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Coordination of multiple kinesin and myosin motors is required for intracellular transport, cell motility and mitosis. However, comprehensive resources that allow systems analysis of the localization and interplay between motors in living cells do not exist. Here, we generated a library of 243 amino- and carboxy-terminally tagged mouse and human bacterial artificial chromosome transgenes to establish 227 stably transfected HeLa cell lines, 15 mouse embryonic stem cell lines and 1 transgenic mouse line. The cells were characterized by expression and localization analyses and further investigated by affinity-purification mass spectrometry, identifying 191 candidate protein-protein interactions. We illustrate the power of this resource in two ways. First, by characterizing a network of interactions that targets CEP170 to centrosomes, and second, by showing that kinesin light-chain heterodimers bind conventional kinesin in cells. Our work provides a set of validated resources and candidate molecular pathways to investigate motor protein function across cell lineages.

Original publication

DOI

10.1038/ncb2689

Type

Journal article

Journal

Nat Cell Biol

Publication Date

03/2013

Volume

15

Pages

325 - 334

Keywords

Animals, Biological Transport, Biomarkers, Blotting, Western, Cell Movement, Centrosome, Chromatography, Affinity, Chromosomes, Artificial, Bacterial, Embryonic Stem Cells, Fluorescent Antibody Technique, Gene Expression Profiling, Genomics, Green Fluorescent Proteins, HeLa Cells, Humans, Immunoprecipitation, Kinesin, Mice, Mice, Transgenic, Microtubule-Associated Proteins, Microtubules, Mitosis, Myosins, Neuroblastoma, Neurons, Oligonucleotide Array Sequence Analysis, Phosphoproteins, Phylogeny, Protein Multimerization, RNA, Messenger, Real-Time Polymerase Chain Reaction, Reverse Transcriptase Polymerase Chain Reaction, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization, Stem Cells, Transgenes