Structure-function insights into the yeast Dam1 kinetochore complex

Abstract

Faithful segregation of genetic material during cell division requires the dynamic but robust attachment of chromosomes to spindle microtubules during all stages of mitosis. This regulated attachment occurs at kinetochores, which are complex protein organelles that are essential for cell survival and genome integrity. In budding yeast, in which a single microtubule attaches per kinetochore, a heterodecamer known as the Dam1 complex (or DASH complex) is required for proper chromosome segregation. Recent years have seen a burst of structural and biophysical data concerning this interesting complex, which has caught the attention of the mitosis research field. In vitro, the Dam1 complex interacts directly with tubulin and self-assembles into ring structures around the microtubule surface. The ring is capable of tracking with depolymerizing ends, and a model has been proposed whereby the circular geometry of the oligomeric Dam1 complex allows it to couple the depolymerization of microtubules to processive chromosome movement in the absence of any additional energy source. Although it is attractive and simple, several important aspects of this model remain controversial. Additionally, the generality of the Dam1 mechanism has been questioned owing to the fact that there are no obvious Dam1 homologs beyond fungi. In this Commentary, we discuss recent structure-function studies of this intriguing complex.

Fig. 1.

The hypothetical architecture of the Dam1 complex. This model is based on the Uniprot interaction database (Ito et al., 2001) and on work by Harrison and co-workers (Miranda et al., 2007). The two dashed circles represent the subcomplexes described by Miranda et al., who also observed that Dad1p and Dad3p form a stable subcomplex (green). On the basis of the literature, Dad3p seems to be connected only to Dad1p. The interaction partner of Dad4p is unclear, although it is known that this protein is within a ternary subcomplex with Ask1p and Dad2p.

Fig. 2.

Cryo-EM showing Dam1-complex rings around microtubules. Rings that are tilted out of the plane of the image, which are harder to see, are indicated with arrows.

Fig. 3.

EM reconstruction of the negatively stained, unassembled Dam1 complex. (A) Reconstruction of the Dam1 dimer (green) and the average monomer derived from it (mesh). (B) Illustration of the conformational change that is proposed to occur upon binding of the Dam1 complex to microtubules and its oligomerization into a ring. (C) The wild-type Dam1 complex is shown as a mesh together with the difference map that is derived when the yeast Dam1p-ΔC mutant reconstruction is subtracted from it. The C-terminus of Dam1p (shown in yellow) is located near the site of microtubule interaction and oligomerization of the Dam1 complex.

Fig. 4.

Cryo-EM reconstruction of Dam1-complex double spirals (blue and yellow) around a microtubule (white) (EMDB-ID:1371). Each rotation of the spiral, which corresponds to a skewed single ring, is shown in a different color. Dashed lines separate adjacent Dam1-complex subunits along the spiral, and the solid black line denotes the two-fold axis of symmetry in the double spiral.