Regulated cleavage of the Alzheimer amyloid precursor protein: Molecular and cellular basis

Biochimie (1994) 76, 300-303

© Socirt6 franqaise de biochimie et biologie molrculaire / Elsevier, Paris

Regulated cleavage of the Aizheimer amyloid precursor protein: Molecular and cellular basis S Gandya, P Greengard b aDepartment of Neurology and Neuroscience, Cornell University Medical College, New York, NY 10021; bLaboratory of Molecular and Cellular Neuroscience, The Rockefeller University, New York, NY 10021, USA

(Received 18 February 1994; accepted 11 March 1994)

Summary - - The relative utilization of alternative processing pathways for APP can be regulated by the activation state ot certain protein phosphorylation signal transduction pathways. For example, activation of protein kinase C (PKC), or inactivation of protein phosphatases 1 and 2A, leads to a relative increase in utilization of the nonamyloidogenic, 'tx-secretase' cleavage pathway for APP processing at the expense of other pathways. The molecular and cellular basis for this regulatory event is unknown. The possible mechanisms of regulated APP cleavage include (either singly or in combination): 1) substrate (ie APP) activation; 2) substrate redistribution; 3) enzyme (ie ct-secretase) activation; or 4) enzyme redistribution. APP is a phosphoprotein; however, recent evidence from studies of the metabolism of mutant APP molecules suggests that changes in the APP cytoplasmic tail phosphorylation state may not be necessary for the phosphorylation-dependent activation of 'ct-secretase' cleavage. Further, indirect immunofluorescent studies of the subeellular distribution of APP in the absence or presence of phorbol esters (PKC activators) fail to disclose obvious phorbolinduced redistribution of APP immunoreactivity. Taken together, current data suggest that major candidate phosphorylation-state sensitive targets relevant to the molecular basis of PKC-activated processing (or 'regulated cleavage') of APP include the APP ectodomain as well as secretase enzymes and/or other components of the APP trafficking/processing apparatus. Progress in distinguishing among these possibilities is discussed. protein processing / protein phosphorylation / secretion / exocytosis / Alzheimer disease

Metabolism of the integral transmembrane Alzheimer amyloid precursor protein (APP; for review, see [1-3]) occurs via various alternative trafficking and processing pathways. The 'a-secretase' pathway involves transport of full-length, fully mature APP holoproteins to the cell surface where they are cleaved within the amyloidogenic A~ or I~A4 domain [4-6] to produce a large amino terminal fragment which is released from the cell [4, 5]. The resultant carboxyl terminal fragment [6, 7] is retained by the cell, and transported via clathrin coated vesicles [8] to lysosomes [9-11 ] for degradation. By cleaving within the AI3 (or 13A4) domain, the 'ot-secretase' pathway precludes a contribution to amyloidogenesis for those APP molecules so divided. Other pathways, notably

Abbreviations: APP, Alzheimer amyloid precursor protein;

PKC, protein kinase C; ApoE, apolipoprotein E; A~ or 13A4, amyloid 1~ peptide; GAG, glycosaminoglycan; TGN, transGolgi petwork; TGF-ct, transforming growth factor-o~.

the '[3-secretase' pathway [ 12], specify cleavage at the amino terminus of the A[3 domain, yielding a potentially amyloidogenic carboxyl termina; fragment [9, 13-17]. This and other potentially amyloidogenic fragments are then presumably further cleaved to yield the A~ peptide, which is released into culture medium and body fluids [ 18-20]. Studies of posttranslational modification and regulation of APP revealed N- and O-linked glycosylation [4, 21,221, tyrosyl sulfation [41, and phosphorylation [21, 23-26]. Cytoplasmic tail phosphorylation at Ser6~5 (numbering according to APP695) was predicted initially on the basis of in vitro studies [23, 24] and later confirmed by phosphorylation-state antibody analysis (M Oishi, personal communication). Of note, the apparent consensus sequences for this and other [25] cytoplasmic tail phosphorylation sites are widely conserved a,:ross the known APP homologues [2730]. Extracellular domain phosphorylation was discovered more recently [31-33]; however, to date, neither its precise location within the ectodomain nor it~ significance is clearly understood.

