共查询到10条相似文献,搜索用时 0 毫秒
1.
2.
Masato Maesako Maiko Uemura Yoshitaka Tashiro Kazuki Sasaki Kiwamu Watanabe Yasuha Noda Karin Ueda Megumi Asada-Utsugi Masakazu Kubota Katsuya Okawa Masafumi Ihara Shun Shimohama Kengo Uemura Ayae Kinoshita 《PloS one》2015,10(9)
Obesity and type 2 diabetes are risk factors of Alzheimer’s disease (AD). We reported that a high fat diet (HFD) promotes amyloid precursor protein (APP) cleavage by β-site APP cleaving enzyme 1 (BACE1) without increasing BACE1 levels in APP transgenic mice. However, the detailed mechanism had remained unclear. Here we demonstrate that HFD promotes BACE1/Adaptor protein-2 (AP-2)/clathrin complex formation by increasing AP-2 levels in APP transgenic mice. In Swedish APP overexpressing Chinese hamster ovary (CHO) cells as well as in SH-SY5Y cells, overexpression of AP-2 promoted the formation of BACE1/AP-2/clathrin complex, increasing the level of the soluble form of APP β (sAPPβ). On the other hand, mutant D495R BACE1, which inhibits formation of this trimeric complex, was shown to decrease the level of sAPPβ. Overexpression of AP-2 promoted the internalization of BACE1 from the cell surface, thus reducing the cell surface BACE1 level. As such, we concluded that HFD may induce the formation of the BACE1/AP-2/clathrin complex, which is followed by its transport of BACE1 from the cell surface to the intracellular compartments. These events might be associated with the enhancement of β-site cleavage of APP in APP transgenic mice. Here we present evidence that HFD, by regulation of subcellular trafficking of BACE1, promotes APP cleavage. 相似文献
3.
Christian B. Lessard Barbara A. Cottrell Hiroko Maruyama Suraj Suresh Todd E. Golde Edward H. Koo 《PloS one》2015,10(12)
The relative increase in Aβ42 peptides from familial Alzheimer disease (FAD) linked APP and PSEN mutations can be related to changes in both ε-cleavage site utilization and subsequent step-wise cleavage. Cleavage at the ε-site releases the amyloid precursor protein (APP) intracellular domain (AICD), and perturbations in the position of ε-cleavage are closely associated with changes in the profile of amyloid β-protein (Aβ) species that are produced and secreted. The mechanisms by which γ-secretase modulators (GSMs) or FAD mutations affect the various γ-secretase cleavages to alter the generation of Aβ peptides have not been fully elucidated. Recent studies suggested that GSMs do not modulate ε-cleavage of APP, but the data were derived principally from recombinant truncated epitope tagged APP substrate. Here, using full length APP from transfected cells, we investigated whether GSMs modify the ε-cleavage of APP under more native conditions. Our results confirmed the previous findings that ε-cleavage is insensitive to GSMs. In addition, fenofibrate, an inverse GSM (iGSM), did not alter the position or kinetics of ε-cleavage position in vitro. APH1A and APH1B, a subunit of the γ-secretase complex, also modulated Aβ42/Aβ40 ratio without any alterations in ε-cleavage, a result in contrast to what has been observed with PS1 and APP FAD mutations. Consequently, GSMs and APH1 appear to modulate γ-secretase activity and Aβ42 generation by altering processivity but not ε-cleavage site utilization. 相似文献
4.
Markus P. Kummer Hiroko Maruyama Claudia Huelsmann Sandra Baches Sascha Weggen Edward H. Koo 《The Journal of biological chemistry》2009,284(4):2296-2306
The formation of insoluble cross β-sheet amyloid is pathologically
associated with disorders such as Alzheimer, Parkinson, and Huntington
diseases. One exception is the nonpathological amyloid derived from the
protein Pmel17 within melanosomes to generate melanin pigment. Here we show
that the formation of insoluble MαC intracellular fragments of Pmel17,
which are the direct precursors to Pmel17 amyloid, depends on a novel
juxtamembrane cleavage at amino acid position 583 between the furin-like
proprotein convertase cleavage site and the transmembrane domain. The
resulting Pmel17 C-terminal fragment is then processed by the
γ-secretase complex to release a short-lived intracellular domain
fragment. Thus, by analogy to the Notch receptor, we designate this cleavage
the S2 cleavage site, whereas γ-secretase mediates proteolysis at the
intramembrane S3 site. Substitutions or deletions at this S2 cleavage site,
the use of the metalloproteinase inhibitor TAPI-2, as well as small
interfering RNA-mediated knock-down of the metalloproteinases ADAM10 and 17
reduced the formation of insoluble Pmel17 fragments. These results demonstrate
that the release of the Pmel17 ectodomain, which is critical for melanin
amyloidogenesis, is initiated by S2 cleavage at a juxtamembrane position.Folding of proteins is a highly regulated process ensuring their correct
three-dimensional structure. Under pathological circumstances, a soluble
protein can be folded into highly stable cross β-sheet amyloid
structures, which are believed to play pathological roles in disorders such as
Alzheimer, Parkinson, and Huntington diseases. An exception to this general
concept is the physiological amyloid structure of the melanosomal matrix
formed by the protein Pmel17. Melanosomes are lysosome-related organelles that
contain pigment granules (melanin) in melanocytes and retinal epithelial cells
(reviewed in Ref. 1).
Melanogenesis is believed to proceed through several sequential maturation
steps, classified by melanosomes from stage I to stage IV. Maturation of stage
II melanosomes requires the formation of Pmel17 intralumenal fibers
(2,
3).Pmel17 (also called gp100, ME20, RPE1, or silver) is a type I transmembrane
glycoprotein of up to 668 amino acids in humans (reviewed in Ref.
4). The requirement of Pmel17
for the generation of functional melanin has been shown in a number of
different organisms, because, for example, certain point mutations in the
Pmel17/silver gene result in hypopigmentation phenotypes
(5–7).
The most characteristic domain within Pmel17 is a specific lumenal
proline/serine/threonine rich repeat domain (see
Fig. 1A), that is
imperfectly repeated 13 times in the Mα fragment. Importantly, deletion
of the rich repeat domain results in a complete loss of fibril formation,
pointing to the requirement of Pmel17, and especially the rich repeat domain,
in melanin formation (8).
Pmel17 exists in different isoforms generated by alternative splicing.
Pmel17-i2 is the most
abundant isoform, whereas the Pmel17-l isoform contains a 7-amino acid
insertion close to the transmembrane domain
(9,
10).Open in a separate windowFIGURE 1.Effect of the γ-secretase inhibitor DAPT on Pmel17 processing.
A, schematic diagram of Pmel17 and epitopes of antibodies. Pmel17
contains five potential N-glycosylation sites indicated by branched
structures. The long form of Pmel17, Pmel17-l, is characterized by a seven
amino acid insertion (VPGILLT) within the lumenal domain close to the
transmembrane domain (TM), which is absent in Pmel17-i. NVS marks a
potential N-glycosylation site near this insertion. The epitopes of
antibodies αPep13h and HMB45 are indicated. Cleavage by a furin-like PC
results in the formation of the Mα and the membrane-bound 26-kDa Mβ
fragment, which are connected via disulfide bonds. Release and further
processing of the Mα fragment into MαN and MαC fragments
results in the formation of fibrils and marks the transition of stage I to
stage II melanosomes (dashed line). B, human MNT-1 cells
were incubated with increasing amounts of DAPT for 18 h, and then the lysates
were separated by SDS-PAGE and analyzed by immunoblotting with αPep13h
antibody. DAPT treatment resulted in the accumulation of a C-terminal fragment
of Pmel17 (CTF), whereas Pmel17 P1 and Mβ fragment were unchanged.
C, probing the Triton-soluble fraction with HMB45 revealed increased
amounts of the highly glycosylated P2 form of Pmel17 after DAPT incubation.
