Sequential binding of import ligands to distinct nucleopore regions during their nuclear import

Science

Volumn 273, Issue 5282, 1996, Pages 1729-1732

  Panté, Nelly ^a   Aebi, Ueli ^a  

^a UNIVERSITY OF BASEL   (Switzerland)

Author keywords

[No Author keywords available]

Indexed keywords

NUCLEOPLASMIN;

ARTICLE; CELL MEMBRANE PERMEABILITY; ELECTRON MICROSCOPY; GENE TRANSLOCATION; LIGAND BINDING; NEUROFILAMENT; NONHUMAN; PRIORITY JOURNAL; PROTEIN ASSEMBLY; PROTEIN PROTEIN INTERACTION; PROTEIN TRANSPORT; SEQUENCE HOMOLOGY;

EID: 0029797940     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.273.5282.1729     Document Type: Article

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32. Colloidal gold particles (∼8 nm or ∼14 nm in diameter) were prepared by the method described by J. W. Slot and H. J. Geuze [Eur. J. Cell Biol. 38, 87 (1985)]. Nucleoplasmin was conjugated to colloidal gold particles as described [W. Baschong and N. G. Wrigley, J. Electron Microsc. Tech. 14, 313 (1990)].
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34. Mature (stage six) oocytes were surgically removed from female Xenopus laevis as described (16) and stored in modified Berth's saline [MBS; see (16) for buffer composition]. Oocytes were defolliculated by treatment with collagenase (5 mg/ml) (Sigma) in calcium-free MBS for 3 hours. Oocytes were then washed with MBS and used for microinjection within the next 2 days. NP-gold (50 nl) was microinjected into the cytoplasm of each oocyte far away from the nucleus. The injected oocytes were incubated in MBS buffer at room temperature for the times indicated.
35. We thank U. Sauder for her help with embedding and thin-sectioning, M. Döbeli for help with the data analysis, M. Hall for providing nucleoplasmin, and H. Frefel and M. Zoller for their expert photographic work. Supported by the Canton Basel-Stadt, the M. E. Müller Foundation of Switzerland, and by research grants from the Swiss National Science Foundation and the Human Frontier Science Program Organization.