Assembly, trafficking and function of gamma-secretase

gamma-Secretase catalyzes the final cleavage of the beta-amyloid precursor protein to generate amyloid-beta peptide, the principal component of amyloid plaques in the brains of patients suffering from Alzheimer's disease. Here, we review the identification of gamma-secretase as a protease complex and its assembly and trafficking to its site(s) of cellular function. In reconstitution experiments, gamma-secretase was found to be composed of four integral membrane proteins, presenilin (PS), nicastrin (NCT), PEN-2 and APH-1 that are essential and sufficient for gamma-secretase activity. PS, which serves as a catalytic subunit of gamma-secretase, was identified as a prototypic member of novel aspartyl proteases of the GxGD type. In human cells, gamma-secretase could be further defined as a heterogeneous activity consisting of distinct complexes that are composed of PS1 or PS2 and APH-1a or APH-1b homologues together with NCT and PEN-2. Using green fluorescent protein as a reporter we localized PS and gamma-secretase activity at the plasma membrane and endosomes. Investigation of gamma-secretase complex assembly in knockdown and knockout cells of the individual subunits allowed us to develop a model of complex assembly in which NCT and APH-1 first stabilize PS before PEN-2 assembles as the last component. Furthermore, we could map domains in PS and PEN-2 that govern assembly and trafficking of the complex. Finally, Rer1 was identified as a PEN-2-binding protein that serves a role as an auxiliary factor for gamma-secretase complex assembly. Copyright (c) 2006 S. Karger AG, Basel

Signal peptide peptidases and gamma-secretase: Cousins of the same protease family?

Signal peptide peptidase (SPIP) is an unusual aspartyl protease, which mediates clearance of signal peptides by proteolysis within the endoplasmic reticulum (ER). Like presenilins, which provide the proteolytically active subunit of the,gamma-secretase complex, SPP contains a conserved GxGD motif in its C-terminal domain which is critical for its activity. While SPIP is known to be an aspartyl protease of the GxGD type, several presenilin homologues/SPP-like proteins (PSHs/ SPPL) of unknown function have been identified by database searches. In contrast to SPP and SPPL3, which are both restricted to the endoplasmic reticulum, SPPL2b is targeted through the secretory pathway to endosomes/lysosomes. As suggested by the differential subcellular localization of SPPL2b and SPPL3 distinct phenotypes were found upon antisense gripNA-mediated knockdown in zebrafish. spp and sppl3 knockdowns in zebrafish result in cell death within the central nervous system, whereas reduction of sppl2b expression causes erythrocyte accumulation in an enlarged caudal vein. Moreover, expression of D/A mutants of the putative C-terminal active sites of spp, sppl2, and spp13 produced phenocopies of the respective knockdown phenotypes. These data suggest that all investigated PSHs/SPPLs are members of the novel family of GxGD aspartyl proteases. More recently, it was shown that SPPL2b utilizes multiple intramembrane cleavages to liberate the TNF(x intracellular domain into the cytosol and to release the C-terminal counterpart into the lumen. These findings suggest common principles of intramembrane proteolysis by GxGD type aspartyl proteases. In this article,we will review the similarities of SPPs and gamma-secretase based on recent findings by us and others

Decoupling of optoelectronic properties from morphological changes in sodium treated kesterite thin film solar cells

