-
[show abstract]
[hide abstract]
ABSTRACT: Presenilin-associated protein (PSAP) has been identified as a mitochondrial proapoptotic protein. However, the mechanism by which PSAP induces apoptosis remains unknown. To this end, we have established an inducible expression system. Using this system, we have examined the roles of B-cell lymphoma 2 (Bcl-2) family proteins, cytochrome c, Smac (Smac/Diablo, second mitochondria-derived activator of caspases/direct IAP binding protein with low PI), and Apaf-1 (apoptotic protease-activating factor) in PSAP-induced apoptosis. Our results demonstrate that knockdown of Apaf-1 abolished PSAP-induced caspase activation and poly(ADP ribose) polymerase (PARP) cleavage, indicating that the apoptosome formation triggered by cytochrome c is crucial for PSAP-induced apoptosis. Our data also demonstrate that knockdown of Smac abolished PSAP-induced caspase activation and PARP cleavage, indicating that, in addition to Apaf-1 or apoptosome formation, Smac is also essential for PSAP-induced apoptosis. However, interestingly, our data demonstrate that overexpression of Bcl-2 and Bcl-xL did not protect cells from PSAP-induced apoptosis, and that knockdown of Bid, Bax, and Bak had no effect on PSAP-induced cytochrome c and Smac release, indicating that PSAP-induced apoptosis is not regulated by Bcl-2 family proteins. These results strongly suggest that PSAP evokes mitochondrial apoptotic cascades via a novel mechanism that is not regulated by Bcl-2 family proteins, but that both the formation of cytochrome c-Apaf-1 apoptosome and the presence of Smac are absolutely required for PSAP-induced apoptosis.
Biochimica et Biophysica Acta 11/2012; · 4.66 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The abnormal production and accumulation of β-amyloid peptide (Aβ), which is produced from amyloid precursor protein (APP) by the sequential actions of β-secretase and γ-secretase, are thought to be the initial causative events in the development of Alzheimer's disease (AD). Accumulating evidence suggests that vascular factors play an important role in the pathogenesis of AD. Specifically, studies have suggested that one vascular factor in particular, oxidized low density lipoprotein (oxLDL), may play an important role in regulating Aβ formation in AD. However, the mechanism by which oxLDL modulates Aβ formation remains elusive. In this study, we report several new findings that provide biochemical evidence suggesting that the cardiovascular risk factor oxLDL may contribute to Alzheimer's disease by increasing Aβ production. First, we found that lysophosphatidic acid (LPA), the most bioactive component of oxLDL induces increased production of Aβ. Second, our data strongly indicate that LPA induces increased Aβ production via upregulating β-secretase expression. Third, our data strongly support the notion that different isoforms of protein kinase C (PKC) may play different roles in regulating APP processing. Specifically, most PKC members, such as PKCα, PKCβ, and PKCε, are implicated in regulating α-secretase-mediated APP processing; however, PKCδ, a member of the novel PKC subfamily, is involved in LPA-induced upregulation of β-secretase expression and Aβ production. These findings may contribute to a better understanding of the mechanisms by which the cardiovascular risk factor oxLDL is involved in Alzheimer's disease.
Biochimica et Biophysica Acta 10/2012; · 4.66 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: γ-secretase is a protease complex with at least four components: presenilin, nicastrin (NCT), anterior pharynx-defective 1 (Aph-1), and presenilin enhancer 2 (Pen-2). In this study, using knockout cell lines and small interfering RNA technology, our data demonstrated that the disappeared presenilin 1 C-terminal fragment (PS1C) caused by knockdown of pen-2 or knockout of NCT or Aph-1 was recovered by the addition of proteasome inhibitors, indicating that Pen-2, as well as NCT and Aph-1α, is dispensable for presenilin endoproteolysis. Our data also demonstrate that the formation of the nicastrin-Aph-1 subcomplex plays not only an important role in γ-secretase complex assembly but also in recruiting substrate C-terminal fragment of amyloid precursor protein generated by β-cleavage. Ablating any one component resulted in the instability of other components of the γ-secretase complex, and the presence of all three of the other components is required for full maturation of NCT.
Journal of Neurochemistry 09/2012; · 4.06 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Cells undergo apoptosis through two major pathways, the extrinsic pathway (death receptor pathway) and the intrinsic pathway (the mitochondrial pathway). These two pathways can be linked by caspase-8-activated truncated Bid formation. Very recently, death receptor 6 (DR6) was shown to be involved in the neurodegeneration observed in Alzheimer disease. DR6, also known as TNFRSF21, is a relatively new member of the death receptor family, and it was found that DR6 induces apoptosis when it is overexpressed. However, how the death signal mediated by DR6 is transduced intracellularly is not known. To this end, we have examined the roles of caspases, apoptogenic mitochondrial factor cytochrome c, and the Bcl-2 family proteins in DR6-induced apoptosis. Our data demonstrated that Bax translocation is absolutely required for DR6-induced apoptosis. On the other hand, inhibition of caspase-8 and knockdown of Bid have no effect on DR6-induced apoptosis. Our results strongly suggest that DR6-induced apoptosis occurs through a new pathway that is different from the type I and type II pathways through interacting with Bax.
