Precocious (pre-anaphase) cleavage furrows in Mesostoma spermatocytes
Introduction
Cell cleavage (cytokinesis) in animals cells generally begins during mid-anaphase or late-anaphase: the cleavage furrow indents the cell cortex in the plane that was occupied by the former metaphase plate of chromosomes and it continuously constricts the cytoplasm until the cell is cleaved into two cells (Burgess and Chang, 2005, Rappaport, 1996). The contractile ring that cleaves the cell generates the force using actin and myosin, and contains as well a variety of other proteins [e.g., anillin, septins, formins, Arp2/3, profilin, MLCK, etc. (Eggert et al., 2006, Guertin et al., 2002, Oegama and Hyman, 2006)], including regulatory proteins such as Rho (Bement et al., 2005, von Dassow, 2009). The work we report here deals with two large scale attributes of the cytokinetic machinery: when does the furrow begin to constrict the cell? And how is the furrow position specified?
There is general agreement in the literature that in animal cells furrowing does not begin until after chromosomes separate in anaphase (e.g., reviews in Barr and Gruneberg, 2007, Burgess and Chang, 2005), but specification of the furrow position occurs earlier (e.g., Rappaport, 1981, Rappaport and Rappaport, 1993). As summarised by Rappaport, 1996 (page 138): “The event that establishes the division [furrow position] can be relatively brief, and it normally occurs (in the small number of cell types that have actually been tested) at metaphase–anaphase.”
There is no solid consensus on how the site of the furrow is determined. Most consider that spindle microtubules interact with the cell cortex to specify furrow position (e.g., Albertson et al., 2008, Alsop and Zhang, 2003, Barr and Gruneberg, 2007, Burgess and Chang, 2005, Canman et al., 2003, Chen et al., 2008, D’Avino et al., 2005, Murthy and Wadsworth, 2008, Rappaport and Rappaport, 1993, Rappaport, 1996, Wadsworth, 2005). The spindle-cortex interaction may be generated by spindle microtubules in the spindle equator (e.g., Rappaport, 1996), or associated with the midzone of the anaphase spindle (e.g., Alsop and Zhang, 2003, Wheatley and Wang, 1996), or associated with spindle asters, e.g., transporting components to the cortex or causing relaxation at the poles and thence contraction at the equator (e.g., Oegama and Mitchison, 1997, Rappaport, 1996, Sisson et al., 1999), though recent experiments show that cleavage can take place even when astral microtubules do not extend to the cortex (von Dassow et al., 2009). Or cells may specify furrow positions in more than one way (e.g., Bringmann and Hyman, 2005, Chen et al., 2008).
The work on Mesostoma spermatocytes that we present here raises questions about both these generalisations, that is, that furrow constriction does not begin until after the start of anaphase, and that spindle microtubules specify the furrow position.
The cleavage furrow in the spermatocyte of the flatworm Mesostoma appears in prometaphase, more than one hour before the start of anaphase. Once the furrow appears, its ingression is arrested until anaphase, or continues very slowly until anaphase, after which it cleaves the cell in the usual manner. Precocious cleavage furrows also are seen in diatoms (Pickett-Heaps et al., 1980) and various other algae such as Spirogyra (e.g., McIntosh et al., 1995), in which precocious furrows may appear as early as prophase (e.g., Lokhorst et al., 1988, Mattox and Stewart, 1974, McIntosh et al., 1995, Sawitzky and Grolig, 1995). We know of no animal cells except Mesostoma spermatocytes in which a precocious furrow persists throughout division and eventually cleaves the cell after anaphase.
The furrow's position in a Mesostoma spermatocyte changes when the distribution of chromosomes changes. In early prometaphase each of the 4 univalent chromosomes moves to a pole as the bivalents become bipolarly oriented, but univalent chromosomes often move between the poles during prometaphase in order to achieve their proper segregation (Oakley, 1985). As we illustrate in this article, the prometaphase cleavage furrow is sensitive to these redistributions; its position changes when there are unequal numbers of univalents at the two poles, though the spindle remains in its one, fixed position.