Sodium is typically used during the synthesis of kesterite thin films to enhance the performance of solar cells. As sodium tends to affect grain growth and morphology, it is difficult to analyse solely the electronic effects of sodium as dopant. To decouple the structural and electronic effects from each other, two processes were designed in this work to successfully incorporate sodium into a vacuum-processed Cu2ZnSnSe4absorber without changing the morphology. A thin layer of NaF is deposited before precursor deposition (Pre-NaF) or after absorber synthesis to undergo a post deposition treatment (NaF-PDT). While composition and distribution of matrix elements remain unchanged, the sodium concentration is increased upon sodium treatment up to 140 ppm as measured by inductively coupled plasma mass spectrometry. X-ray photoelectron spectroscopy showed that the surface composition was not altered. Within its detection limit, sodium was not present at the absorber surface. For a Pre-NaF sample measured with atom probe tomography a sodium concentration of 30 ppm was measured in a grain, suggesting that sodium might segregate at grain boundaries. The additional sodium content in the film leads to an increased acceptor concentration, which results in improved open-circuit voltage and fill factor.Financial support from the Swiss National Science Foundation (SNF) in the network of the Indo-Swiss Joint Research Programme (ISJRP) [IZLIZ2_157140/1] is gratefully acknowledged. T. Schwarz is grateful for the support of the German Research Foundation (DFG) [Contract GA 2450/1-1]. R. Caballero acknowledges financial support from Spanish MINECO within the Ramón y Cajal program [RYC-2011-08521], MINECO project WINCOST [ENE2016-80788-C5-2-R] and from Spanish Ministry of Education, Culture and Sport within the José Castillejo program [CAS 15/00070

A study of the selective hydroconversion of biocarboxylic acids to bioalcohols over novel indium-nickel/zeolite catalysts using octanoic acid as model reactant

Octanoic acid (OA) was hydrotreated in a flow-through reactor at 21 bar total pressure and 240-340 °C over supported metal catalysts prepared from Ni-zeolites (A, X, P) by indium modification. The Ni-zeolites were activated first in H2 flow at 21 bar and 450 °C. While a fraction of the nickel got fully reduced, the zeolite structure became partially destructed. However, some nickel cations remained unreduced, therefore a large fraction of the crystalline zeolite structure was retained. The indium modification of the reduced Ni-zeolites generated bimetallic NiIn/Ni,H-zeolite catalysts having higher stability, hydroconversion activity, octanol selectivity, and lower hydrodecarbonylation activity than the parent partially destructed Ni-zeolite and the Adkins-type catalysts, commercially applied for the conversion of fatty acids to alcohols. © 2013 Akadémiai Kiadó, Budapest, Hungary

Generic Mechanism of Emergence of Amyloid Protofilaments from Disordered Oligomeric aggregates

The presence of oligomeric aggregates, which is often observed during the process of amyloid formation, has recently attracted much attention since it has been associated with neurodegenerative conditions such as Alzheimer's and Parkinson's diseases. We provide a description of a sequence-indepedent mechanism by which polypeptide chains aggregate by forming metastable oligomeric intermediate states prior to converting into fibrillar structures. Our results illustrate how the formation of ordered arrays of hydrogen bonds drives the formation of beta-sheets within the disordered oligomeric aggregates that form early under the effect of hydrophobic forces. Initially individual beta-sheets form with random orientations, which subsequently tend to align into protofilaments as their lengths increases. Our results suggest that amyloid aggregation represents an example of the Ostwald step rule of first order phase transitions by showing that ordered cross-beta structures emerge preferentially from disordered compact dynamical intermediate assemblies.Comment: 14 pages, 4 figure

Chronic y-secretase inhibition reduces amyloid plaque-associated instability of pre- and postsynaptic structures

The loss of synapses is a strong histological correlate of the cognitive decline in Alzheimer’s disease (AD). Amyloid bpeptide (Ab), a cleavage product of the amyloid precursor protein (APP), exerts detrimental effects on synapses, a process thought to be causally related to the cognitive deficits in AD. Here, we used in vivo two-photon microscopy to characterize the dynamics of axonal boutons and dendritic spines in APP/Presenilin 1 (APPswe/PS1L166P)–green fluorescent protein (GFP) transgenic mice. Time-lapse imaging over 4 weeks revealed a pronounced, concerted instability of pre- and postsynaptic structures within the vicinity of amyloid plaques. Treatment with a novel sulfonamide-type g-secretase inhibitor (GSI) attenuated the formation and growth of new plaques and, most importantly, led to a normalization of the enhanced dynamics of synaptic structures close to plaques. GSI treatment did neither affect spines and boutons distant from plaques in amyloid precursor protein/presenilin 1-GFP (APPPS1-GFP) nor those in GFP-control mice, suggesting no obvious neuropathological side effects of the drug

