling, although some assembling microtubules can transiently exist. This view fits the bimodal distribution seen in the percentages of microtubules displaying the curled end morphology, and explains the previous observation that tubulin subunits can be incorporated into kinetochore microtubules even during early anaphase [5]. A key element of this new view is that microtubule elongation and shortening can be five-fold faster than the rate of chromosome movement [6]. Thus, as chromosomes plod toward the pole, the microtubules in the leading kinetochore can rapidly switch between assembly and disassembly while remaining ensnared by the kinetochore. The preponderance of microtubules in the curled, presumably, disassembling morphology further suggests that kinetochores retard the dissociation of disassembling microtubules from the kinetochore. This point is critical for several reasons arising from the importance of microtubule dissociation from kinetochores. First, in somatic cells, chromosome segregation in anaphase is driven predominantly by the disassembly of microtubule ends bound to kinetochores; thus, kinetochores must have mechanisms to maintain tight attachment to the depolymerizing microtubule ends
A
B
Current Biology
Figure 1. Kinetochores (blue) bind multiple microtubules in animal cells. (A) Previous models proposed that, as chromosomes move (yellow arrows), all kinetochore microtubules in the leading kinetochore disassembled (curled ends) while all microtubules in the lagging kinetochore assembled (blunt ends). (B) Models based on new data show that kinetochore microtubules are not well coordinated and kinetochores contain both assembling and disassembling microtubules. These data demonstrate that kinetochores represent groups of independently acting microtubule binding sites despite their cohesive appearance.
to prevent detachment from the spindle that would result in chromosome mis-segregation. Second, it explains why kinetochore microtubules are stable to external perturbations (cold, calcium, and so on) and exhibit significantly slower tubulin subunit turnover kinetics relative to non-kinetochore microtubules [7]. And third, microtubule dissociation from kinetochores is essential to correct errors in chromosome attachment to the spindle, but a careful balance must be struck between attachment and detachment for optimal mitotic fidelity [8]. Images of kinetochore microtubules with bent ends provide a provocative suggestion for how kinetochores may prevent disassembling microtubules from dissociating. Unquestionably, this work underscores the importance of determining how kinetochores attach to microtubules and how kinetochores retard detachment of microtubules. These data add to a growing evidence that kinetochores in
animal cells which appear as cohesive units when viewed by electron microscopy actually represent independently acting microtubule binding sites that are loosely grouped together (Figure 1B). This idea is not new to scientists studying the holocentric chromosomes in Caenorhabditis elegans, but has been slow to take root with investigators studying mitosis in other organisms. Nevertheless, previous data showed that mammalian kinetochores break up into apparently repetitive units when chromosomes are dispersed by hypotonic treatment [9]. Such kinetochore fragments retain microtubule binding and force generating activities, demonstrating that the repetitive units are functional [10]. In this context, one may ask if a single microtubule attachment is sufficient for chromosome movement? Recent data show that force from the disassembly of a single microtubule may be sufficient to drive chromosome movement [11], and it is well
Current Biology Vol 16 No 13 R496
documented that kinetochores in budding yeast rely on just one microtubule attachment for segregation [12]. In mammalian cells, it has been shown that initial poleward chromosome movement can occur along the sidewall of a single microtubule [13], and one kinetochore microtubule can direct chromosome alignment to the metaphase plate [14]. Thus, it seems that a single microtubule may be sufficient for chromosome movement. However, when 20–30 microtubules are attached to a mammalian kinetochore, it remains unclear how many disassembling microtubule ends are engaged in force generation at any given moment and how the microtubule binding sites are linked to one another.
2.
3.
4.
5.
6.
7.
8.
References 1. Gorbsky, G.J., Sammak, P.J., and Borisy, G.G. (1987). Chromosomes move poleward in anaphase along stationary
9.
