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Science 14 October 2005:
Vol. 310. no. 5746, pp. 310 - 314
DOI: 10.1126/science.1113230
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Reports

Counting Cytokinesis Proteins Globally and Locally in Fission Yeast

Jian-Qiu Wu1 and Thomas D. Pollard1,2*

We used fluorescence microscopy to measure global and local concentrations of 28 cytoskeletal and signaling proteins fused to yellow fluorescent protein (YFP) in the fission yeast Schizosaccharomyces pombe. Native promoters controlled the expression of these functional YFP fusion proteins. Fluorescence measured by microscopy or flow cytometry was directly proportional to protein concentration measured by quantitative immunoblotting. Global cytoplasmic concentrations ranged from 0.04 (formin Cdc12p) to 63 micromolar (actin). Proteins concentrated up to 100 times in contractile rings and 7500 times in spindle pole bodies at certain times in the cell cycle. This approach can be used to measure the global and local concentrations of any fusion protein.

1 Department of Molecular, Cellular, and Developmental Biology, Yale University, New Haven, CT 06520–8103, USA.
2 Departments of Cell Biology and Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520–8103, USA.

* To whom correspondence should be addressed. E-mail: thomas.pollard{at}yale.edu

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Science. ISSN 0036-8075 (print), 1095-9203 (online)