D, detection of Pmel17 amyloidogenic fragments (MαC) in the
SDS-extracted insoluble pellet using antibody HMB45. E, murine B16-FO
cells treated with increasing concentrations of DAPT. Immunoblotting using
antibodyαPep13h revealed the formation of CTF of similar size as in
MNT-1 cells. F, time course analysis of Pmel17, Mβ, and
Pmel17-CTF after DAPT treatment. The cell lysates were immunoblotted using
αPep13h. Pmel17-CTF was detectable after 10 min of incubation with 1
μm DAPT. G, the size of the Pmel17-CTF was determined
using an unstained low molecular range peptide standard. The marker peptides
were detected by Ponceau S staining and Pmel17-CTF were detected by immunoblot
using αPep13h.Pmel17 traffics through the secretory pathway as a 100-kDa protein (called
P1). In the late Golgi compartment it undergoes further glycosylation,
resulting in a short lived 120-kDa protein (called P2). P2 is rapidly cleaved
within the post-Golgi by a furin-like proprotein convertase (PC) to generate
two fragments that remain tethered to each other by disulfide bonds: a
C-terminal polypeptide containing the transmembrane domain (Mβ) and a
large N-terminal ectodomain (Mα)
(2)
(Fig. 1A).
Consequently, inhibition of this furin-like activity not only prevents the
generation of Mα and Mβ fragments but also inhibits the formation
of melanosomal striation in HeLa cells
(3). These findings suggest
that Mα must first be dissociated from the Mβ for melanogenesis to
proceed. It is unclear how Mα is released from the membrane. Reduction
of disulfide bonds would release Mα from Mβ; alternatively,
proteolytic digestion of Mβ should also free Mα from the membrane
tether. It has been speculated that, given the presence of lysosomal
hydrolases in melanosomes and proteolytic maturation of Pmel17, proteolysis is
the more likely mechanism (4).
Recently, it was shown that recombinant Mα is able to form amyloid
structures in vitro in an unprecedented rapidity, and furthermore,
Pmel17 amyloid also accelerated melanin formation
(11). These findings
demonstrate that mammalian amyloid formed by Pmel17 is functional and
physiological.The insoluble pool of Pmel17 in cells consists mostly of truncated Mα
C-terminal fragments (MαC) of heterogeneous sizes, indicating that
further processing of Mα occurs after its release from the membrane
(8,
12). MαC fragments are
found in the insoluble fraction of melanocytes as well as in nonmelanotic
cells, the latter after overexpression of Pmel17
(8), and are reduced or absent
in amelanotic cells (8,
13,
14). Meanwhile, the C-terminal
fragment derived from the Mβ fragment and recognized by a C-terminal
specific epitope antibody is less stable, indicating rapid turnover
(2).The presenilin (PS) family of proteins consists of two homologous integral
transmembrane proteins, PS1 and PS2, which are part of the γ-secretase
complex. The latter consists of presenilin 1 or 2, nicastrin, APH-1, and PEN-2
(15) and catalyzes the
cleavage of the hydrophobic transmembrane domain of a burgeoning list of
proteins, also called regulated intramembrane cleavage. Other substrates for
the γ-secretase-mediated intramembrane cleavage include Notch, amyloid
precursor protein (APP), cadherin (E-cadherin), nectin-1, the low density
lipoprotein-related receptor, CD44, ErbB-4, the voltage-gated sodium channel
β2-subunit, and the Notch ligands Delta and Jagged. Importantly, in
Alzheimer disease, the presenilin-mediated γ-secretase cleavage of APP
releases the amyloid β-protein fragment, a peptide believed to play a key
role in Alzheimer disease pathogenesis. Interestingly, a recent report
described the absence of melanin pigment in presenilin-deficient animals, an
observation confirmed by the lack of melanin formation in cells treated with
γ-secretase inhibitors
(16). The mechanism
responsible for this finding is unclear, leading us to ask whether Pmel17
processing is a presenilin-dependent process and, if so, whether this cleavage
is involved in melanogenesis.In this study, we show the presence of an endoproteolytic activity that
cleaves the extracellular domain of Pmel17-i at a juxtamembrane position
between the known PC cleavage site and the transmembrane domain, which we term
the S2 cleavage site, by a TAPI-sensitive ADAM (a disintegrin
and metalloproteinase protein) protease. This
intracellular shedding of Pmel17 after S2 cleavage results in the liberation
of the Mα N-terminal ectodomain, the precursor to Pmel17 amyloid, which
is able to form insoluble Pmel17 aggregates. The C-terminal transmembrane
fragment generated by S2 cleavage is further processed by γ-secretase
(S3 cleavage) to release the Pmel17 intracellular domain, which is then
rapidly degraded. 相似文献
5.