FET proteins TAF15 and EWS are selective markers that distinguish FTLD with FUS pathology from amyotrophic lateral sclerosis with FUS mutations

Accumulation of the DNA/RNA binding protein fused in sarcoma as cytoplasmic inclusions in neurons and glial cells is the pathological hallmark of all patients with amyotrophic lateral sclerosis with mutations in FUS as well as in several subtypes of frontotemporal lobar degeneration, which are not associated with FUS mutations. The mechanisms leading to inclusion formation and fused in sarcoma-associated neurodegeneration are only poorly understood. Because fused in sarcoma belongs to a family of proteins known as FET, which also includes Ewing's sarcoma and TATA-binding protein-associated factor 15, we investigated the potential involvement of these other FET protein family members in the pathogenesis of fused in sarcoma proteinopathies. Immunohistochemical analysis of FET proteins revealed a striking difference among the various conditions, with pathology in amyotrophic lateral sclerosis with FUS mutations being labelled exclusively for fused in sarcoma, whereas fused in sarcoma-positive inclusions in subtypes of frontotemporal lobar degeneration also consistently immunostained for TATA-binding protein-associated factor 15 and variably for Ewing's sarcoma. Immunoblot analysis of proteins extracted from post-mortem tissue of frontotemporal lobar degeneration with fused in sarcoma pathology demonstrated a relative shift of all FET proteins towards insoluble protein fractions, while genetic analysis of the TATA-binding protein-associated factor 15 and Ewing's sarcoma gene did not identify any pathogenic variants. Cell culture experiments replicated the findings of amyotrophic lateral sclerosis with FUS mutations by confirming the absence of TATA-binding protein-associated factor 15 and Ewing's sarcoma alterations upon expression of mutant fused in sarcoma. In contrast, all endogenous FET proteins were recruited into cytoplasmic stress granules upon general inhibition of Transportin-mediated nuclear import, mimicking the findings in frontotemporal lobar degeneration with fused in sarcoma pathology. These results allow a separation of fused in sarcoma proteinopathies caused by FUS mutations from those without a known genetic cause based on neuropathological features. More importantly, our data imply different pathological processes underlying inclusion formation and cell death between both conditions; the pathogenesis in amyotrophic lateral sclerosis with FUS mutations appears to be more restricted to dysfunction of fused in sarcoma, while a more global and complex dysregulation of all FET proteins is involved in the subtypes of frontotemporal lobar degeneration with fused in sarcoma patholog

Human Hsp70 Disaggregase reverses Parkinson’s-linked α-Synuclein Amyloid Fibrils

Intracellular amyloid fibrils linked to neurodegenerative disease typically accumulate in an age-related manner, suggesting inherent cellular capacity for counteracting amyloid formation in early life. Metazoan molecular chaperones assist native folding and block polymerization of amyloidogenic proteins, preempting amyloid fibril formation. Chaperone capacity for amyloid disassembly, however, is unclear. Here, we show that a specific combination of human Hsp70 disaggregase-associated chaperone components efficiently disassembles α-synuclein amyloid fibrils characteristic of Parkinson’s disease in vitro. Specifically, the Hsc70 chaperone, the class B J-protein DNAJB1, and an Hsp110 family nucleotide exchange factor (NEF) provide ATP-dependent activity that disassembles amyloids within minutes via combined fibril fragmentation and depolymerization. This ultimately generates non-toxic α-synuclein monomers. Concerted, rapid interaction cycles of all three chaperone components with fibrils generate the power stroke required for disassembly. This identifies a powerful human Hsp70 disaggregase activity that efficiently disassembles amyloid fibrils and points to crucial yet undefined biology underlying amyloid-based diseases