Journal of Biological Chemistry 07/2012; 287(34):29125-33. · 4.77 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Lysophosphatidic acid (LPA) modulates vascular cell function in vitro and in vivo via regulating the expression of specific genes. Previously, we reported that a transcriptional mechanism controls LPA-induced expression of Egr-1 in vascular smooth muscle cells. Egr-1 is a master transcription factor mediating the expression of various genes that have been implied to modulate a broad spectrum of vascular pathologies. In this study, we determined the essential intracellular signaling pathway leading to LPA-induced Egr-1 expression. Our data demonstrate that activation of ERK1/2 and JNK, but not p38 MAPK, is required for LPA-induced Egr-1 expression in smooth muscle cells. We provide the first evidence that MEK-mediated JNK activation leads to LPA-induced gene expression. JNK2 is required for Egr-1 induction. Examining the upstream kinases that mediate ERK and JNK activation, leading to Egr-1 expression, we found that LPA-induced activation of MAPKs and expression of Egr-1 are dependent on PKC activation. We observed that LPA rapidly activates PKCδ and PKCθ. Overexpression of dominant-negative PKCδ, but not dominant-negative PKCθ, diminished activation of ERK and JNK and blocked LPA-induced expression of Egr-1 mRNA and protein. We also evaluated LPA receptor involvement. Our data reveal an intracellular regulatory mechanism: LPA induction of Egr-1 expression is via LPA cognate receptor (LPA receptor 1)-dependent and PKCδ-mediated ERK and JNK activation. This study provides the first evidence that PKCδ mediates ERK and JNK activation in the LPA signaling pathway and that this pathway is required for LPA-induced gene regulation as evidenced by Egr-1 expression.
Journal of Biological Chemistry 05/2012; 287(27):22635-42. · 4.77 Impact Factor
-
Mei-Zhen Cui
[show abstract]
[hide abstract]
ABSTRACT: Lysophosphatidic acid (LPA) has been found to accumulate in high concentrations in atherosclerotic lesions. LPA is a bioactive phospholipid produced by activated platelets and formed during the oxidation of LDL. Accumulating evidence suggests that this lipid mediator may serve as an important risk factor for development of atherosclerosis and thrombosis. The role of LPA in atherogenesis is supported by the evidence that LPA: stimulates endothelial cells to produce adhesion molecules and chemoattractants; induces smooth muscle cells to produce inflammatory cytokines; stimulates smooth muscle cell dedifferentiation, proliferation, and migration; increases monocyte migration and decreases monocyte-derived cell emigration from the vessel wall; induces hypertension and vascular neointimal formation in vivo; and promotes plaque progression in a mouse atherosclerosis model. The role of LPA in thrombogenesis is supported by the evidence that LPA markedly induces the aggregation of platelets and the expression of tissue factor, which is the principal initiator of blood coagulation. Recent experimental data indicate that LPA is produced by specific enzymes and that LPA binds to and activates multiple G-protein-coupled receptors, leading to intracellular signaling. Therapeutics targeting LPA biosynthesis, metabolism and signaling pathways could be viable for prevention and treatment of atherosclerosis and thrombosis.
Clinical lipidology. 08/2011; 6(4):413-426.
-
[show abstract]
[hide abstract]
ABSTRACT: Lysophosphatidic acid (LPA) is a potent bioactive lysophospholipid. Accumulated evidence supports a role for LPA in inflammation. To profile LPA-induced cytokine production in vascular smooth muscle cells (SMCs), we used a cytokine antibody array system and found that LPA prominently induces the secretion of IL-6 and monocyte chemoattractant protein (MCP)-1 from human aortic SMCs (HASMCs). The mechanism by which LPA induces MCP-1 expression in SMCs has been previously reported. However, LPA induction of IL-6 secretion from vascular SMCs and its regulatory mechanism are unknown. The present study reveals that LPA induces the expression of IL-6 mRNA and protein in HASMCs as well as the secretion of IL-6 protein in a time-dependent manner. Our results demonstrate that LPA-specific receptor 1 (LPA(1)) mediates LPA-induced IL-6 secretion and that LPA induction of IL-6 is independent of the EGF receptor pathway. Our data further show that PKC-mediated p38 MAPK is responsible for the IL-6 secretion. Finally, small interfering RNA depletion experiments revealed that p38alpha is specifically responsible for the LPA-induced IL-6 secretion. The present study profiles the regulatory relationship between LPA and multiple cytokines in vascular SMCs for the first time, provides the first evidence that LPA upregulates IL-6 in vascular SMCs, and reveals the regulatory mechanism of LPA-induced IL-6 production in HASMCs. In light of the emerging roles of LPA and IL-6 in vascular inflammation, the understanding of the regulatory mechanism may contribute to the treatment and prevention of cardiovascular disorders.
