title: Regulation of functional nuclear pore size in fibroblasts.
abstract: Protein-NLS-coated gold particles up to approximately 250 A in diameter are transported through the nuclear pores in normal, proliferating BALB/c 3T3 cells. This size can increase or decrease, depending on cellular activity. It has been suggested that increases in functional pore size are related to a reduction in the amount of available p53. To further test this hypothesis, we investigated the effects of cycloheximide and pifithrin-alpha, which inhibits p53-dependent transcriptional activation, on nuclear transport. After 3 hours in cycloheximide, there was a significant increase in the size of the gold particles that entered the nucleoplasm. When the incubation period was extended to 6 hours or longer, transport capacity returned to the control level. By using proteasome inhibitors, it was shown that the cycloheximide-dependent increase in functional pore size was due to the inhibition of protein synthesis, consistent with the fact that p53 is a short-lived protein, and requires the activity of at least two different factors. Although cycloheximide increases the functional diameter of the channel available for signal-mediated transport by approximately 60 A, it had no significant effect on either the import rate of small NLS-containing substrates (FITC-BSA-NLS), or passive diffusion of fluorescent-labeled proteins across the envelope. This suggests that changes in transport capacity were not caused by an increase in overall pore diameter but instead are due to a transient increase in pore size that accompanies signal-mediated transport. Pifithrin-alpha also caused an increase in functional pore diameter without altering the import rate of FITC-BSA-NLS, providing further support for the view that p53 can initiate changes in nuclear transport capacity.
authors.affiliation: Department of Anatomy and Cell Biology, University of Florida, College of Medicine, Gainesville, FL 32610, USA. feldherr@anatomy.med.ufl.edu
authors.forename: C M
authors.initials: CM
authors.lastname: Feldherr
authors.forename: D
authors.initials: D
authors.lastname: Akin
authors.forename: R J
authors.initials: RJ
authors.lastname: Cohen
mesh_terms: D016475:3T3 Cells; D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000502:physiology; D000818:Animals; D052160:Benzothiazoles; D003513:Cycloheximide / Q000494:pharmacology; D003546:Cysteine Endopeptidases / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004058:Diffusion / Q000187:drug effects; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism / Q000502:physiology* / Q000648:ultrastructure*; D017979:Gold Colloid / Q000378:metabolism; D007976:Leupeptins / Q000494:pharmacology; D051379:Mice; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism; D019913:Nuclear Localization Signals / Q000187:drug effects / Q000502:physiology; D022022:Nuclear Pore / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D046988:Proteasome Endopeptidase Complex; D013844:Thiazoles / Q000494:pharmacology; D014050:Toluene / Q000031:analogs & derivatives / Q000494:pharmacology; D016159:Tumor Suppressor Protein p53 / Q000502:physiology
chemical_list: D052160:Benzothiazoles; D015853:Cysteine Proteinase Inhibitors; D017979:Gold Colloid; D007976:Leupeptins; D009097:Multienzyme Complexes; D019913:Nuclear Localization Signals; D013844:Thiazoles; D016159:Tumor Suppressor Protein p53; D014050:Toluene; D003513:Cycloheximide; C121565:pifithrin; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde