This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.During prometaphase in vertebrate cells, kinetochores interact with microtubules and frequently exhibit poleward movement (PM) prior to bi-orientation. In fission yeast there are three presumptive minus-end directed motors (dynein and two KAR3 homologs, Pkl1p and Klp2p), each of which is a good candidate for PM. To study their respective contributions we have analyzed the kinetochore dynamics in cells whose kinetochore-pole connections were disrupted prior to mitosis using the cold-sensitive mutation in beta-tubulin nda3. Live imaging and immunofluorescence of fixed samples have strongly suggested that Klp2p promotes processivity of the PM, while Pkl1p and dynein ensure efficient bi-orientation. We have used EM tomography on S. pombe cells that have been prepared by rapid freezing and freeze substitution to seek structural evidence for such roles In klp2 deletion cells delayed in kinetochore retrieval we have found long intranuclear astral microtubules, which are likely to connect poles and distant kintochores. These microtubules frequently contain abnormal occlusions at their plus ends, suggesting that Klp2p promotes processivity of kinetochore PM by sustaining normal kinetochore microtubules disassembly. In the absence of Pkl1p and dynein motors the overall spindle structure is normal but the pole organization is disrupted. Based on these structural findings we propose that Pklp1 contributes to pole by preventing splitting of the pole-associated material, due to spindle-generated forces, while dynein promotes attachments between the pole and microtubule minus ends.
| Giddings Jr, Thomas H; Morphew, Mary K; McIntosh, J Richard (2017) Preparing Fission Yeast for Electron Microscopy. Cold Spring Harb Protoc 2017: |
| Zhao, Xiaowei; Schwartz, Cindi L; Pierson, Jason et al. (2017) Three-Dimensional Structure of the Ultraoligotrophic Marine Bacterium ""Candidatus Pelagibacter ubique"". Appl Environ Microbiol 83: |
| Brown, Joanna R; Schwartz, Cindi L; Heumann, John M et al. (2016) A detailed look at the cytoskeletal architecture of the Giardia lamblia ventral disc. J Struct Biol 194:38-48 |
| Saheki, Yasunori; Bian, Xin; Schauder, Curtis M et al. (2016) Control of plasma membrane lipid homeostasis by the extended synaptotagmins. Nat Cell Biol 18:504-15 |
| Höög, Johanna L; Lacomble, Sylvain; Bouchet-Marquis, Cedric et al. (2016) 3D Architecture of the Trypanosoma brucei Flagella Connector, a Mobile Transmembrane Junction. PLoS Negl Trop Dis 10:e0004312 |
| Park, J Genevieve; Palmer, Amy E (2015) Properties and use of genetically encoded FRET sensors for cytosolic and organellar Ca2+ measurements. Cold Spring Harb Protoc 2015:pdb.top066043 |
| McCoy, Kelsey M; Tubman, Emily S; Claas, Allison et al. (2015) Physical limits on kinesin-5-mediated chromosome congression in the smallest mitotic spindles. Mol Biol Cell 26:3999-4014 |
| Höög, Johanna L; Lötvall, Jan (2015) Diversity of extracellular vesicles in human ejaculates revealed by cryo-electron microscopy. J Extracell Vesicles 4:28680 |
| Marc, Robert E; Anderson, James R; Jones, Bryan W et al. (2014) The AII amacrine cell connectome: a dense network hub. Front Neural Circuits 8:104 |
| Weber, Britta; Tranfield, Erin M; Höög, Johanna L et al. (2014) Automated stitching of microtubule centerlines across serial electron tomograms. PLoS One 9:e113222 |
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