29 Apr 2016

Intracellular trafficking of the amyloid precursor protein (APP) is thought to go awry in Alzheimer’s disease, but most studies focus on later stages of the process, when this membrane protein recycles through endosomes. Could earlier phases of APP transit also influence disease? A pair of papers published online April 25 in the Proceedings of the National Academy of Sciences suggest just that. Scientists led by Marc Flajolet and Paul Greengard at The Rockefeller University, New York, report that preventing the retrograde movement of nascent APP from the Golgi to the endoplasmic reticulum (ER) reduces the amount of mature protein at the cell surface and the production of Aβ. In AD mouse models, blocking this retrograde transit reduced plaque load and rescued memory deficits in transgenic mice. In AD cohorts, genetic variants among eight proteins that regulate this retrograde transport segregated with disease. 

“These two papers report a comprehensive investigation into how perturbation in early secretory trafficking could have a profound impact on the amyloidogenic processing of APP in vitro and in vivo,” noted Gopal Thinakaran, University of Chicago, to Alzforum. “While there is a large body of literature implicating endocytic trafficking in AD pathogenesis, these papers shine the light on the significance of bidirectional trafficking between the ER and Golgi, and illustrate that the biosynthetic trafficking pathway might be equally important.”

After APP is synthesized and folded in the endoplasmic reticulum, it proceeds, as do many other membrane-bound proteins, through the Golgi, where it gets modified post-translationally. From there, proteins make the journey to the plasma membrane. Any that have not folded properly travel back to the ER to be refolded, in what is known as retrograde transport. Coat protein complex I (COPI), a seven-subunit protein, lines specialized vesicles and helps orchestrate this retrograde travel. Flajolet and colleagues wondered if the function of this complex might affect how much Aβ is made down the road.

To find out, Karima Bettayeb, who is first author on both papers, tested what would happen if she reduced expression of individual COPI subunits with small interfering RNAs. She did this in mouse neuroblastoma cells that overexpressed APP-695 (N2a-695). She found that knocking down several of the subunits was too toxic for cells, but she could safely reduce one—the δ subunit. Without δ-COP, these cells produced less Aβ40 and Aβ42; if it was overexpressed, the cell produced more. Tagging APP with a green florescent protein revealed that reduced δ-COP left more APP in the Golgi, and that the two proteins co-localized there. According to biotinylation experiments, less APP reached the cell surface.

Why would reduced retrograde transport lead to less Aβ? The effect seemed to depend in part on N-glycosylation of APP—the attachment of sugar molecules to amino side chains in the protein. This occurs in the ER. When the researchers removed APP’s two N-glycosylation sites, then deleting δ-COP made no difference to Aβ output. The results imply that N-glycosylation promotes Aβ production and that blocking retrograde trafficking reduces this post-translational modification and Aβ cleavage. N-glycosylation may keep APP from being flagged for degradation once it reaches the plasma membrane, Flajolet suggested. Without glycosylation, less APP is endocytosed and so less Aβ produced, he told Alzforum. Scientists believe that most Aβ production occurs during endocytosis, when APP and β-secretase cross paths in acidic lysosomes.

What does deleting δ-COP do in vivo? In their second paper, Bettayeb and colleagues examined this in animal models. They crossed the APPSwe/PS1deltaE9 mouse with nur17 mice, which carry a T-to-C missense mutation in δ-COP that partially disrupts intracellular trafficking (Xu et al., 2010). Compared to control APP/PS1 mice, the crosses had less Aβ40 and Aβ42 in the hippocampus, and fewer plaques in both the hippocampus and piriform cortex. These mice also explored an unfamiliar object longer than a familiar one in the novel object recognition test, indicating that they remember which object they previously saw as well as wild-type animals do.

Could COPI trafficking contribute to AD pathogenesis? The researchers wondered if any genetic variants in the COPI genes might be risk factors for AD. Meta-analysis of six different AD cohorts revealed 12 SNPs among genes for five subunits of the COPI complex that trended toward statistical significance for increasing AD risk. To get more detailed data, the researchers examined whole genome sequence data from 941 affected and 404 unaffected people from the National Institute of Mental Health (NIMH) Genetics Initiative Alzheimer’s Disease Study. They uncovered 24 variants among nine COPI genes that segregated with AD.

"Taken together, the findings reported in these two papers strongly favor a role for the COPI complex in Alzheimer’s,” wrote Lindsay Farrer, Boston University School of Medicine, to Alzforum. “These studies also confirm the important role of cellular trafficking and recycling mechanisms in AD.” Other scientists noted, however, that none of the COPI loci turned up in the International Genomics of Alzheimer’s Project (IGAP) GWAS meta-analysis of more than 74,000 patients and controls (see Lambert et al., 2013). 

