Rings, bracelets, sleeves, and chevrons: new structures of kinetochore proteins

Abstract

Electron microscopy has recently revealed striking structural orderliness in kinetochore proteins and protein complexes that associate with microtubules. In addition to their astonishing appearance and intrinsic beauty, the structures are functionally informative. The Dam1 and Ndc80 complexes bind to the microtubule lattice as rings and chevrons, respectively. These structures give insight into how the kinetochore couples to dynamic microtubules, a process crucial to the accurate segregation of chromosomes. HURP and kinesin-13 arrange tubulin into sleeves and bracelets surrounding the microtubule lattice. These structures might reflect the ability of these proteins to modulate microtubule dynamics by interacting with specialized tubulin configurations. In this review, we compare and contrast the structure of these proteins and their interactions with microtubules to illustrate how they attach to and modulate the dynamics of microtubules.

Figure 1. Microtubule Structures

A. Polymerizing microtubules. Microtubules assemble from a sheet of 13 protofilaments, which roll up to form a tubular structure. The helical pitch intrinsic to the polymerizing tubulin dimers gives rise to a noticeable seam in the microtubule that has been visualized in the electron microscope. The fundamental unit of microtubule assembly is the αβ-tubulin dimer (light green box). B. Depolymerizing microtubules. Individual protofilaments peel apart during microtubule disassembly. The outwardly curved protofilaments, which form naturally during disassembly due to conformational change in the tubulin dimer triggered by the hydrolysis of bound GTP, can be stabilized by the motor domains of kinesin-13 family members (shown in green). In the presence of ATP the disassembly of microtubules is promoted by kinesin-13s due to their ability to form a high affinity complex with curved protofilaments. C. Tubulin rings. The products of microtubule disassembly in vitro can be individual tubulin dimers or curved protofilaments of various sizes depending on the buffer constituents. These protofilament rings can be stabilized by motor domains of kinesin-13 family proteins (shown in green) in the presence of non-hydrolyzable ATP. This promotes a strong rigor association of the motor domain preferentially with curved, rather than straight, protofilaments. D. HURP-induced sheets. Sheets of tubulin that form in the presence of HURP and preformed microtubules consist of antiparallel protofilaments similar to zinc sheets with the exception that the luminal surfaces are all on the same side of the sheet. It is this surface (the back side of the sheet shown here) that interacts with HURP bound to microtubules. E. Zinc sheets. Microtubules assembled in the presence of zinc ions will form antiparallel, interdigitating straight protofilament sheets (sometimes referred to as macrotubes). Neighboring protofilaments show opposite surfaces; a protofilament showing what would be the luminal surface in a MT (shaded dark) is neighbor to protofilaments showing what would be the outer surface in a MT (not shaded). These structures are useful for structural studies and as unusual substrates for motor activity.

Figure 2. Rings, bracelets, sleeves and chevrons formed by kinetochore proteins

A. Dam1 complex rings around microtubules. Recombinant Dam1 complex bound to paclitaxel-stabilized microtubule, Images are electron micrographs of negative stained material. Bar represents 50 nm. Reprinted from (8) with permission from Nature. B. MCAK stabilized tubulin bracelets. Paclitaxel stabilized microtubule in the presence of recombinant MCAK (core motor domain plus 64 residues of the neck domain) and AMPPNP. Image is an electron micrograph of negative stained material. Bar represents 50 nm. Reprinted from (5) with permission from Landes. C. HURP-induced tubulin sheets. Recombinant HURP was incubated with tubulin in the presence of GTP. Vitrification by quick-freezing was followed by freeze-drying and then unidirectional metal shadowing before cryo elecron microscopy. The red box indicates a region of the microtubule covered with a sleeve. Bar represents 50 nm. Reprinted from (9) with permission from Elsevier. D. Ndc80 chevrons bound to microtubules. Recombinant Ndc80 complex bound to microtubules stabilized with GMPCPP. Image is an electron micrograph of negative stained material. Bar represents 50 nm. Reprinted from (10) with permission from Elsevier.