A Single-Cell Biochemistry Approach Reveals PAR Complex Dynamics during Cell Polarization

被引:94
作者
Dickinson, Daniel J. [1 ,2 ,7 ]
Schwager, Francoise [3 ]
Pintard, Lionel [4 ,5 ,6 ]
Gotta, Monica [3 ]
Goldstein, Bob [1 ,2 ]
机构
[1] Univ N Carolina, Dept Biol, Chapel Hill, NC 27599 USA
[2] Univ N Carolina, Lineberger Comprehens Canc Ctr, Chapel Hill, NC 27599 USA
[3] Univ Geneva, Dept Cell Physiol & Metab, Med Fac, 1 Rue Michel Servet, CH-1211 Geneva, Switzerland
[4] CNRS, Inst Jacques Monod, Cell Cycle & Dev Team, F-75013 Paris, France
[5] Univ Paris Diderot, F-75013 Paris, France
[6] Sorbonne Paris Cite, UMR7592, F-75013 Paris, France
[7] Univ Texas Austin, Dept Mol Biosci, NHB 2-606,Stop A5000,100 East 24th St, Austin, TX 78712 USA
基金
瑞士国家科学基金会;
关键词
C-ELEGANS EMBRYOS; N-TERMINAL DOMAIN; PROTEIN-KINASE-C; CAENORHABDITIS-ELEGANS; PARTICLE TRACKING; ESCHERICHIA-COLI; LIVE-CELL; POLARITY; LOCALIZATION; ASSOCIATION;
D O I
10.1016/j.devcel.2017.07.024
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
Regulated protein-protein interactions are critical for cell signaling, differentiation, and development. For the study of dynamic regulation of protein interactions in vivo, there is a need for techniques that can yield time-resolved information and probe multiple protein binding partners simultaneously, using small amounts of starting material. Here we describe a single-cell protein interaction assay. Single-cell lysates are generated at defined time points and analyzed using single-moleculepull-down, yielding information about dynamic protein complex regulation in vivo. We established the utility of this approach by studying PAR polarity proteins, which mediate polarization of many animal cell types. We uncovered striking regulation of PAR complex composition and stoichiometry during Caenorhabditis elegans zygotepolarization, which takes place in less than 20 min. PAR complex dynamics are linked to the cell cycle by Polo-like kinase 1 and govern the movement of PAR proteins to establish polarity. Our results demonstrate an approach to study dynamic biochemical events in vivo.
引用
收藏
页码:416 / +
页数:30
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