301 The initial studies of APP cytoplasmic tail phosphorylation suggested an important role for protein kinase C (PKC) [23, 24], prompting studies of the effect of phorbol esters on APP processing and traflicking, in order to investigate the possibility that PKC might modulate APP turnover. Indeed, PKC activation, or protein phosphatase inactivation (with okadaic acid), proved to be a highly reliable stimulator of APP processing via the 'tx-secretase' pathway [34-40]. These studies were extended to provide clear evidence that the 'tx-secretase' pathway was not the sole pathway for APP processing [9-12, 15-20, 41, 421, and that the phorbol effects could be mimicked by neurotransmitters or other first messengers whose receptors were linked to PKC [43-46]. The stimulation of 'tx-secretase' cleavage [34--40] is accompanied by a corresponding diminution in utilization of other APP processing pathways, including potentially amyloidogenic pathways [46--48]. The implication of regulated cleavage of APP for the pathogenesis of AD is not yet clear, although it is notable that defective PKC signalling has been frequently associated with AD [49]. In addition, with regard to possible therapeutic exploitation of regulated cleavage to diminish production of soluble A[3 peptide, it is interesting to note that signalling compounds have recently been suggested to regulate APP metabolism in humans in vivo [50]. What is the PKC substrate responsible for this regulation of APP cleavage? Since APP was an effective PKC substrate in vitro [23, 24], mutational analyses of the putative PKC target residues in the APP cytoplasmic tail were undertaken. However, PKC sensitivity was retained by APP molecules lacking the appropriate predicted PKC phosphorylation acceptor sites [33, 51]. Moreover, APP molecules lacking the entire cytoplasmic tail retained sensitivity to PKCregulated cleavage [51]. Along this line of inquiry, other investigators have provided similar evidence, and also demonstrated that membrane anchorage is required for PKC-regulated cleavage [33, 52]. Thus, unlike other examples (eg the polyimmunoglobulin receptor [53]) in which phosphorylation of the cytoplasmic tail of the substrate targets the molecule for rapid ectodomain cleavage, the APP cytoplasmic tail plays little, if any, role in its regulated cleavage. Since 't~-secretase' cleavage of APP occurs in large part at the cell-surface [54], a possible cellular basis for regulated cleavage is that PKC activation leads to redistribution of APP to the plasma membrane. Light microscopic indirect immunofluorescence analysis has not supported this model [22], but more sophisticated approaches are underway. Of note, as proposed by Luini and De Matteis [55], PKC may promote vesicular traffic from the trans-Golgi network (TGN) to the plasma membrane, as evidenced in their demon-

stration that PKC stimulates glycosaminoglycan (GAG) secretion [56]. This stimulated GAG release is associated with redistribution of non-clathrin coatomers from the cytosol to the Golgi region of the cell, perhaps inducing or participating in an enhancement of vesicle budding or flux at the TGN [56]. The 'o~-secretase' is also a possible target for PKC: such a regulatory mechanism has not previously been described for a proteinase but has been strongly implicated in the case of the proTGF-tx processing proteinase [57]. Experiments are underway for testing the possible similarity of regulated proTGF-o~ cleavage to regulated APP cleavage. To date, neither APP o~-secretase nor the proTGF-~ processing proteinase has been characterized at the molecular level, and it is this information which will be required in order to definitively establish that the activity of either proteinase is indeed directly regulated by the state of phosphorylation of the enzyme. The importance of APP processing for the pathogenesis of Alzheimer disease (AD) is not yet clear. Although some early-onset FAD kindreds provide linkage of clinical dementia to APP mutations which generate excess AI3/I3A4 peptide [58, 59], the most common APP mutation lacks an obvious proamyloidogenic processing phenotype [59], and recent linkage of late-onset and sporadic AD to apolipoprotein E (ApoE; [60--62]) raises a number of crucial issues about the role of APP processing and amyloid in AD: is AI3/I3A4 aggregation state a more important determinant of AD than is the level of AI3/~A4 or the number of AI3/I3A4 deposits? Does ApoE exert its AD-risk-factor effects strictly via promoting amyloidogenesis, or are there more important activities of ApoE relating to its roles in neuronal plasticity, in the injury response [63], or in maintaining the cytoskeleton [64], which, in turn, are more crucial for determining clinical dementia in AD than is the 'burden' of cerebral amyloid plaques? The tools for addressing these issues are now in place, and the coming years should yield clarification of these points. It is anticipated that particularly valuable elucidation of these issues will emerge from the eventual discovery of the major early-onset FAD gene on chromosome 14 [65].

