title: CRM1 protein-mediated regulation of nuclear clusterin (nCLU), an ionizing radiation-stimulated, Bax-dependent pro-death factor.
abstract: Expression of the clusterin (CLU) gene results in the synthesis of a conventional secretory isoform set (pre- and mature secretory clusterin proteins, psCLU/sCLU), as well as another set of intracellular isoforms, appearing in the cytoplasm (pre-nuclear CLU, pnCLU) and in the nucleus as an ∼55-kDa mature nuclear clusterin (nCLU) form. These two isoform sets have opposing cell functions: pro-survival and pro-death, respectively. Although much is known about the regulation and function of sCLU as a pro-survival factor, the regulation and function of endogenous nCLU in cell death are relatively unexplored. Here, we show that depletion of endogenous nCLU protein using siRNA specific to its truncated mRNA increased clonogenic survival of ionizing radiation (IR)-exposed cells. nCLU-mediated apoptosis was Bax-dependent, and lethality correlated with accumulation of mature nCLU protein. nCLU accumulation was regulated by CRM1 because binding between CRM1 and nCLU proteins was significantly diminished by leptomycin B (LMB), and nuclear levels of nCLU protein were significantly enhanced by LMB and IR co-treatment. Moreover, LMB treatment significantly enhanced IR-induced nCLU-mediated cell death responses. Importantly, bax(-/-) and bax(-/-)/bak(-/-) double knock-out cells were resistant to nCLU-mediated cell death, whereas bak(-/-) or wild-type bax(+/+)/bak(+/+) cells were hypersensitive. The regulation of nCLU by CRM1 nuclear export/import may explain recent clinical results showing that highly malignant tumors have lost the ability to accumulate nCLU levels, thereby avoiding growth inhibition and cell death.
authors.affiliation: Department of Pharmacology, Case Western Reserve University, Cleveland, Ohio 44109, USA. ksl9@case.edu
authors.forename: Konstantin S
authors.initials: KS
authors.lastname: Leskov
authors.forename: Shinako
authors.initials: S
authors.lastname: Araki
authors.forename: John-Paul
authors.initials: JP
authors.lastname: Lavik
authors.forename: Jose A
authors.initials: JA
authors.lastname: Gomez
authors.forename: Vivian
authors.initials: V
authors.lastname: Gama
authors.forename: Efstathios S
authors.initials: ES
authors.lastname: Gonos
authors.forename: Ioannis P
authors.initials: IP
authors.lastname: Trougakos
authors.forename: Shigemi
authors.initials: S
authors.lastname: Matsuyama
authors.forename: David A
authors.initials: DA
authors.lastname: Boothman
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects*; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D051152:Clusterin / Q000235:genetics / Q000378:metabolism*; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D005720:Gamma Rays; D006801:Humans; D028884:Karyopherins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011836:Radiation Tolerance / Q000187:drug effects / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000235:genetics / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism*; D000097606:Exportin 1 Protein
chemical_list: D000903:Antibiotics, Antineoplastic; D051152:Clusterin; D005231:Fatty Acids, Unsaturated; D028884:Karyopherins; D020033:Protein Isoforms; D018160:Receptors, Cytoplasmic and Nuclear; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein; D000097606:Exportin 1 Protein; C494772:BAK1 protein, human; C494826:BAX protein, human; C494773:Bak1 protein, mouse; C494827:Bax protein, mouse; C495242:CLU protein, human; C495243:Clu protein, mouse; C038753:leptomycin B