microtubules that coordinately disassemble from their kinetochore ends. J. Cell Biol. 104, 9–18. VandenBeldt, K.J., Barnard, R.M., Hergert, P.J., Meng, X., Maiato, H., and McEwen, B.F. (2006). Kinetochores use a novel mechanism for coordinating the dynamics of individual microtubules. Curr. Biol. 16, 1217–1223. Howard, J., and Hyman, A.A. (2003). Dynamics and mechanics of the microtubule plus end. Nature 422, 753–758. McIntosh, R., Nicastro, D., and Mastronarde, D. (2005). New views of cells in 3D: an introduction to electron tomography. Trends Cell Biol. 15, 43–51. Wadsworth, P., Shelden, E., Rupp, G., and Rieder, C.L. (1989). Biotin-tubulin incorporates into kinetochore fiber microtubules during early but not late anaphase. J. Cell Biol. 109, 309–321. Rusan, N.M., Fagerstrom, C.J., Yvon, A.M., and Wadsworth, P. (2001). Cell cycle-dependent changes in microtubule dynamics in living cells expressing green fluorescent proteinalpha tubulin. Mol. Biol. Cell 12, 971–980. Zhai, Y., Kronebusch, P.J., and Borisy, G.G. (1995). Kinetochore microtubule dynamics and the metaphase-anaphase transition. J. Cell Biol. 131, 721–734. Nicklas, R.B., and Ward, S.C. (1994). Elements of error correction in mitosis: microtubule capture, release, and tension. J. Cell Biol. 126, 1241–1253. Zinkowski, R.P., Meyne, J., and Brinkley, B.R. (1991). The centromere-
Meiotic Diapause: How a Sperm Signal Sets You Free Major sperm protein, a cytoskeletal molecule required for the amoeboid motility of sperm in Caenorhabditis elegans, also functions as a signaling molecule that regulates the rates of meiotic maturation and ovulation. Recent work has begun to uncover new genes required for the response to this signal in both somatic and germ line cells. Indrani Chatterjee, Pavan Kadandale, and Andrew Singson In the model nematode Caenorhabditis elegans, both sexes — males and hermaphrodites — produce sperm [1,2]. The hermaphrodite can only produce several hundred sperm during its last larval stage; it then uses these sperm to fertilize the oocytes that it produces as an adult. This makes sperm a limited resource in the unmated hermaphrodite. Hermaphrodites therefore need to limit the number of metabolically costly oocytes that are ovulated in the absence of sperm.
The regulation of meiotic maturation by hormonal signaling is a highly conserved process [3,4]. The progression of meiotic maturation is needed to prepare the oocyte for successful fertilization. Pioneering work in C. elegans demonstrated that the presence of properly differentiated proximal gonad sheath cells and sperm are required for mitogen-activated protein kinase (MAPK) activation in oocytes, meiotic progression and high rates of ovulation [5–7]. In ‘genetic female’ mutants that lack sperm, oocytes remain arrested in diakinesis for extended periods and the rate of sheath cell contractions is also very low [6].
10.
11.
12.
13.
14.
kinetochore complex: a repeat subunit model. J. Cell Biol. 113, 1091–1110. Khodjakov, A., Cole, R.W., McEwen, B.F., Buttle, K.F., and Rieder, C.L. (1997). Chromosome fragments possessing only one kinetochore can congress to the spindle equator. J. Cell Biol. 136, 229–240. Grishchuk, E.L., Molodtsov, M.I., Ataullakhanov, F.I., and McIntosh, J.R. (2005). Force production by disassembling microtubules. Nature 438, 384–388. McAinsh, A.D., Tytell, J.D., and Sorger, P.K. (2003). Structure, function, and regulation of budding yeast kinetochores. Annu. Rev. Cell Dev. Biol. 19, 519–539. Rieder, C.L., and Alexander, S.P. (1990). Kinetochores are transported poleward along a single astral microtubule during chromosome attachment to the spindle in newt lung cells. J. Cell Biol. 110, 81–95. McEwen, B.F., Heagle, A.B., Cassels, G.O., Buttle, K.F., and Rieder, C.L. (1997). Kinetochore fiber maturation in PtK1 cells and its implications for the mechanisms of chromosome congression and anaphase onset. J. Cell Biol. 137, 1567–1580.
Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755, USA. E-mail: [email protected]
DOI: 10.1016/j.cub.2006.06.010
Further studies showed that sheath cells are in direct communication with oocytes through gap junctions [8]. A great leap forward for the field came from a landmark study by Miller et al. [9], who showed that an activity in sperm-conditioned media can stimulate oocyte maturation and ovulation. This activity turned out to be the major sperm protein (MSP), which was previously known as a cytoskeletal protein required for the amoeboid motility of nematode sperm [10]. Further, it was shown using recombinant molecules that MSP provides two distinct signaling activities: an amino-terminal activity promotes oocyte meiotic maturation, while a carboxy-terminal activity stimulates the rate of sheath cell contractions. The question of how MSP gets out of sperm to mediate its signaling function was answered in a recent study [11] which showed that MSP is released from sperm via a novel double-membrane vesicle budding mechanism.