Acknowledgment This w o r k w a s supported by U S P H S AG09464, A G I 0 4 9 1 and AG11508.

References I Gandy SE, Greengard P (1992) Amyioidogenesis in Alzheimer disease: some possible therapeutic opportunities. Trends Pharmacol Sci 13, 108-113 2 Gandy S, Caporaso G, Buxbaum J, Frangione B, Greengard P (1994) APP proc~ssing, AI3 amyloidogenesis, and the pathogenesis of Alzheimer's disease. NeurobioiAging 15, 253-256

302 3 Gandy S. Greengard P (1994) Processing of A~-amyloid precursor protein: Cell biology, regulation, and role in Alzheimer disease, lnI Rev Neurobiol. in press 4 Weidemann A. Konig G. Bunke D. Fischer P. Salbaum JM. Masters CL. Beyreuther K (1989) Identification. biogenesis and Iocal;zation of precursors of Alzheimer's disease A4 amyloid protein. Cell 57. ! 15-126 5 Stscdia SS. Koo Eli. Beyreuther K. Unterheck A. Price DL (1990) Evidence that I$-amyloid r~.~eia in Alzheimer's disease is not derived by normal processing. Science 248.492-495 6 Esch FS. Keim PS. Beattie EC. Blacher RW. Culwell AR. Oltersdorf T. Me(lure D. Ward PJ (1990) Cleavage of amyloid peptide during constitutive processing of its precursor. Sciem'e 248. ! 122-1124 7 Ramabhadran TV. Gandy SE. Ghiso J. Czeroik A. Ferris D. Bhasin R. Goldgaber D. Frangiune B. Greengard P (1993) Proteolytic processing of human amyloid [$ protein precursor in insect cells: major carboxyl terminal fragment is identical to its human counterpart. J Biol (hem 268. 2009-2012 8 Nordstedt C. Caporaso GL. Thyberg J. Gandy SE. Greengard P (1993) Identification of the Alzheimer [$A4 amyloid precursor protein in clathrin-coated vesicles purified from PC 12 cells. $ Biol {?hem 268. 608-612 9 Golde T. Estus S. Younkin L. Selkoe D. Younkin S (1992) Processing of the amyloid protein precursor to potentially amyloidogenic derivatives. Science "J~5. 728--739 I0 ~-~,assC. Koo EH. Mellon A. Hung AY. Selkoe DJ (1992) Targeting of cellsurface [$-amyloid precursor protein to lysosomes: Alternative processing into amyloid.bearing fragments. Nature 357. 500-503 I! Caporaso GL. Gandy SE. Buxbaum JD. Greengard P (1992) Chloroquine inhibits intracellular degradation but not secretion of Alzheimer ~A4 amyloid precursor protein. Proc Nail Acad Sci USA 89. 2252-2256 12 Seubert P. Oltersdorf T. Lee MG. Barbour R. Blomquist C. Davis DL. Bryant K. Galasko D. Thai 13. Fritz L. Lieberbta'g !. Schenk D (1993) Secretion of [$-amyloid precursor protein cleaved at the amino terminus of the ~amyloid peptide. Nature 361. 260-263 13 Nordsledt C. Gandy S. Alafuzoff 1. Caporaso G. lveffeldt K. Grebb J. Winblad B. Greengard P (1991) Alzheimer [~A4-amyloid precursor protein in human brain: Aging-associated increases in holoprotein and in a proteolytic fragment. Proc Nail Acad Sci USA ~:~.8910-8914 14 Gandy SE. Bhasin R. Ramabhadran TV. Koo EH. Price DL. Goldgaber D. Greengard P (1992) Alzheimer [$A4 amyloid precursor protein: Evidence for putative amyloidogenic fragment. J Ncurm'hem 58. 383-386 15 Estus S. Golde T. Kanishita T. Blades D. Lowery D. Eisen M. Usiak M. Qu X. Tabira 1". Greenberg B. Younkin S (1992) Potentially amyloidogenic, carboxyl-terminal derivatives of the amyloid prote~n precursor. Science 255. 726--728 16 Tama~Ka A. Kalaria R. Lieherburg i. Selkoe D (1992) Identification of a stable fragment of the Alzheimer amyloid precursor containing the IS-protein in brain microvessels. Proc Nail Acad Sci USA 89. 1345-1349 17 Kr~ops J. Lieberburg !. Sinha S (1992) Evidence for a nonsecretory, acidic degradation pathway for amyloid precursor protein in 293 cells: Identification of a novel. 22-kDa. heta-peptide containing intermediate. J Biol Chem 267. 16022-16024 18 Haass C. Schlossmacher MG. Hung AY. Vigo-Peifrey C. Mellon A. Ostaszewski BC. Lieberburg I. Koo EH. Schenk D. Teplow D. Selkoe DJ (1992) Amyloid ~peptide is produced by cultured cells during normal metabolism. Nature 359. 322-325 19 Seuben P. Vigo-Pelfrey C. Esch IF. Lee M. Dovey H. Davis D. Sinha S. Schlossmacher M. Whaley J. Swindlehurst C. McCormack R. Wolfert R. Selkoe D. Lieherburg !. Schenk D (1992) Isolation and quantification of soluble Alzheimer's 13-peptide from biological fluids. Nature 359. 325-327 20 Shoji M. Golde TE. Ghiso J. Cheung "IT. Estus S. ShatTer LM. Cai X. McKay DM. Tintner R. Frangione B. Younkin SG (1992) Production of the Alzheimet amyloid ~ protein by normal proteolytic processing. Science 258. 126--129 21 Oltersdorf TL. Ward PJ. Henriksson T. Beattie EC. Neve RL. Lieberburg IL. Fritz LC (1990) The Alzheimer amyloid precursor protein: Identification of a stable intermediate in the biosynthetic/degradative pathway. J Biol Chem 265.4492--4497 22 Caporaso G. Takei K. Gandy S. Matteoli M. Mandigl O. Greengard P. de Camilli P (1904) Morphologic and biochemical analysis of the intracellular traf~cking of the Alzheimer [~/A4 amyloid precursor protein. J Neurosci 14. 3122-3138

