RESUMO
Mutations in the human presenilin genes (PS1 or PS2) have been linked to autosomal dominant, early onset Alzheimer's disease (AD). Presenilins, probably as an essential part of gamma-secretase, modulate gamma-cleavage of the amyloid protein precursor (APP) to the amyloid beta-peptide (Abeta). Mutations in sel-12, a Caenorhabditis elegans presenilin homologue, cause a defect in egg laying that can be suppressed by loss of function mutations in a second gene, SEL-10. SEL-10 protein is a homologue of yeast Cdc4, a member of the SCF (Skp1-Cdc53/CUL1-F-box protein) E2-E3 ubiquitin ligase family. In this study, we show that human SEL-10 interacts with PS1 and enhances PS1 ubiquitination, thus altering cellular levels of unprocessed PS1 and its N- and C-terminal fragments. Co-transfection of sel-10 and APP cDNAs in HEK293 cells leads to an alteration in the metabolism of APP and to an increase in the production of amyloid beta-peptide, the principal component of amyloid plaque in Alzheimer's disease.
Assuntos
Peptídeos beta-Amiloides/biossíntese , Proteínas de Caenorhabditis elegans , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas F-Box , Proteínas de Helminto/genética , Proteínas de Helminto/metabolismo , Proteínas de Membrana/metabolismo , Ubiquitina-Proteína Ligases , Ubiquitina/metabolismo , Sequência de Aminoácidos , Animais , Caenorhabditis elegans , Proteínas de Ciclo Celular/farmacologia , Linhagem Celular , Clonagem Molecular , DNA Complementar/genética , Proteína 7 com Repetições F-Box-WD , Proteínas de Helminto/farmacologia , Humanos , Rim/citologia , Rim/metabolismo , Camundongos , Dados de Sequência Molecular , Especificidade de Órgãos , Presenilina-1 , Ligação Proteica/fisiologia , Saccharomyces cerevisiae , Proteínas de Saccharomyces cerevisiae , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , TransfecçãoRESUMO
Nicastrin is a component of the gamma-secretase complex that has been shown to adhere to presenilin-1 (PS1), Notch, and APP. Here we demonstrate that Nicastrin-deficient mice showed a phenotype that is indistinguishable from PS1/PS2 double knock-out mice, whereas heterozygotes were healthy and viable. Fibroblasts derived from Nicastrin-deficient embryos were unable to generate amyloid beta-peptide and failed to release the intracellular domain of APP- or Notch1-Gal4-VP16 fusion proteins. Additionally, C- and N-terminal fragments of PS1 and the C-terminal fragments of PS2 were not detectable in Nicastrin-null fibroblasts, whereas full-length PS1 accumulated in null fibroblasts, indicating that Nicastrin is required for the endoproteolytic processing of presenilins. Interestingly, cells derived from Nicastrin heterozygotes produced relatively higher levels of amyloid beta-peptide whether the source was endogenous mouse or transfected human APP. These data demonstrate that Nicastrin is essential for the gamma-secretase cleavage of APP and Notch in mammalian cells and that Nicastrin has both positive and negative functions in the regulation of gamma-secretase activity.