Jose Henrique Pereira Richard A. Heins Daniel L. Gall Ryan P. McAndrew Kai Deng Keefe C. Holland Timothy J. Donohue Daniel R. Noguera Blake A. Simmons Kenneth L. Sale John Ralph Paul D. Adams 《The Journal of biological chemistry》2016,291(19):10228-10238
There has been great progress in the development of technology for the conversion of lignocellulosic biomass to sugars and subsequent fermentation to fuels. However, plant lignin remains an untapped source of materials for production of fuels or high value chemicals. Biological cleavage of lignin has been well characterized in fungi, in which enzymes that create free radical intermediates are used to degrade this material. In contrast, a catabolic pathway for the stereospecific cleavage of β-aryl ether units that are found in lignin has been identified in Sphingobium sp. SYK-6 bacteria. β-Aryl ether units are typically abundant in lignin, corresponding to 50–70% of all of the intermonomer linkages. Consequently, a comprehensive understanding of enzymatic β-aryl ether (β-ether) cleavage is important for future efforts to biologically process lignin and its breakdown products. The crystal structures and biochemical characterization of the NAD-dependent dehydrogenases (LigD, LigO, and LigL) and the glutathione-dependent lyase LigG provide new insights into the early and late enzymes in the β-ether degradation pathway. We present detailed information on the cofactor and substrate binding sites and on the catalytic mechanisms of these enzymes, comparing them with other known members of their respective families. Information on the Lig enzymes provides new insight into their catalysis mechanisms and can inform future strategies for using aromatic oligomers derived from plant lignin as a source of valuable aromatic compounds for biofuels and other bioproducts. 相似文献
6.
Wanxia He Jinxuan Hu Yuxing Xia Riqiang Yan 《The Journal of biological chemistry》2014,289(30):20630-20637
BACE1 is a type I transmembrane aspartyl protease that cleaves amyloid precursor protein at the β-secretase site to initiate the release of β-amyloid peptide. As a secretase, BACE1 also cleaves additional membrane-bound molecules by exerting various cellular functions. In this study, we showed that BACE1 can effectively shed the membrane-anchored signaling molecule Jagged 1 (Jag1). We also mapped the cleavage sites of Jag1 by ADAM10 and ADAM17. Although Jag1 shares a high degree of homology with Jag2 in the ectodomain region, BACE1 fails to cleave Jag2 effectively, indicating a selective cleavage of Jag1. Abolished cleavage of Jag1 in BACE1-null mice leads to enhanced astrogenesis and, concomitantly, reduced neurogenesis. This characterization provides biochemical evidence that the Jag1-Notch pathway is under the control of BACE1 activity. 相似文献
7.