23 Gandy S. Czeroik AJ. Greengard P (1988) Phosphorylation of Alzheimer disease amyloid precursor peptide by protein kinase C and Ca2÷/calmodulin dependent protein kinase II. Proc Nail Acad Sci USA 85. 6218--6221 24 Suzuki T. Nairo AC. Gandy SE. Greengard P (1992) Phosphorylation of Alzheimer amyloid precursor protein by protein kinase C. Neuroscience 48. 755-76 ! 25 Suzuki T. Oishi M. Marshak DR. Czemik AJ. Nairn AC. Greengard P (1993) Cell cycle-dependent regulation of the phosphorylation and metabolism of the Alzheimer amyloid precursor protein. EMBO l i 3. ! 114-1122 26 Johnson G. Refolo LM. Merril CR. Wallace W (1993) Altered expression and phosphorylation of amyloid precursor protein in heat shocked neuronal PCI2 cells. Moi Brain Res 19.140-148 27 Rosen DR. Martin-Morris L. Luo L. White K (1989) A Drosophila gene encoding a protein resembling the human [$-amyioid protein precursor. Proc Nail Acad Sci USA 86. 2478-2482 28 Okado H. Okamoto H (1992) A Xenopus homologue of the human beta-amyloid precursor protein: developmental regulation of its gene expression. Biochem Biophys Res Commun 189. 1561-1568 29 Wasco W. Bupp K. Magendantz M. Gusella JF. Tanzi RE. Solomon F (1992) Identification of a mouse brain cDNA that encodes a protein related to the Alzheimer disease-associated amyloid [3 protein precursor. Proc Nail Acad Sci USA 89. 10758-10762 30 Wasco W. Gurubhagavatula S. Paradis Md. Romano DM. Sisodia SS. Hyman BT. Neve RL. Tanzi RE (1993) Isolation and characterization of APLP2 encoding a homologue of the Aizheimer's associated amyloid [5 protein precursor. Proc Nail Acad Sci USA 5.95-100 31 Knops J. Gandy S. Greengard P. Lieberburg 1. Sinha S (1993) Serine phosphorylation of the secreted extracellular domain of APP. Biochem Biophys Res Commun 197. 380-385 32 Gabuzda D. Busciglio J. Yankner BA (1993) Inhibition of [l-amyloid production by activation of protein kinase C. J Neurochem 61. 2326--2329 33 Hung AY. Selkoe DJ (1994) Selective ectodomain pbosphorylation and regulated cleavage of ~amyloid precursor protein. EMBO l 13. 534-542 34 Buxbaum JD. Gandy SE. Cicchetti P. Ehrlich ME. Czeroik AJ. Fracasso RP. Ramabhadran TV. Unterbeck AJ. Greengard P (1990) Processing of Alzheimer [~A4 amyloid precursor protein: Modulation by agents that regulate protein phosphorylation. Proc Nail Acad Sci USA 87. 6003--6006 35 Caporaso G. Gandy S. Buxbaum J. Ramabhadran T. Greengard P (1992) Protein phosphorylation regulates secretion of Alzheimer ~A4 amyloid precursor protein. Proc Nail Acad Sci USA 89. 3055-3059 36 Gillespie SL. Golde "rE. Younkin SG (1992) Secretory processing of the Alzhein~er amyioid [3A4 protein precursor is increased by protein phosphory.'~tion. Biochem Biophys Res Commun 187. 1285-1290 37 Sinha S. Lieherburg i (1992) Normal mtetabolism of the Alzheimer amyloid precursor protein. Neurodegeneralion 1.169-175 38 Loftier J. Huber G (1993) Modulation of [$-amyioid precursor protein secretion in differentiated and non-differentiated cells. Biochem Biophys Res Commun 195.97-103 39 Slack BE0 Nits~:h RM. Livneh E. Kunz GM. Breu J. Eldar H. Wurtman RJ (1993) Regulation by phorbol esters of amyloid precursor protein release from Swiss 3T3 fibroblasts overexpressing protein kinase C-alpha. J Biol Chem 268. 21097-21101 40 Bieger S. Klafki HW. Unsicker K (1993) Synthesis and release of the p-amyIoid precursor protein by bovine chromaffin cells. Neurosci Leu 162. 173175 41 Gandy SE. Caporaso GL. Ramabhadran TV. Buxbaum JD. Suzuki T. Nordstedt C. lverfeldt K. Czemik AJ. Nairo AC. Greengard P (1992) Characterization of alternative routes for processing of Alzheimer [5A4 amyloid precursor protein: Differential effects of phorboi esters and chloroquine. Ann NY Acad Sci 674. 203-217 42 Gandy S. Caporaso G. Buxbaum J. da Cruz e Silva O. Iveffeldt K. Nordstedt C. Suzuki T. Czeroik A. Nairo A. Greengard P (1993) Protein phosphorylation regulates relative utilization of processing pathways for Alzheimer [~A4 amyloid precursor protein. Ann NY Acad Sci 695. 117-121 43 Buxbaum JD. Oishi M. Chen HI. Pinkas-Kramarski R. Jaffe EA. Gaudy SE. Greengard P (1992) Cholinergic agonists and interleukin ! regulate processing and secretion of the Alzheimer [$A4 amyloid protein precursor. Proc Nail Acad Sci USA 89. 10075-10078