Dispatches
Mitosis: Disorderly Conduct at the Kinetochore Conventional models posit that microtubules bound to kinetochores act coordinately during chromosome movement. Such models need to be revised in the light of new data demonstrating uncoordinated behavior among kinetochore-associated microtubules. Duane A. Compton Chromosome alignment and segregation are performed by the microtubule-based structure called the spindle. The primary attachment site for spindle microtubules on chromosomes is the kinetochore, a discrete structure associated with the centromeric heterochromatin of each sister chromatid. Kinetochores can interact with microtubule sidewalls, but the insertion of microtubule ends into the kinetochore marks a mature interaction. Kinetochores in animal cells bind multiple microtubules (w25 in human cells), and microtubule marking experiments in the late 1980s demonstrated that chromosomes in cultured cells move poleward as their associated spindle microtubules shorten through loss of tubulin subunits from their attached kinetochore ends [1]. Those data were interpreted to suggest that microtubules bound to a kinetochore behaved in a cohesive and coordinated fashion. When chromosomes move poleward all microtubules bound to the leading kinetochore lost tubulin subunits. Surprising new data, reported recently in Current Biology [2], call that interpretation into question by showing that the conduct of microtubules attached to kinetochores is uncoordinated. Microtubules are dynamic polymers that elongate and shrink by addition and loss of tubulin subunits (ab heterodimers) from their ends. Tubulin subunits associate in a head-to-tail fashion to build protofilaments that associate laterally to form the cylindrical microtubule. When studied under controlled in vitro
conditions, disassembling microtubules display curved or curled ends as tubulin subunits peel back from the microtubule end before dissociating. Assembling microtubules display blunt ends or ends that are transiently flat sheets prior to rolling into a cylinder. Thus, morphological differences can be used to distinguish between microtubules that are assembling or disassembling [3]. It was against that backdrop that McEwen and colleagues [2] used computerized electron tomography to examine the morphology of microtubule ends bound to kinetochores during mitosis. Tomography compiles multiple images of the same object to construct a threedimensional view of the object [4]. The increased contrast that tomography provides, compared to conventional electron microscopy, made it possible to visualize microtubule end morphology in kinetochores in cells. When the microtubule ends observed with this technique in metaphase cells were categorized according to morphological criteria (blunt versus curled), the startling result emerged that roughly two-thirds of kinetochore-bound microtubules had a curled conformation at the time the cells were fixed. To rule out technical artifacts, McEwen and colleagues [2] used two different methods of fixation and observed similar distributions in microtubule end morphology. Also, they observed a predictable shift in microtubule end morphology when mitotic cells were treated with microtubule targeting drugs. Kinetochore-bound microtubule ends became predominantly curled in nocodazole-treated cells and
predominantly blunt in taxol-treated cells. Thus, assuming that the microtubule end morphology observed in kinetochores in vivo reflects the properties of microtubule ends previously documented in vitro, these data indicate that at any given time, the ends of most kinetochore microtubules are in a state of disassembly. McEwen and colleagues [2] then focused their attention on the morphology of microtubule ends within individual kinetochores. They observed only one kinetochore that was occupied by microtubules with exclusively blunt morphology and one kinetochore occupied by microtubules with exclusively curled morphology. All other kinetochores (eight) contained a mixture of microtubule end morphologies. Thus, individual kinetochores contain both assembling and disassembling microtubules, although a slight bimodal distribution in the percentage of microtubule ends with curled morphology suggests that kinetochores prefer either most or just a few microtubules disassembling at any time. Also, in a few cases, sister kinetochores could be examined and there was little coordination in the microtubule end morphology between sister kinetochores. Importantly, these trends were observed in both mammalian PtK1 cells and Drosophila S2 cells, demonstrating the generality of this phenomenon. These observations run counter to conventional models that assumed all kinetochore microtubules act coordinately during chromosome movement (Figure 1A), and indicate that kinetochore microtubules are not well coordinated (Figure 1B). To explain these observations McEwen and colleagues [2] suggest that microtubule ends can continuously switch between
Dispatch R495
