Chapter title |
Nuclear Protein Transport in Digitonin Permeabilized Cells.
|
---|---|
Chapter number | 29 |
Book title |
The Nuclear Envelope
|
Published in |
Methods in molecular biology, January 2016
|
DOI | 10.1007/978-1-4939-3530-7_29 |
Pubmed ID | |
Book ISBNs |
978-1-4939-3528-4, 978-1-4939-3530-7
|
Authors |
Stephen A. Adam Ph.D., Stephen A. Adam |
Editors |
Sue Shackleton, Philippe Collas, Eric C. Schirmer |
Abstract |
The high concentration of cholesterol in the plasma membrane relative to the endomembranes of eukaryotic cells allows the selective permeabilization of the plasma membrane with the glycoside digitonin leaving the intracellular membrane bound organelles intact. In this chapter, we describe the basic method to use digitonin permeabilized cells to reconstitute the transport of proteins containing nuclear localization signals into the nucleus. The assay requires only a target cell line that can be permeabilized with digitonin, a source of soluble transport factors, typically provided by the cytosol fraction of cultured cells, and a cargo protein of interest. No other specialized equipment is required other than a fluorescence microscope. The assay can be used to identify transport factors required to transport specific proteins, to study the regulation of protein transport, or to study nuclear protein transport under different conditions. |
Mendeley readers
Geographical breakdown
Country | Count | As % |
---|---|---|
Unknown | 9 | 100% |
Demographic breakdown
Readers by professional status | Count | As % |
---|---|---|
Student > Ph. D. Student | 2 | 22% |
Researcher | 2 | 22% |
Other | 1 | 11% |
Student > Master | 1 | 11% |
Unknown | 3 | 33% |
Readers by discipline | Count | As % |
---|---|---|
Biochemistry, Genetics and Molecular Biology | 4 | 44% |
Agricultural and Biological Sciences | 1 | 11% |
Chemistry | 1 | 11% |
Unknown | 3 | 33% |