title: SILAC-based proteomic quantification of chemoattractant-induced cytoskeleton dynamics on a second to minute timescale.
abstract: Cytoskeletal dynamics during cell behaviours ranging from endocytosis and exocytosis to cell division and movement is controlled by a complex network of signalling pathways, the full details of which are as yet unresolved. Here we show that SILAC-based proteomic methods can be used to characterize the rapid chemoattractant-induced dynamic changes in the actin-myosin cytoskeleton and regulatory elements on a proteome-wide scale with a second to minute timescale resolution. This approach provides novel insights in the ensemble kinetics of key cytoskeletal constituents and association of known and novel identified binding proteins. We validate the proteomic data by detailed microscopy-based analysis of in vivo translocation dynamics for key signalling factors. This rapid large-scale proteomic approach may be applied to other situations where highly dynamic changes in complex cellular compartments are expected to play a key role.
authors.affiliation: Division of Cell and Developmental Biology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, UK.
authors.forename: Grzegorz J
authors.initials: GJ
authors.lastname: Sobczyk
authors.affiliation: 1] Division of Cell and Developmental Biology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, UK [2].
authors.forename: Jun
authors.initials: J
authors.lastname: Wang
authors.affiliation: Division of Cell and Developmental Biology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, UK.
authors.forename: Cornelis J
authors.initials: CJ
authors.lastname: Weijer
mesh_terms: D000596:Amino Acids / Q000378:metabolism; D002630:Chemotactic Factors / Q000494:pharmacology*; D000242:Cyclic AMP / Q000494:pharmacology; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism; D003599:Cytoskeleton / Q000187:drug effects* / Q000378:metabolism; D004023:Dictyostelium / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D055786:Gene Knockout Techniques; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007553:Isotope Labeling / Q000379:methods; D007700:Kinetics; D013058:Mass Spectrometry; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D009154:Mutation; D021381:Protein Transport / Q000187:drug effects; D020543:Proteome / Q000235:genetics / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors
chemical_list: D000596:Amino Acids; D002630:Chemotactic Factors; D003598:Cytoskeletal Proteins; D020543:Proteome; D015800:Protozoan Proteins; D049452:Green Fluorescent Proteins; D000242:Cyclic AMP