Kutay lec notes - Kutay et al. 1997 Export of importin from...

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Unformatted text preview: Kutay et al. 1997 Export of importin from the nucleus is mediated by a speciFc nuclear transport factor. Cell 86:1061-1071. Kutay et al. 1997 Export of importin from the nucleus is mediated by a speciFc nuclear transport factor. Cell 86:1061-1071. A. Major question There is an assymetric import/re-export cycle: The import of cargo and the re- export of the receptors has to be coupled for import to be efFcient. How is importin efFciently recycled to the cytoplasm for further rounds of import? Importin is part of the NLS receptor that mediates nuclear import of cargo proteins with a classic NLS. Importin Importin receptor binds NLS-containing cargo and mediates translocation across the nuclear pore The complex is disrupted after importin binds RanGTP in the nucleus. Importin acts as an export receptor when bound to RanGTP, but how is importin exported? B. Hypothesis : There must be an export receptor for importin just like there is an import receptor for proteins that must enter the nucleus an importin- export receptor would have certain properties Bind importin together with RanGTP (nucleus) release importin when RanGTP is converted to RanGDP (in cytoplasm) *note:NTF2 is the import receptor for Ran Set up a system where proteins are transported across a nuclear membrane (a HeLa cell) C. Approach: Cell Biology and Biochemistry Experimental Strategy: C. Approach - permeabilization. System must be able to be manipulated Cells permeabilized Is cytoplasmic envelope intact? Is nuclear envelope intact? C. Approach - permeabilization. System must be able to be manipulated Cells permeabilized C. Approach - permeabilization. System must be able to be manipulated Cells permeabilized Soluble factors are lost Allows authors to add deFned components to the system while using the native nuclear envelope for transport C. Reconstitutionon Add back energy (GTP and way to regenerate it from ATP - more phosphate available from creatine phosphate) AND...
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This note was uploaded on 10/19/2009 for the course B M B 460 taught by Professor Hanna-rose,w during the Spring '08 term at Pennsylvania State University, University Park.

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Kutay lec notes - Kutay et al. 1997 Export of importin from...

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