Section snippets
Materials and methods
Mesostoma were reared from diapausing (overwintering) eggs originally provided by Dr. Paul Hebert, University of Guelph (e.g., Hebert and Beaton, 1990) and kept in pond water at 4 °C for several months before being placed in a jar that contained algae and Daphnia; the algae were food for the Daphnia which in turn were food for the Mesostoma that hatched from the eggs. Mesostoma are hermaphrodites, with both testes and ovaries; adults can be viviparous, or can bear diapausing eggs (Ferguson and
Results
Mesostoma spermatocytes contain three bivalent and four univalent chromosomes (Fig. 1). The three bivalents (arrows in Fig. 1) are bipolarly oriented from early prometaphase onwards; they are monochiasmatic, with prominent free arms at each end (the grey arrowheads in Fig. 1 point to kinetochores from which the free arms extend). The four univalents are at the poles from early prometaphase onwards (Oakley, 1985) and thus would seem to be unipolarly oriented. The univalents (open arrows in Fig. 1
Precocious formation of the cleavage furrow
Precocious cleavage furrows are initiated in early prometaphase Mesostoma spermatocytes, nearly two hours before anaphase; furrow ingrowth proceeds for a short time but is arrested until telophase when ingression recommences and cleaves the cell (e.g., Fig. 3, Fig. 5, Fig. 10). Initiation of furrows in prometaphase is an apparent exception to the general consensus that cleavage begins as cells transit through anaphase into telophase. From viewing cleavage in several types of entirely unrelated
Acknowledgements
This work was supported by grants from the Natural Sciences and Engineering Council of Canada to A.F. We thank Karen Rethoret, York University, for sectioning the cells and taking the electron micrographs.
References (68)
Formation of the first cleavage spindle in nematode embryos
Dev. Biol.
(1984)- et al.
Cytokinesis: placing and making the final cut
Cell
(2007) - et al.
Site selection for the cleavage furrow at cytokinesis
Trends Cell Biol.
(2005) - et al.
Mitosis-specific mechanosensing and contractile-protein redistribution control cell shape
Curr. Biol.
(2006) - et al.
Fibrin clots keep non-adhering living cells in place on glass for perfusion or fixation
Cell Biol. Int.
(2005) - et al.
Duration of division-related events in cleaving sand dollar eggs
Dev. Biol.
(1993) - et al.
Asymmetrization of first cleavage by transient disassembly of one spindle pole aster in the leech Helobdella robusta
Dev. Biol.
(2006) - et al.
Pseudocleavage is dispensible for polarity and development in C. elegans embryos
Dev. Biol.
(1995) - et al.
Cytokinesis: lessons from Rappaport and the Drosophila blastoderm embryo
Cell Biol. Int.
(1999) Concurrent cues for cytokinetic furrow induction in animal cells
Trends Cell Biol.
(2009)
Astral signals spatially bias cortical myosin recruitment to break symmetry and promote cytokinesis
Curr. Biol.
Vesicles and actin are targeted to the cleavage furrow via furrow microtubules and the central spindle
J. Cell Biol.
Microtubules are the only structural constituent of the spindle apparatus required for induction of cell cleavage
J. Cell Biol.
A microtubule-dependent zone of active RhoA during cleavage plane specification
J. Cell Biol.
A cytokinesis furrow is positioned by two consecutive signals
Nature
Determining the position of the cell division plane
Nature
Redundant mechanisms recruit actin into the contractile ring in silkworm spermatocytes
PLoS Biol.
Early spindle assembly in Drosophila embryos: role of a force balance involving cytoskeletal dynamics and nuclear mechanics
Mol. Biol. Cell
Cleavage furrow formation and ingression during animal cytokinesis: a microtubule legacy
J. Cell Sci.
Animal cytokinesis: from parts list to mechanisms
Annu. Rev. Biochem.
Redundant mechanisms for anaphase chromosome movements: crane-fly spermatocyte spindles normally use actin filaments but also can function without them
Protoplasma
Titin in insect spermatocyte spindle fibers associates with actin, myosin and the matrix proteins skeletor, megator and chromator
J. Cell Sci.
A synopsis of the genus Mesostoma Ehrenberg 1835
J. Elisha Mitchell Scientific Soc.
Microtubules and mitotic cycle phase modulate spatiotemporal distributions of F-actin and myosin II in Drosophila syncytial blastoderm embryos
Development
Does 2,3-butanedione monoxime inhibit nonmuscle myosin?
Protoplasma
Structure of kinetochore fibres in crane-fly spermatocytes after irradiation with an ultraviolet microbeam: neither microtubules nor actin filaments remain in the irradiated region
Cell Motil. Cytoskel.
What generates flux of tubulin in kinetochore microtubules?
Protoplasma
Oscillatory movements of bipolarly-oriented bivalent kinetochores and spindle forces in male meiosis of Mesostoma ehrenbergii
Europ. J. Cell Biol.
Rapid kinetochore movements in Mesostoma ehrenbergii spermatocytes: action of antagonistic chromosome fibres
Cell Motil. Cytoskel.
Morphological aspects of chromosome spindle fibres in Mesostoma: “microtubular fir-tree” structures and microtubule association with kinetochores and chromatin
Protoplasma
Cleavage furrow positioning
J. Cell Biol.
Polarity controls forces governing asymmetric spindle positioning in the Caenorhabditis elegans embryo
Nature
Cytokinesis in eukaryotes
Microbiol. Mol. Biol. Rev.
Breeding system and genome size of the rhabdocoel turbellarian Mesostoma ehrenbergii
Genome
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