states of assembly and disassembly while remaining within the confines of the kinetochore. Thus, at any given time, both assembling and disassembling microtubules are observed in kinetochores. In this scenario, kinetochores do not act deterministically on microtubule ends: instead, they influence the probability with which microtubules switch from assembly to disassembly and vice versa. For example, during poleward chromosome movement, the leading kinetochore is in a state that increases the probability that microtubules will switch into disassembly. In that situation, as was observed in anaphase kinetochores, most kinetochoreassociated microtubules are disassembling, although some assembling microtubules can transiently exist. This view fits the bimodal distribution seen in the percentages of microtubules displaying the curled end morphology, and explains the previous observation that tubulin subunits can be incorporated into kinetochore microtubules even during early anaphase [5]. A key element of this new view is that microtubule elongation and shortening can be five-fold faster than the rate of chromosome movement [6]. Thus, as chromosomes plod toward the pole, the microtubules in the leading kinetochore can rapidly switch between assembly and disassembly while remaining ensnared by the kinetochore. The preponderance of microtubules in the curled, presumably, disassembling morphology further suggests that kinetochores retard the dissociation of disassembling microtubules from the kinetochore. This point is critical for several reasons arising from the importance of microtubule dissociation from kinetochores. First, in somatic cells, chromosome segregation in anaphase is driven predominantly by the disassembly of microtubule ends bound to kinetochores; thus, kinetochores must have mechanisms to maintain tight attachment to the depolymerizing microtubule ends
A
B
Current Biology
Figure 1. Kinetochores (blue) bind multiple microtubules in animal cells. (A) Previous models proposed that, as chromosomes move (yellow arrows), all kinetochore microtubules in the leading kinetochore disassembled (curled ends) while all microtubules in the lagging kinetochore assembled (blunt ends). (B) Models based on new data show that kinetochore microtubules are not well coordinated and kinetochores contain both assembling and disassembling microtubules. These data demonstrate that kinetochores represent groups of independently acting microtubule binding sites despite their cohesive appearance.
to prevent detachment from the spindle that would result in chromosome mis-segregation. Second, it explains why kinetochore microtubules are stable to external perturbations (cold, calcium, and so on) and exhibit significantly slower tubulin subunit turnover kinetics relative to non-kinetochore microtubules [7]. And third, microtubule dissociation from kinetochores is essential to correct errors in chromosome attachment to the spindle, but a careful balance must be struck between attachment and detachment for optimal mitotic fidelity [8]. Images of kinetochore microtubules with bent ends provide a provocative suggestion for how kinetochores may prevent disassembling microtubules from dissociating. Unquestionably, this work underscores the importance of determining how kinetochores attach to microtubules and how kinetochores retard detachment of microtubules. These data add to a growing evidence that kinetochores in
animal cells which appear as cohesive units when viewed by electron microscopy actually represent independently acting microtubule binding sites that are loosely grouped together (Figure 1B). This idea is not new to scientists studying the holocentric chromosomes in Caenorhabditis elegans, but has been slow to take root with investigators studying mitosis in other organisms. Nevertheless, previous data showed that mammalian kinetochores break up into apparently repetitive units when chromosomes are dispersed by hypotonic treatment [9]. Such kinetochore fragments retain microtubule binding and force generating activities, demonstrating that the repetitive units are functional [10]. In this context, one may ask if a single microtubule attachment is sufficient for chromosome movement? Recent data show that force from the disassembly of a single microtubule may be sufficient to drive chromosome movement [11], and it is well
Current Biology Vol 16 No 13 R496
documented that kinetochores in budding yeast rely on just one microtubule attachment for segregation [12]. In mammalian cells, it has been shown that initial poleward chromosome movement can occur along the sidewall of a single microtubule [13], and one kinetochore microtubule can direct chromosome alignment to the metaphase plate [14]. Thus, it seems that a single microtubule may be sufficient for chromosome movement. However, when 20–30 microtubules are attached to a mammalian kinetochore, it remains unclear how many disassembling microtubule ends are engaged in force generation at any given moment and how the microtubule binding sites are linked to one another.
2.
3.
4.
5.
6.
7.
8.
References 1. Gorbsky, G.J., Sammak, P.J., and Borisy, G.G. (1987). Chromosomes move poleward in anaphase along stationary
9.
