A non-ring-like form of the Dam1 complex modulates microtubule dynamics in fission yeast.
The Dam1 complex is a kinetochore component that couples chromosomes to the dynamic ends of kinetochore microtubules (kMTs). Work in the budding yeast Saccharomyces cerevisiae has shown that the Dam1 complex forms a 16-unit ring encircling and tracking the tip of a MT in vitro, consistent with its cellular function as a coupler. Dam1 also forms smaller, nonring patches in vitro that track the dynamic ends of MTs. However, the identity of Dam1's functional form in vivo remains unknown. Here we report a comprehensive in vivo characterization of Dam1 in the fission yeast Schizosaccharomyces pombe. In addition to their dense localizations on kinetochores and spindle MTs during mitosis, we identify that Dam1 is also localized onto cytoplasmic MTs as discrete spots in interphase, providing the unique opportunity to analyze Dam1 oligomers at the single-particle resolution in live cells. Such analysis shows that each oligomer contains one to five copies of Dam1, and is able to "switch-rail" while moving along MTs, precluding the possibility of a 16-unit encircling structure. Dam1 patches track the plus ends of the shortening, but not the elongating, MTs and retard MT depolymerization. Together with Mal3, the EB1-like MT-interacting protein, cytoplasmic Dam1 plays an important role in maintaining proper cell shape. In mitosis, kinetochore-associated Dam1 appears to facilitate kMT depolymerization. Together, our findings suggest that patches, instead of rings, are the physiologically functional forms of Dam1 in pombe. Our findings help establish the benchmark parameters of the Dam1 coupler and elucidate the mechanism of its functions.
Gao
Qi
Q
Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030, USA.
Courtheoux
Thibault
T
Gachet
Yannick
Y
Tournier
Sylvie
S
He
Xiangwei
X
D020090:Chromosome Segregation / Q000502:physiology; D003593:Cytoplasm / Q000378:metabolism; D018628:Gene Dosage; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D018386:Kinetochores / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism*; D008938:Mitosis / Q000502:physiology; D009154:Mutation; D011108:Polymers / Q000378:metabolism; D011485:Protein Binding; D012568:Schizosaccharomyces / Q000235:genetics / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics / Q000378:metabolism*
C527984:Dam1 protein, S pombe; D008869:Microtubule-Associated Proteins; D011108:Polymers; D029702:Schizosaccharomyces pombe Proteins; D049452:Green Fluorescent Proteins