A 3D map of the yeast kinetochore reveals the presence of core and accessory centromere-specific histone.
The budding yeast kinetochore is ~68 nm in length with a diameter slightly larger than a 25 nm microtubule. The kinetochores from the 16 chromosomes are organized in a stereotypic cluster encircling central spindle microtubules. Quantitative analysis of the inner kinetochore cluster (Cse4, COMA) reveals structural features not apparent in singly attached kinetochores. The cluster of Cse4-containing kinetochores is physically larger perpendicular to the spindle axis relative to the cluster of Ndc80 molecules. If there was a single Cse4 (molecule or nucleosome) at the kinetochore attached to each microtubule plus end, the cluster of Cse4 would appear geometrically identical to Ndc80. Thus, the structure of the inner kinetochore at the surface of the chromosomes remains unsolved. We have used point fluorescence microscopy and statistical probability maps to deduce the two-dimensional mean position of representative components of the yeast kinetochore relative to the mitotic spindle in metaphase. Comparison of the experimental images to three-dimensional architectures from convolution of mathematical models reveals a pool of Cse4 radially displaced from Cse4 at the kinetochore and kinetochore microtubule plus ends. The pool of displaced Cse4 can be experimentally depleted in mRNA processing pat1Δ or xrn1Δ mutants. The peripheral Cse4 molecules do not template outer kinetochore components. This study suggests an inner kinetochore plate at the centromere-microtubule interface in budding yeast and yields information on the number of Ndc80 molecules at the microtubule attachment site.
Department of Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-3280, USA.
Julian
J
Haase
Prashant K
PK
Mishra
Andrew
A
Stephens
Rachel
R
Haggerty
Cory
C
Quammen
Russell M
RM
Taylor
Elaine
E
Yeh
Munira A
MA
Basrai
Kerry
K
Bloom
D002503:Centromere / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005095:Exoribonucleases / Q000235:genetics; D006657:Histones / Q000378:metabolism*; D018386:Kinetochores / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000235:genetics; D008870:Microtubules; D009687:Nuclear Proteins / Q000378:metabolism*; D020836:Protein Structure, Quaternary; D016601:RNA-Binding Proteins / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D008941:Spindle Apparatus / Q000378:metabolism
C092746:CSE4 protein, S cerevisiae; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; D006657:Histones; D008869:Microtubule-Associated Proteins; C112472:NDC80 protein, S cerevisiae; D009687:Nuclear Proteins; C103266:PAT1 protein, S cerevisiae; D016601:RNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; D005095:Exoribonucleases; C068398:XRN1 protein, S cerevisiae