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Analysis of interphase node proteins in fission yeast by quantitative and super resolution fluorescence microscopy

Matthew Akamatsu, Yu Lin, Joerg Bewersdorf, Thomas D. Pollard
doi: https://doi.org/10.1101/137794
Matthew Akamatsu
1Departments of Molecular Cellular and Developmental Biology, Yale University, PO Box 208103 New Haven, CT 06520-8103 USA
2Departments of Molecular Biophysics and Biochemistry, Yale University, PO Box 208103 New Haven, CT 06520-8103 USA
4Integrated Graduate Program in Physical and Engineering Biology, Yale University, PO Box 208103 New Haven, CT 06520-8103 USA
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Yu Lin
3Department of Cell Biology, Yale University, PO Box 208103 New Haven, CT 06520-8103 USA
4Integrated Graduate Program in Physical and Engineering Biology, Yale University, PO Box 208103 New Haven, CT 06520-8103 USA
5Department of Biomedical Engineering, Yale University, PO Box 208103 New Haven, CT 06520-8103 USA
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Joerg Bewersdorf
4Integrated Graduate Program in Physical and Engineering Biology, Yale University, PO Box 208103 New Haven, CT 06520-8103 USA
5Department of Biomedical Engineering, Yale University, PO Box 208103 New Haven, CT 06520-8103 USA
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Thomas D. Pollard
1Departments of Molecular Cellular and Developmental Biology, Yale University, PO Box 208103 New Haven, CT 06520-8103 USA
2Departments of Molecular Biophysics and Biochemistry, Yale University, PO Box 208103 New Haven, CT 06520-8103 USA
3Department of Cell Biology, Yale University, PO Box 208103 New Haven, CT 06520-8103 USA
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  • For correspondence: thomas.pollard@yale.edu
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Abstract

We used quantitative confocal microscopy and FPALM super resolution microscopy of live fission yeast to investigate the structures and assembly of two types of interphase nodes, multiprotein complexes associated with the plasma membrane that merge together and mature into the precursors of the cytokinetic contractile ring. During the long G2 phase of the cell cycle seven different interphase node proteins maintain constant concentrations as they accumulate in proportion to cell volume. During mitosis the total numbers of type 1 node proteins (cell cycle kinases Cdr1p, Cdr2p, Wee1p, and anillin Mid1p) are constant even when the nodes disassemble. Quantitative measurements provide strong evidence that both types of nodes have defined sizes and numbers of constituent proteins, as observed for cytokinesis nodes. Type 1 nodes assemble in two phases, a burst at the end of mitosis, followed by steady increase during interphase to double the initial number. Type 2 nodes containing Blt1p, Rho-GEF Gef2p, and kinesin Klp8p remain intact throughout the cell cycle and are constituents of the contractile ring. They are released from the contractile ring as it disassembles and then associate with type 1 nodes around the equator of the cell during interphase.

Highlight summary FPALM super resolution microscopy and quantitative confocal microscopy reveal that interphase nodes, the precursors to the fission yeast cytokinetic contractile ring, are discrete unitary structures with defined sizes and ratios of component proteins. Type 1 nodes disassemble during mitosis, but type 2 nodes remain intact throughout the cell cycle.

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Posted May 14, 2017.
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Analysis of interphase node proteins in fission yeast by quantitative and super resolution fluorescence microscopy
Matthew Akamatsu, Yu Lin, Joerg Bewersdorf, Thomas D. Pollard
bioRxiv 137794; doi: https://doi.org/10.1101/137794
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Analysis of interphase node proteins in fission yeast by quantitative and super resolution fluorescence microscopy
Matthew Akamatsu, Yu Lin, Joerg Bewersdorf, Thomas D. Pollard
bioRxiv 137794; doi: https://doi.org/10.1101/137794

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