303 44 Nitsch RM, Slack BE, Wurtman Pal, Growdon JH (1992) Release of Alzheimer amyioid precursor derivatives stimulated by activation of muscarinic acetyicholine receptors. Science 258, 304-307 45 Emmerling MR, Moore CJ, Doyle PD, Carroll RT, Davis RE (1993) Phospholipase A2 activation influences the processing and secretion of the amyIoid precursor protein. Biochem Biophys Res Commun 197, 292-297 46 Hung AY, Haass C, Nitsch RM, Qiu WQ, Citron M, Wurtman ILl, Growdon JH, Selkoe DJ (1993) Activation of protein kinase C inhibits cellular production of the amyloid [s-protein. J Biol Chem 268, 22959-22962 47 Buxbaum JD, Koo EH, Greengard P (1993) Protein phosphorylation inhibits production of Alzheimer amyloi0 [SA4 peptide. Proc Natl Acad Sci USA 90, 9195-9198 48 Fukushima D, Konishi M, Mamyama K, Miyamoto T, Ishiura S, Suzuki K (1993) Activation of the secretory pathway leads to a decrease in the intracellular amyloidogenic fragments genera!ed from the amyloid protein precursor. Biochem Biophys Res Commun 194, 202-207 49 Huang HM, Martins R, Gandy S, Blas~ J, Gibson G (1994) The use of cultured fibroblasts in elucidating the pathophysiology and diagnosis of Aizheimer's disease. Ann NY Acad Sci, in press 50 Clarke NA, Webster MT, Francis PT, Procter AW, Hodgkiss AD, Bowen DM (1993) [s-Amyloid precursor protein-like immunoreactivity can be altered in humans by drugs affecting neurotransmitter function. Neurodegeneration 2, 243-248 51 da Cruz e Silva O, Iveffeldt K, Oitersdorf T, Sinha S, Lieberburg I, Ramabhadran T, Suzuki T, Sisodia S, Gandy S, Greengard P (1993) Regulated cleavage of Alzheimer [s-amyloid precursor protein in the absence of the cytoplasmic tail. Neuroscience 57, 873-877 52 Demaerschlack 1, Delvaux A, Octave JN (1993) Activation of protein kinase C increases the extracellular release of the transmembrane amyloid protein precursr of Alzheimer's disease. Biochim Biophys Acta 118 i, 214--218 53 Casanova JE, Breitfield PP, Ross SA, Mostov KE (1990) Phosphorylation of the polyimmunoglobulin receptor required for its efficient transcytosis. Science 248, 742-745 54 Sisodia SS (1992) [s-Amyloid precursor protein cleavage by a membranebound protease. Proc Natl Acad Sci USA 89, 6075--6079 55 Luini A, De Matteis MA (1993) Receptor-mediated regulation of constitutive secretion. Trends Cell Biol 3, 290-292

