Evidence for intracellular cleavage of the Alzheimer's amyloid precursor in PC12 cells

The Alzheimer's amyloid precursor (APP) is cleaved by an unidentified enzyme (APP secretase) to produce soluble APP. Fractionation of PC12 cell homogenates in a detergent‐free buffer showed the presence of the Kunitz protease inhibitor (KPI)‐containing soluble APP (nexin II) in the particulate fraction. Digitonin or sodium carbonate treatment of this fraction solubilized nexin II suggesting that it is contained in the lumen of vesicles. Nexin II production was not affected by lysosomotropic agents, suggesting that APP secretase is not a lysosomal enzyme. Labelling of cell surface proteins by iodination failed to detect full‐length APP on the surface of PC12 cells, suggesting that most of this protein is located intracellularly. Furthermore, pulse‐chase experiments showed that nexin II is detected in cell extracts before it appears in the culture medium. Cellular nexin II was detected at zero time of chase after only 5 min of pulse labelling with 35S‐sulfate, indicated that APP secretase cleavage takes place immediately after APP is sulfated. Temperature block, pulse‐chase, and 35S‐sulfate‐labelling experiments suggested that APP is cleaved by APP secretase intracellularly in the trans‐Golgi network (TGN) or in a post‐Golgi compartment. © Wiley‐Liss, Inc.

[1]  P. Greengard,et al.  Chloroquine inhibits intracellular degradation but not secretion of Alzheimer beta/A4 amyloid precursor protein. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[2]  D. Selkoe,et al.  Processing of the amyloid protein precursor to potentially amyloidogenic derivatives. , 1992, Science.

[3]  L. Refolo,et al.  The Alzheimer amyloid precursor is associated with the detergent- insoluble cytoskeleton , 1991, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[4]  D. Selkoe,et al.  Processing of beta-amyloid precursor protein in microglia and astrocytes favors an internal localization over constitutive secretion , 1991, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[5]  K. Tanaka,et al.  "Secretase," Alzheimer amyloid protein precursor secreting enzyme is not sequence-specific. , 1991, Biochemical and biophysical research communications.

[6]  E. Johnson,et al.  Evidence for endocytosis-dependent proteolysis in the generation of soluble truncated nerve growth factor receptors by A875 human melanoma cells. , 1991, The Journal of biological chemistry.

[7]  N. Robakis,et al.  Exact cleavage site of Alzheimer amyloid precursor in neuronal PC-12 cells , 1991, Neuroscience Letters.

[8]  M. Pericak-Vance,et al.  Segregation of a missense mutation in the amyloid precursor protein gene with familial Alzheimer's disease , 1991, Nature.

[9]  D. Rappolee,et al.  Platelet alpha-granule fibrinogen, albumin, and immunoglobulin G are not synthesized by rat and mouse megakaryocytes. , 1990, The Journal of clinical investigation.

[10]  T. Oltersdorf,et al.  Cleavage of amyloid beta peptide during constitutive processing of its precursor. , 1990, Science.

[11]  I. Lieberburg,et al.  Mutation of the Alzheimer's disease amyloid gene in hereditary cerebral hemorrhage, Dutch type. , 1990, Science.

[12]  A. Schmaier,et al.  Protease nexin-II (amyloid beta-protein precursor): a platelet alpha-granule protein. , 1990, Science.

[13]  D. Price,et al.  Evidence that beta-amyloid protein in Alzheimer's disease is not derived by normal processing. , 1990, Science.

[14]  Y. Takahashi,et al.  Enzyme specificity of proteinase inhibitor region in amyloid precursor protein of Alzheimer's disease: different properties compared with protease nexin I. , 1990, Biochimica et biophysica acta.

[15]  W. Huttner,et al.  Cell-free protein sorting to the regulated and constitutive secretory pathways , 1990, Cell.

[16]  H. Lodish,et al.  Intracellular degradation of unassembled asialoglycoprotein receptor subunits: a pre-Golgi, nonlysosomal endoproteolytic cleavage , 1989, The Journal of cell biology.

[17]  V. Haroutunian,et al.  Differential brain expression of the Alzheimer's amyloid precursor protein. , 1989, The EMBO journal.

[18]  S. Younkin,et al.  Soluble derivatives of the β amyloid protein precursor of Alzheimer's disease are labeled by antisera to the β amyloid protein , 1989 .

[19]  N. Robakis,et al.  Nerve and epidermal growth factors induce the release of the Alzheimer amyloid precursor from PC 12 cell cultures. , 1989, Biochemical and biophysical research communications.

[20]  Carl W. Cotman,et al.  Protease nexin-II, a potent anti-chymotrypsin, shows identity to amyloid β-protein precursor , 1989, Nature.

[21]  K. Johnson-wood,et al.  The secreted form of the Alzheimer's amyloid precursor protein with the Kunitz domain is protease nexin-II , 1989, Nature.

[22]  R. Fine.  Vesicles without clathrin: Intermediates in bulk flow exocytosis , 1989, Cell.

