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A fluorescent reporter for mapping cellular protein-protein interactions in time and space.


ABSTRACT: We introduce a fluorescent reporter for monitoring protein-protein interactions in living cells. The method is based on the Split-Ubiquitin method and uses the ratio of two auto-fluorescent reporter proteins as signal for interaction (SPLIFF). The mating of two haploid yeast cells initiates the analysis and the interactions are followed online by two-channel time-lapse microscopy of the diploid cells during their first cell cycle. Using this approach we could with high spatio-temporal resolution visualize the differences between the interactions of the microtubule binding protein Stu2p with two of its binding partners, monitor the transient association of a Ran-GTPase with its receptors at the nuclear pore, and distinguish between protein interactions at the polar cortical domain at different phases of polar growth. These examples further demonstrate that protein-protein interactions identified from large-scale screens can be effectively followed up by high-resolution single-cell analysis.

SUBMITTER: Moreno D 

PROVIDER: S-EPMC3619943 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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A fluorescent reporter for mapping cellular protein-protein interactions in time and space.

Moreno Daniel D   Neller Joachim J   Kestler Hans A HA   Kraus Johann J   Dünkler Alexander A   Johnsson Nils N  

Molecular systems biology 20130101


We introduce a fluorescent reporter for monitoring protein-protein interactions in living cells. The method is based on the Split-Ubiquitin method and uses the ratio of two auto-fluorescent reporter proteins as signal for interaction (SPLIFF). The mating of two haploid yeast cells initiates the analysis and the interactions are followed online by two-channel time-lapse microscopy of the diploid cells during their first cell cycle. Using this approach we could with high spatio-temporal resolution  ...[more]

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