Regulated fast nucleocytoplasmic shuttling observed by reversible protein highlighting.
The observation of the regulation of fast protein dynamics in a cellular context requires the development of reliable technologies. Here, a signal regulation cascade reliant on the stimulus-dependent acceleration of the bidirectional flow of mitogen-activated protein kinase (extracellular signal-regulated kinase) across the nuclear envelope was visualized by reversible protein highlighting. Light-induced conversion between the bright and dark states of a monomeric fluorescent protein engineered from a novel coral protein was employed. Because of its photochromic properties, the protein could be highlighted, erased, and highlighted again in a nondestructive manner, allowing direct observation of regulated fast nucleocytoplasmic shuttling of key signaling molecules.
Ando
Ryoko
R
Laboratory for Cell Function and Dynamics, Advanced Technology Development Group, Brain Science Institute, RIKEN, 2-1 Hirosawa, Wako-city, Saitama, 351-0198, Japan.
Mizuno
Hideaki
H
Miyawaki
Atsushi
A
D021581:Active Transport, Cell Nucleus; D000595:Amino Acid Sequence; D000818:Animals; D037421:Anthozoa; D019556:COS Cells; D002467:Cell Nucleus / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005453:Fluorescence; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008027:Light; D008164:Luminescent Proteins / Q000737:chemistry / Q000378:metabolism*; D020935:MAP Kinase Signaling System; D018613:Microscopy, Confocal; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D008969:Molecular Sequence Data; D009685:Nuclear Envelope / Q000378:metabolism*; D010766:Phosphorylation; D021381:Protein Transport; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014162:Transfection; D028961:beta Karyopherins / Q000378:metabolism
D008164:Luminescent Proteins; D011994:Recombinant Proteins; D028961:beta Karyopherins; D004815:Epidermal Growth Factor; D048052:Mitogen-Activated Protein Kinase 3