[23]  D. Selkoe,et al.  The beta-amyloid protein precursor of Alzheimer disease has soluble derivatives found in human brain and cerebrospinal fluid. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[24]  Andreas Weidemann,et al.  Identification, biogenesis, and localization of precursors of Alzheimer's disease A4 amyloid protein , 1989, Cell.

[25]  Curtis,et al.  Dissection of Semliki Forest virus glycoprotein delivery from the trans-Golgi network to the cell surface in permeabilized BHK cells. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[26]  S. Shiojiri,et al.  Novel precursor of Alzheimer's disease amyloid protein shows protease inhibitory activity , 1988, Nature.

[27]  L. Villa-komaroff,et al.  Protease inhibitor domain encoded by an amyloid protein precursor mRNA associated with Alzheimer's disease , 1988, Nature.

[28]  B. Greenberg,et al.  A new A4 amyloid mRNA contains a domain homologous to serine proteinase inhibitors , 1988, Nature.

[29]  D. L. Quinn,et al.  The trans-most cisternae of the Golgi complex: A compartment for sorting of secretory and plasma membrane proteins , 1987, Cell.

[30]  W. Huttner,et al.  Tyrosine sulfation is a trans-Golgi-specific protein modification , 1987, The Journal of cell biology.

[31]  J. Tooze,et al.  An antibody specific for an endoproteolytic cleavage site provides evidence that pro-opiomelanocortin is packaged into secretory granules in AtT20 cells before its cleavage , 1987, The Journal of cell biology.

[32]  S. Pfeffer.  The endosomal concentration of a mannose 6-phosphate receptor is unchanged in the absence of ligand synthesis , 1987, The Journal of cell biology.

[33]  Richard G. W. Anderson,et al.  Proteolytic maturation of insulin is a post-Golgi event which occurs in acidifying clathrin-coated secretory vesicles , 1987, Cell.

[34]  P. S. St George-Hyslop,et al.  Amyloid beta protein gene: cDNA, mRNA distribution, and genetic linkage near the Alzheimer locus. , 1987, Science.

[35]  M. Lerman,et al.  Characterization and chromosomal localization of a cDNA encoding brain amyloid of Alzheimer's disease. , 1987, Science.

[36]  K. Grzeschik,et al.  The precursor of Alzheimer's disease amyloid A4 protein resembles a cell-surface receptor , 1987, Nature.

[37]  K. Simons,et al.  The trans Golgi network: sorting at the exit site of the Golgi complex. , 1986, Science.

[38]  G. Palade,et al.  Temperature-sensitive steps in the transport of secretory proteins through the Golgi complex in exocrine pancreatic cells. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[39]  P. Srivastava,et al.  Tumor rejection antigens of chemically induced sarcomas of inbred mice. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[40]  G. Glenner,et al.  Alzheimer's disease and Down's syndrome: sharing of a unique cerebrovascular amyloid fibril protein. , 1984, Biochemical and biophysical research communications.

[41]  G. Glenner,et al.  Alzheimer's disease: Initial report of the purification and characterization of a novel cerebrovascular amyloid protein , 1984 .

[42]  K. Simons,et al.  Reduced temperature prevents transfer of a membrane glycoprotein to the cell surface but does not prevent terminal glycosylation , 1983, Cell.

[43]  Y. Fujiki,et al.  Isolation of intracellular membranes by means of sodium carbonate treatment: application to endoplasmic reticulum , 1982, The Journal of cell biology.

[44]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[45]  G. Caporaso.  Chloroquine inhibits intracellular segradation but not decretion of Alzheimerβ/A4 amyloid precursor protein. , 1992 .

[46]  E. Neufeld,et al.  Lysosomal storage diseases. , 1991, Annual review of biochemistry.

[47]  N. Robakis,et al.  Effects of Neurotrophic Factors on the Secretion and Metabolism of the Alzheimer Amyloid Precursor , 1991 .

[48]  P. Stahl,et al.  Digitonin permeabilization procedures for the study of endosome acidification and function. , 1989, Methods in cell biology.

[49]  E. Kuismanen,et al.  Low temperature-induced transport blocks as tools to manipulate membrane traffic. , 1989, Methods in cell biology.

[50]  C. Cotman,et al.  Protease nexin-II, a potent antichymotrypsin, shows identity to amyloid beta-protein precursor. , 1989, Nature.

[51]  B. Greenberg,et al.  A new A4 amyloid mRNA contains a domain homologous to serine protease inhibitors , 1988 .

[52]  M. Lerman,et al.  Characterization and chromosomal localization of a cDNA encoding brain amyloid of Alzheimerʼs disease , 1988 .

[53]  S. Kornfeld,et al.  Assembly of asparagine-linked oligosaccharides. , 1985, Annual review of biochemistry.

[54]  T. Morrison,et al.  Intracellular processing of the vesicular stomatitis virus glycoprotein and the Newcastle disease virus hemagglutinin-neuraminidase glycoprotein. , 1984, Virus research.

[55]  P. Seglen.  Inhibitors of lysosomal function. , 1983, Methods in enzymology.

[56]  G H Sato,et al.  Growth of a rat neuroblastoma cell line in serum-free supplemented medium. , 1979, Proceedings of the National Academy of Sciences of the United States of America.