Unless you are in a field of study related to cell biology, you most likely
have never heard of Ndc80. Yet this protein complex is essential to mitosis,
the process by which a living cell separates its chromosomes and distributes
them equally between its two daughter cells.
Now, through a combination of cryo-electron microscopy and three-dimensional
image reconstruction, a team of researchers with the Lawrence
Berkeley National Laboratory (Berkeley Lab) and the University of California
(UC) Berkeley have produced a subnanometer resolution model of human Ndc80 that
reveals how this unsung hero carries out its essential tasks.
“Our model suggests that Ndc80 oligomerizes on the surface of the microtubule
via a segment of the protein that is regulated so that correct attachments are
maintained and incorrect attachments are discarded,” says biophysicist
Eva Nogales who led this study.
“What we propose is that this oligomerization is an important part of
the mechanism by which Ndc80 is able to utilize the energy of microtubule disassembly
to move chromosomes towards the spindle poles during mitosis. This oligomerization
will only happen for correctly attached microtubules”
Nogales holds joint appointments with Berkeley Lab’s Life Sciences Divisions,
UC Berkeley’s Molecular and Cell Biology Department, and the Howard Hughes
Medical Institute. An expert on electron microscopy and image analysis and an
authority on the structure and dynamics of microtubules, she is the corresponding
author of a paper published in the journal Nature titled, “The Ndc80 kinetochore
complex forms oligomeric arrays along microtubules.”
Co-authoring the paper with Nogales were Gregory Alushin, Vincent Ramey, Sebastiano
Pasqualato, David Ball, Nikolaus Grigorieff and Andrea Musacchio.
Biological cells have a cytoskeleton that gives shape to membrane walls and
other cellular structures and also controls the transportation of substances
in and out of the cell. This cytoskeleton is spun from tiny fibers of tubulin
protein called microtubles. During mitosis, microtubules disassemble and reform
into spindles across which duplicate sets of chromosomes line up and migrate
to opposite poles. After chromosome migration is complete, the microtubules
disassemble and reform back into skeletal systems for the two new daughter cells.
Mistakes in the distribution of chromosomes from a parent cell to its daughter
cells can lead to birth defects, cancer and other disorders. To ensure that
each daughter cell receives a single copy of each chromosome, microtubule spindles
dock with each chromosome’s centromere – the central region where
its two chromatids connect. The microtubule spindles connect with the centromere
through a network of proteins called the kinetochore. Ndc80 is a key member
of the kinetochore network and serves as a sort of “landing pad”
for the microtubule-centromere connection. Although Ndc80’s genetics and
biochemistry have been extensively characterized, the mechanisms behind its
activities have until now remained a mystery.
“Our first ever subnanometer model of Ndc80 shows that the protein complex
binds the microtubule with a tubulin monomer repeat that is sensitive to tubulin
conformation,” Nogales says. “Furthermore, Ndc80 complexes self-associate
along microtubule protofilaments via interactions that are mediated by the amino-terminal
tail of the Ndc80 protein, which is the site of phospho-regulation by the Aurora
B kinase.”
The Aurora B kinase is an enzyme that ensures the correction of any improper
microtubule-kinetochore attachments – faulty attachments will result in
unequal segregation of the genetic material, such as both chromatides going
to the same daughter cell. In their paper, Nogales and her co-authors contend
that Ndc80’s mode of interaction with the microtubule and its oligomerization
provide a means by which the Aurora B kinase can regulate the stability of the
load-bearing Ndc80-microtubule attachments.
“The Aurora B kinase corrects wrong microtubule-kinetochore attachments
by phosphorylating proteins in the kinetochore,” Nogales says. “Ndc80
is a major substrate of this regulation. Our work shows that if phosphorylated
by Aurora B, attachments are not robust because there is no oligomerization
of Ndc80s.”