56 De Matteis MA, San~ini G, Kahn RA, Di Tullio G, Luini A (1993) Receptor and protein kinase C-mediated regulation of ARF binding to the Golgi complex. Nature 364, 818-821 57 Bosenberg M, PandieUa A, Massagu¢ J (1993) Activated release of memhrane-anchored TGF-a in the absence of cytosol. J Cell Biol 122, 95-101 58 Citron M, OItersdoff T, Haass C, McConlogue L, Hung AY, Seubert P, VigoPalfrey C, Lieberburg I, Selkoe DJ (1992) Mutation of the [s-amyloid precursor protein in familial Alzheimer's disease causes increased [s-protein production. Nature 360, 672-674 59 Cai XD, Golde TE, Younkin SG (1993) Release of excess amyloid IS-protein from mutant amyloid [s-protein precursor. Science 259, 514-516 60 Namba Y, Tomonaga M, Kawasaki H, Otomo E, Ikeda K (1991) Apolipoprorein E immunoreactivity in cerebral amyloid deposits and neurofibriilary angles in Alzheimer's disease and kuru plaque amyloid in Creutzfeldt-Jakob disease. Brain Res 54 l, 163-166 61 Wisniewski T, Frangione B (1992) Apolipoprotein E: A pathological chaperone protein in patients with cerebral and systemic amyloid. Neurosci Lett 135, 235-238 62 Saunders AM, Strittmatter WJ, Schraechel D, St George-Hyslop PH, PericakVance MA, Joos SH, Rosi BL, Gusella JF, Crapper-MacLachlan DR, Alberts MJ, Hulette C, Crain B, Goldgaber D, Roses AD (1993) Association of ,polipoprotein E allele ~4 with late-onset familial and sporadic Alzbeimer's disease. Neurology 43, 1467-1472 63 Ignatius MJ, Gebicke-Harter PJ, Skene JHR Schilling JW, Weisgraber KH, Mahley RW, Shooter EM (1986) Expression of apolipoprotein E during nerve degeneration and regeneration. Proc Natl Acad Sci USA 83, 1125-1129 64 Strittmatter WJ, Weisgraber KH, Goedert M, Saunders AM, Huang D, Corder EH, Dong LM, Jakes R, Alberts MI, Gilbert JR, Hart SH, Hulette C, Einstein G, Schmechei DE, Pericak-Vance MA, Roses AD (1994) Hypothesis: Microtubule instability and paired helical filament formation :~n 1he Alziteimer disease brain as a function of apolipoprotein E genotype. Exp Neurol 125, 163-171 65 Scheilenberg GD, Bird TD, Wijsman EM, Orr HT, Anderson L, Nemens E. White JA, Bonnycastle L, Weber JL, AIonso ME, Potter H, Heston LL, Martin GM (1992) Genetic linkage evidence for a familial Alzfieimer's disease locus on chromosome 14. Science 258,668-671