Cell：“分子发动机和刹车”作用机制的研究

Bias in Crosslinking Polarity Aids Protein Localization and MT Self-Organization

We found that the MT bundling protein ase1p has the inherent ability to distinguish between parallel and antiparallel MTs. A similar specificity had been proposed for the binding of α-actinin isoforms to actin filaments (Meyer et al., 1990), but the potential of polarity specificity of cytoskeletal bundling proteins to control self-organization processes in cells has remained largely unaddressed. Because of its polarity preference and its weak binding to single MTs, ase1p localizes with high fidelity to antiparallel interdigitated MTs. Interestingly, ase1p localizes to overlapping minus ends during interphase and overlapping plus ends in anaphase spindles. The difference in localization in both cell stages may be purely determined by molecular motors and geometry effects that bring either antiparallel minus ends (interphase/klp2p) or plus ends together (anaphase/opposing spindle poles; Figure 6). The specialized MT binding properties of ase1p/PRC1 may aid the localization of various proteins. Molecular binding partners of PRC1 in mitosis, which intriguingly include various kinesin motors (Gruneberg et al., 2006, Kurasawa et al., 2004, Zhu et al., 2005) and a RhoA GAP (Ban et al., 2004), may associate with PRC1 for ATP-independent transport to the spindle midzone. In addition, other spindle midzone components like the centralspindlin complex (Hizlan et al., 2006) may similarly use multiple antiparallel-oriented MT binding sites for their localization.

Figure 6. Model for MT Organization with Competing Sliding and Friction Forces

(A) MT plus ends are indicated by arrow heads, minus ends by spheres. MT nucleation along interphase bundles occurs from MT-bound nucleation complexes (purple). After nucleation, MTs are stabilized in the antiparallel configuration by polarity-specific ase1p (green). The minus-end-directed kinesin-14 klp2p (red) subsequently transports MTs to the bundle midzone. As the new MT grows, additional ase1p binds, increasing the friction against a length-independent number of motors at MT plus ends. Consequently, the speed of transport decreases and finally becomes zero when motors lose contact with antiparallel MTs.

(B) Possible mechanisms, based on length-dependent and -independent forces, for the regulation of overlap between antiparallel overlapping MT plus ends. The bundling of antiparallel MTs by ase1p (observed in anaphase spindles) would not interfere with the focusing of parallel MTs toward the spindle poles by minus-end-directed motors (yellow). Other minus-end-directed motors may specifically bind to MT plus ends (blue) and pull poles together with an overlap-independent force. An increase in overlap recruits additional ase1p proteins, generating friction that resists MT sliding and slows down pole-to-pole motion. Plus-end-directed motors (black) may bind in a length-dependent manner along MTs, a process potentially regulated by the binding of motors to ase1p. For large overlap, enough plus-end-directed motors may bind to push the poles outward. An equilibrium overlap can then exist at which forces generated by plus- and minus-end-directed motors and ase1p are balanced.

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Apart from aiding localization, a polarity bias in MT crosslinking also enables ase1p to polarize self-organization processes. Newly nucleated MTs along interphase MT bundles, for example, are orientated antiparallel with respect to the underlying MT (Janson et al., 2005), even in the absence of klp2p (Figure S1B). This geometry, which may in part be initiated by a preferred orientation of nucleation complexes, is stabilized specifically by ase1p (see model, Figure 6A). In computer model 2, where as

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