Flajolet said he’s unsure if these findings will have therapeutic implications, as total knockout of the COPI complex is lethal. Any therapeutic approach should only dampen COPI trafficking, not block it.—Gwyneth Dickey Zakaib

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References

Research Models Citations

1. APPswe/PSEN1dE9 (line 85)

Paper Citations

1. Xu X, Kedlaya R, Higuchi H, Ikeda S, Justice MJ, Setaluri V, Ikeda A. Mutation in archain 1, a subunit of COPI coatomer complex, causes diluted coat color and Purkinje cell degeneration. PLoS Genet. 2010 May;6(5):e1000956. PubMed.
2. Lambert JC, Ibrahim-Verbaas CA, Harold D, Naj AC, Sims R, Bellenguez C, DeStafano AL, Bis JC, Beecham GW, Grenier-Boley B, Russo G, Thorton-Wells TA, Jones N, Smith AV, Chouraki V, Thomas C, Ikram MA, Zelenika D, Vardarajan BN, Kamatani Y, Lin CF, Gerrish A, Schmidt H, Kunkle B, Dunstan ML, Ruiz A, Bihoreau MT, Choi SH, Reitz C, Pasquier F, Cruchaga C, Craig D, Amin N, Berr C, Lopez OL, De Jager PL, Deramecourt V, Johnston JA, Evans D, Lovestone S, Letenneur L, Morón FJ, Rubinsztein DC, Eiriksdottir G, Sleegers K, Goate AM, Fiévet N, Huentelman MW, Gill M, Brown K, Kamboh MI, Keller L, Barberger-Gateau P, McGuiness B, Larson EB, Green R, Myers AJ, Dufouil C, Todd S, Wallon D, Love S, Rogaeva E, Gallacher J, St George-Hyslop P, Clarimon J, Lleo A, Bayer A, Tsuang DW, Yu L, Tsolaki M, Bossù P, Spalletta G, Proitsi P, Collinge J, Sorbi S, Sanchez-Garcia F, Fox NC, Hardy J, Deniz Naranjo MC, Bosco P, Clarke R, Brayne C, Galimberti D, Mancuso M, Matthews F, European Alzheimer's Disease Initiative (EADI), Genetic and Environmental Risk in Alzheimer's Disease, Alzheimer's Disease Genetic Consortium, Cohorts for Heart and Aging Research in Genomic Epidemiology, Moebus S, Mecocci P, Del Zompo M, Maier W, Hampel H, Pilotto A, Bullido M, Panza F, Caffarra P, Nacmias B, Gilbert JR, Mayhaus M, Lannefelt L, Hakonarson H, Pichler S, Carrasquillo MM, Ingelsson M, Beekly D, Alvarez V, Zou F, Valladares O, Younkin SG, Coto E, Hamilton-Nelson KL, Gu W, Razquin C, Pastor P, Mateo I, Owen MJ, Faber KM, Jonsson PV, Combarros O, O'Donovan MC, Cantwell LB, Soininen H, Blacker D, Mead S, Mosley TH Jr, Bennett DA, Harris TB, Fratiglioni L, Holmes C, de Bruijn RF, Passmore P, Montine TJ, Bettens K, Rotter JI, Brice A, Morgan K, Foroud TM, Kukull WA, Hannequin D, Powell JF, Nalls MA, Ritchie K, Lunetta KL, Kauwe JS, Boerwinkle E, Riemenschneider M, Boada M, Hiltuenen M, Martin ER, Schmidt R, Rujescu D, Wang LS, Dartigues JF, Mayeux R, Tzourio C, Hofman A, Nöthen MM, Graff C, Psaty BM, Jones L, Haines JL, Holmans PA, Lathrop M, Pericak-Vance MA, Launer LJ, Farrer LA, van Duijn CM, Van Broeckhoven C, Moskvina V, Seshadri S, Williams J, Schellenberg GD, Amouyel P, Wang J, Uitterlinden AG, Rivadeneira F, Koudstgaal PJ, Longstreth WT Jr, Becker JT, Kuller LH, Lumley T, Rice K, Garcia M, Aspelund T, Marksteiner JJ, Dal-Bianco P, Töglhofer AM, Freudenberger P, Ransmayr G, Benke T, Toeglhofer AM, Bressler J, Breteler MM, Fornage M, Hernández I, Rosende Roca M, Ana Mauleón M, Alegrat M, Ramírez-Lorca R, González-Perez A, Chapman J, Stretton A, Morgan A, Kehoe PG, Medway C, Lord J, Turton J, Hooper NM, Vardy E, Warren JD, Schott JM, Uphill J, Ryan N, Rossor M, Ben-Shlomo Y, Makrina D, Gkatzima O, Lupton M, Koutroumani M, Avramidou D, Germanou A, Jessen F, Riedel-Heller S, Dichgans M, Heun R, Kölsch H, Schürmann B, Herold C, Lacour A, Drichel D, Hoffman P, Kornhuber J, Gu W, Feulner T, van den Bussche H, Lawlor B, Lynch A, Mann D, Smith AD, Warden D, Wilcock G, Heuser I, Wiltgang J, Frölich L, Hüll M, Mayo K, Livingston G, Bass NJ, Gurling H, McQuillin A, Gwilliam R, Deloukas P, Al-Chalabi A, Shaw CE, Singleton AB, Guerreiro R, Jöckel KH, Klopp N, Wichmann HE, Dickson DW, Graff-Radford NR, Ma L, Bisceglio G, Fisher E, Warner N, Pickering-Brown S. Meta-analysis of 74,046 individuals identifies 11 new susceptibility loci for Alzheimer's disease. Nat Genet. 2013 Dec;45(12):1452-8. Epub 2013 Oct 27 PubMed.

Further Reading

Papers

1. Selivanova A, Winblad B, Farmery MR, Dantuma NP, Ankarcrona M. COPI-mediated retrograde transport is required for efficient gamma-secretase cleavage of the amyloid precursor protein. Biochem Biophys Res Commun. 2006 Nov 10;350(1):220-6. PubMed.

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