Exportin 1 inhibition attenuates nuclear factor-kappaB-dependent gene expression.
Activation of nuclear factor (NF)-kappaB is mediated by signal-induced phosphorylation of IkappaBalpha, subsequent IkappaBalpha degradation, and then translocation of unbound NF-kappaB to the nucleus. Termination of gene expression occurs when IkappaBalpha binds NF-kappaB subunits (Rel A) in the nucleus. Leptomycin B specifically inhibits export of IkappaBalpha and the inactive IkappaBalpha/Rel A complex via the nuclear export protein exportin 1. We hypothesized that inhibition of IkappaBalpha nuclear export would increase nuclear IkappaBalpha and attenuate NF-kappaB inflammatory gene expression in pulmonary microvascular endothelial cells. We found that inhibition of exportin 1 causes nuclear accumulation of both endogenous NF-kappaB (Rel A) and IkappaBalpha. IL-1beta causes nuclear accumulation of NF-kappaB (Rel A) but does not increase nuclear IkappaBalpha. Inhibition of exportin 1 before IL-1beta prevented an increase in the nuclear ratio of NF-kappaB (Rel A) to IkappaBalpha and decreases NF-kappaB DNA binding. Furthermore, inhibition of exportin 1 attenuates IL-1beta-induced phosphorylation of IkappaBalpha without affecting IkappaB kinase phosphorylation. Lastly, inhibition of exportin 1 attenuates monocyte chemoattractant protein, IL-8, and intercellular adhesion molecule expression in response to IL-1beta stimulation. We suggest that the decrease in cell activation due to exportin 1 inhibition is a result of termination of NF-kappaB DNA binding due to increased concentration of IkappaBalpha in the nucleus.
Walsh
Mark D
MD
Department of Surgery, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA. mark.walsh@uchsc.edu
Hamiel
Christine R
CR
Banerjee
Anirban
A
Cheng
Aaron M
AM
Escobar
Guillermo
G
McIntyre
Robert C
RC
D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism*; D015151:Immunoblotting; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D053583:Interleukin-1beta / Q000494:pharmacology; D016209:Interleukin-8 / Q000378:metabolism; D028884:Karyopherins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D018160:Receptors, Cytoplasmic and Nuclear / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D000097606:Exportin 1 Protein
D005231:Fatty Acids, Unsaturated; D051550:I-kappa B Kinase; D018799:Intercellular Adhesion Molecule-1; D053583:Interleukin-1beta; D016209:Interleukin-8; D028884:Karyopherins; D016328:NF-kappa B; D018160:Receptors, Cytoplasmic and Nuclear; D000097606:Exportin 1 Protein; C038753:leptomycin B