microtubules that coordinately disassemble from their kinetochore ends. J. Cell Biol. 104, 9–18. VandenBeldt, K.J., Barnard, R.M., Hergert, P.J., Meng, X., Maiato, H., and McEwen, B.F. (2006). Kinetochores use a novel mechanism for coordinating the dynamics of individual microtubules. Curr. Biol. 16, 1217–1223. Howard, J., and Hyman, A.A. (2003). Dynamics and mechanics of the microtubule plus end. Nature 422, 753–758. McIntosh, R., Nicastro, D., and Mastronarde, D. (2005). New views of cells in 3D: an introduction to electron tomography. Trends Cell Biol. 15, 43–51. Wadsworth, P., Shelden, E., Rupp, G., and Rieder, C.L. (1989). Biotin-tubulin incorporates into kinetochore fiber microtubules during early but not late anaphase. J. Cell Biol. 109, 309–321. Rusan, N.M., Fagerstrom, C.J., Yvon, A.M., and Wadsworth, P. (2001). Cell cycle-dependent changes in microtubule dynamics in living cells expressing green fluorescent proteinalpha tubulin. Mol. Biol. Cell 12, 971–980. Zhai, Y., Kronebusch, P.J., and Borisy, G.G. (1995). Kinetochore microtubule dynamics and the metaphase-anaphase transition. J. Cell Biol. 131, 721–734. Nicklas, R.B., and Ward, S.C. (1994). Elements of error correction in mitosis: microtubule capture, release, and tension. J. Cell Biol. 126, 1241–1253. Zinkowski, R.P., Meyne, J., and Brinkley, B.R. (1991). The centromere-
Meiotic Diapause: How a Sperm Signal Sets You Free Major sperm protein, a cytoskeletal molecule required for the amoeboid motility of sperm in Caenorhabditis elegans, also functions as a signaling molecule that regulates the rates of meiotic maturation and ovulation. Recent work has begun to uncover new genes required for the response to this signal in both somatic and germ line cells. Indrani Chatterjee, Pavan Kadandale, and Andrew Singson In the model nematode Caenorhabditis elegans, both sexes — males and hermaphrodites — produce sperm [1,2]. The hermaphrodite can only produce several hundred sperm during its last larval stage; it then uses these sperm to fertilize the oocytes that it produces as an adult. This makes sperm a limited resource in the unmated hermaphrodite. Hermaphrodites therefore need to limit the number of metabolically costly oocytes that are ovulated in the absence of sperm.
The regulation of meiotic maturation by hormonal signaling is a highly conserved process [3,4]. The progression of meiotic maturation is needed to prepare the oocyte for successful fertilization. Pioneering work in C. elegans demonstrated that the presence of properly differentiated proximal gonad sheath cells and sperm are required for mitogen-activated protein kinase (MAPK) activation in oocytes, meiotic progression and high rates of ovulation [5–7]. In ‘genetic female’ mutants that lack sperm, oocytes remain arrested in diakinesis for extended periods and the rate of sheath cell contractions is also very low [6].
10.
11.
12.
13.
14.
kinetochore complex: a repeat subunit model. J. Cell Biol. 113, 1091–1110. Khodjakov, A., Cole, R.W., McEwen, B.F., Buttle, K.F., and Rieder, C.L. (1997). Chromosome fragments possessing only one kinetochore can congress to the spindle equator. J. Cell Biol. 136, 229–240. Grishchuk, E.L., Molodtsov, M.I., Ataullakhanov, F.I., and McIntosh, J.R. (2005). Force production by disassembling microtubules. Nature 438, 384–388. McAinsh, A.D., Tytell, J.D., and Sorger, P.K. (2003). Structure, function, and regulation of budding yeast kinetochores. Annu. Rev. Cell Dev. Biol. 19, 519–539. Rieder, C.L., and Alexander, S.P. (1990). Kinetochores are transported poleward along a single astral microtubule during chromosome attachment to the spindle in newt lung cells. J. Cell Biol. 110, 81–95. McEwen, B.F., Heagle, A.B., Cassels, G.O., Buttle, K.F., and Rieder, C.L. (1997). Kinetochore fiber maturation in PtK1 cells and its implications for the mechanisms of chromosome congression and anaphase onset. J. Cell Biol. 137, 1567–1580.
Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755, USA. E-mail: [email protected]
DOI: 10.1016/j.cub.2006.06.010
Further studies showed that sheath cells are in direct communication with oocytes through gap junctions [8]. A great leap forward for the field came from a landmark study by Miller et al. [9], who showed that an activity in sperm-conditioned media can stimulate oocyte maturation and ovulation. This activity turned out to be the major sperm protein (MSP), which was previously known as a cytoskeletal protein required for the amoeboid motility of nematode sperm [10]. Further, it was shown using recombinant molecules that MSP provides two distinct signaling activities: an amino-terminal activity promotes oocyte meiotic maturation, while a carboxy-terminal activity stimulates the rate of sheath cell contractions. The question of how MSP gets out of sperm to mediate its signaling function was answered in a recent study [11] which showed that MSP is released from sperm via a novel double-membrane vesicle budding mechanism.