Cornelia M. Wilson Amandine Magnaudeix Catherine Yardin Faraj Terro 《The Journal of biological chemistry》2011,286(36):31080-31091
The oligosaccharyltransferase complex catalyzes the transfer of oligosaccharide from a dolichol pyrophosphate donor en bloc onto a free asparagine residue of a newly synthesized nascent chain during the translocation in the endoplasmic reticulum lumen. The role of the less known oligosaccharyltransferase (OST) subunits, DC2 and KCP2, recently identified still remains to be determined. Here, we have studied DC2 and KCP2, and we have established that DC2 and KCP2 are substrate-specific, affecting amyloid precursor protein (APP), indicating that they are not core components required for N-glycosylation and OST activity per se. We show for the first time that DC2 and KCP2 depletion affects APP processing, leading to an accumulation of C-terminal fragments, both C99 and C83, and a reduction in full-length mature APP. This reduction in mature APP levels was not due to a block in secretion because the levels of sAPPα secreted into the media were unaffected. We discover that DC2 and KCP2 depletion affects only the γ-secretase complex, resulting in a reduction of the PS1 active fragment blocking Aβ production. Conversely, we show that the overexpression of DC2 and KCP2 causes an increase in the active γ-secretase complex, particularly the N-terminal fragment of PS1 that is generated by endoproteolysis, leading to a stimulation of Aβ production upon overexpression of DC2 and KCP2. Our findings reveal that components of the OST complex for the first time can interact with the γ-secretase and affect the APP processing pathway. 相似文献
8.
Hyo-Jin Park Daniil Shabashvili Michael D. Nekorchuk Eva Shyqyriu Joo In Jung Thomas B. Ladd Brenda D. Moore Kevin M. Felsenstein Todd E. Golde Seong-Hun Kim 《The Journal of biological chemistry》2012,287(48):40629-40640
The presence of neuritic plaques containing aggregated amyloid-β (Aβ) peptides in the brain parenchyma is a pathological hallmark of Alzheimer disease (AD). Aβ is generated by sequential cleavage of the amyloid β precursor protein (APP) by β- and γ-secretase, respectively. As APP processing to Aβ requires transport through the secretory pathway, trafficking of the substrate and access to the secretases are key factors that can influence Aβ production (Thinakaran, G., and Koo, E. H. (2008) Amyloid precursor protein trafficking, processing, and function. J. Biol. Chem. 283, 29615–29619). Here, we report that retention in endoplasmic reticulum 1 (RER1) associates with γ-secretase in early secretory compartments and regulates the intracellular trafficking of γ-secretase. RER1 overexpression decreases both γ-secretase localization on the cell surface and Aβ secretion and conversely RER1 knockdown increases the level of cell surface γ-secretase and increases Aβ secretion. Furthermore, we find that increased RER1 levels decrease mature APP and increase immature APP, resulting in less surface accumulation of APP. These data show that RER1 influences the trafficking and localization of both γ-secretase and APP, thereby regulating the production and secretion of Aβ peptides. 相似文献
9.
《The Journal of biological chemistry》2014,289(26):17970
10.
We present the results obtained with an in vitro model system that resembles the in vivo tumour microenvironment, where malignant
cells are in close contact with the infiltrating lymphocytes. Unmanipulated blood lymphocytes were cytotoxic against the autologous
ex vivo tumour cells in 3/19 patients and this function was generated in 6-day mixed cultures in five additional cases. Production
of transforming growth factor β (TGFβ) by the freshly separated tumour cells was determined in parallel. Cytotoxicity was
generated by a small number of tumour cells (2–5/100 lymphocytes), while a large number (10–20/100 lymphocytes) inhibited
not only the generation but also the existing lytic activity. The presence of a neutralising TGFβ-specific mAb (2G7) potentiated
the activation of lymphocytes and suspended the suppression inflicted by the tumour cells. In those tumours, which expressed
relatively high levels of MHC class I and ICAM-1 molecules, the quantity of secreted TGFβ interfered with the ability of tumour
cells to generate cytotoxic lymphocytes. In the tumours with low expression of class I, such a correlation was not detected,
indicating the primordial role of MHC class I expression in the regulation of autologous tumour recognition. Our results demonstrate
the involvement of TGFβ in the impaired lymphocyte-mediated reactivity against immunogenic tumours and support a mechanism
that contrasts the tolerance or anergy. Since presence of TGFβ in the microenvironment of tumours counteracts the function
of cytotoxic T lymphocytes, production of this cytokine can contribute to the failure of immunotherapy.
Received: 19 June 1997 / Accepted: 14 August 1997 相似文献