Organization Of A Bipolar Spindle
In oocytes, the spindle lacks centrosomes (see 1.2.1.). Therefore, the bipolar organization of the spindle in oocytes, as in acentrosomal mitotic systems, is not pre-determined but becomes established as the spindle microtubules assemble around chromosomes (see 1.1.2.2.; Karsenti and Vernos, 2001). Originally randomly oriented microtubules undergo re-arrangement according to their polarity (Heald et al., 1996; Walczak et al., 1998). Microtubules are bundled into parallel and antiparallel arrays and, ultimately, become focused at the poles with their minus ends (Walczak et al., 1998). This self-organization of microtubules is mediated by microtubule-associated and motor proteins (see 1.1.1.2.; Walczak et al., 1998; Matthies et al., 1996).
A kinesin-5 family member, Eg5/Klp61F, has been shown to sort microtubules by sending microtubule minus ends away from the chromosomes in Xenopus egg extract (Walczak et al., 1998). Eg5/Klp61F has been proposed to promote formation of antiparallel microtubules within the spindle by microtubule crosslinking activity (Walczak et al., 1998; Kashina et al., 1996). Eg5/Klp61F is a homotetrameric motor, which shares its motor dimers between microtubules of opposite polarity (Sawin et al., 1992; Kashina et al., 1996). Its plus end-directed activity has been shown to be used to slide the antiparallel microtubules in vitro (Kapitein et al., 2005).
In Drosophila oocytes, nucleated microtubules are proposed to first arrange into a structure around chromosomes composed of an antiparallel microtubule array (Matthies et al., 1996; Skold et al., 2005; Jang et al., 2005). The structure is called the metaphase meiotic central spindle, MMCS. MMCS has been suggested to define the axis of the bipolar spindle organization and it is important for organization and/or maintenance of the spindle (Jang et al., 2005). Formation of the central spindle depends on a kinesin-6/MKLP1 family member, Subito (Giunta et al. 2002; Jang et al., 2005). Subito localizes to the central spindle where it has been proposed to crosslink the antiparallel microtubules (Jang et al., 2005; Jang et al., 2007). A subito mutation results in oocytes with frayed spindles and with undefined poles (Giunta et al., 2002, Jang et al., 2005). Requirement of Subito for the spindle bipolarity seems specific to meiosis, as dysfunction of Subito/MKLP1 in mitosis does not result in the loss of bipolarity (Cesario et al, 2006). Central spindle is also a site of Incenp and Aurora B recruitment in Drosophila oocytes, which is dependent on Subito (see 1.1.2.2. and 1.1.3.5.; Jang et al., 2005). Incenp maintains spindle stability by preventing formation of ectopic poles (see 1.3.1. and 1.3.2.1.; Colombie et al., 2008).
Further sorting and pole focusing of spindle microtubules was shown to be mediated by minus-end directed motors, such as dynein and a minus-end directed kinesin-14 family member, Nonclaret disjunctional, Ncd (Walczak et al., 1998; Hatsumi and Endow, 1992; Endow et al., 1994; Matthies et al., 1996). Ncd localizes uniformly to spindle microtubules and is crucial for organization of spindle poles in Drosophila oocytes (Hatsumi and Endow, 1992; Matthies et al., 1996). Ncd homolog, XCTK2, also sorts microtubules in Xenopus egg extract (Walczak et al., 1998). Similarly to dysfunction of Ncd, loss of dynein function in Xenopus egg extract results in asing spindle poles (Heald et al., 1996; Heald et al., 1997; Walczak et al., 1998). Ncd/XCTK2 and dynein have been proposed to act by cross-linking microtubules and translocating them toward minus ends, thus focusing the poles (see 1.1.1.2.; Hatsumi and Endow, 1992; Matthies et al., 1996; Walczak et al., 1998).
The integrity of the poles has been proposed to be stabilized by two non-motor proteins, dTACC and Msps in Drosophila oocytes (Cullen and Ohkura, 2001). Both localize to the spindle poles, but Msps localization depends on dTACC and Ncd. Nevertheless, the mode of dTACC and Msps function is not well understood.
Little is known about the cooperation and regulation of the microtubule associated and motor proteins in sorting spindle microtubules independently of centrosomes. Differences in initial conditions, such as presence or absence of MTOCs, may contribute to variability in spindle organization and the simpulan shape of the spindle between different systems (see 1.2.1.).
Interestingly, a membrane-associated protein, Axs, has been shown to influence the spindle size and shape in Drosophila oocytes (Kramer and Hawley, 2003). Mutation of Axs results in shorter spindles than in wild type. The spindles adopt a barrel shaped form in place of spindles with sharply tapered poles. Axs seems to have affinity for microtubules and it has been shown to encapsulate the female meiotic spindle in a manner dependent on microtubules
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