AJP Heart and Circulatory Physiology 03/2010; 298(3):H974-83. · 3.71 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Most of the Alzheimer's disease (AD)-linked mutations in amyloid precursor protein (APP), which cause abnormal production of beta-amyloid (Abeta), are localized at the major beta-secretase-and gamma-secretase cleavage sites. In this study, using an APP-knockout mouse neuronal cell line, our data demonstrated that at the P2-P1 positions of the epsilon-cleavage site at Abeta49 and the zeta-cleavage site at Abeta46, aromatic amino acids caused a strong reduction in total Abeta. On the other hand, residues with a long side chain caused a decrease in Abeta(40) and a concomitant increase in Abeta(42) and Abeta(38). These findings indicate that the structures of the substituting residues at these key positions strongly determine the efficiency and preference of gamma-secretase-mediated APP processing, which determines the ratio of different secreted Abeta species, a crucial factor in the disease development. Our findings provide new insight into the mechanisms of gamma-secretase-mediated APP processing and, specifically, into why most AD-linked APP mutations are localized at major gamma-secretase cleavage sites. This information may contribute to the development of methods of prevention and treatment of Alzheimer's disease aimed at modulating gamma-secretase activity.
Neurobiology of Disease 09/2009; 36(3):453-60. · 5.40 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Gamma-secretase-mediated processing of the amyloid-beta protein precursor (AbetaPP) is a crucial step in the formation of the amyloid-beta peptide (Abeta), but little is known about how the substrate AbetaPP interacts with the gamma-secretase complex. To understand the molecular events involved in gamma-secretase-mediated AbetaPP processing and Abeta formation, in the present study we determined the role of a well conserved GxxxG motif in the transmembrane domain of AbetaPP. Our data clearly demonstrate that substitution of aspartic acid for the key glycine residues in the GxxxG motif almost completely abolished the formation of Abeta. Furthermore, our data revealed that substitution of aspartic acid for the glycine in this GxxxG motif disrupts the interaction of AbetaPP with the gamma-secretase complex. Thus, the present study revealed an essential role for the GxxxG motif in the interaction of AbetaPP with the gamma-secretase complex and the formation of Abeta.
Journal of Alzheimer's disease: JAD 08/2009; 18(1):167-76. · 3.74 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: OBJECTIVE: Monocyte activation and migration are crucial events in the development of atherosclerosis and other inflammatory diseases. This study examined the role of protein kinase D (PKD) in monocyte migration. Method and Results- PKD2 is the predominant isoform of PKD expressed in monocytic THP-1 cells and primary human monocytes. Lysophosphatidylcholine (lysoPC), a prominent component of oxidized low-density lipoprotein, induces rapid and marked PKD activation in these cells. Using multiple approaches, including dominant-negative mutants and small interfering RNA knock-down, we found that lysoPC-induced PKD2 activation was required for the activation of both ERK and p38 MAPK. p38 MAPK mediation of lysoPC-induced monocytic cell migration was reported previously; our results reveal that the lysoPC-induced PKD2-p38 pathway controls monocyte migration. CONCLUSIONS: This study provides the first evidence that (1) lysoPC activates PKD, (2) PKD2 has a novel role in p38 activation, and (3) the PKD2-activated p38 pathway is responsible for lysoPC-induced migration of THP-1 cells and human monocytes. Thus, PKD is a novel and functional intracellular regulator in both lysoPC signaling and monocyte migration. These results suggest a new role for PKD2 in the development of atherosclerosis and other inflammatory diseases.
Arteriosclerosis Thrombosis and Vascular Biology 07/2009; 29(9):1376-82. · 6.37 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Histamine, a potent inflammatory mediator, has multiple effects on the pathogenesis of atherosclerosis. This study investigates the effect of histamine on the expression of early growth response factor 1 (Egr-1), a master transcription factor that regulates the expression of an array of atherogenic genes in atherosclerotic lesions. Histamine markedly and rapidly induces Egr-1 mRNA and protein expression in primary human aortic endothelial cells (HAECs). Histamine-induced Egr-1 expression is dependent on the activation of the H1 receptor. Histamine also rapidly and transiently activates protein kinase C-delta (PKCdelta), extracellular signal-regulated kinase (ERK)1/2, p38 kinase, and c-Jun N-terminal kinase (JNK) prior to Egr-1 induction. Using specific pharmacological inhibitors and small interfering RNA technology, we determined that PKCdelta and ERK, but not p38 and JNK, mediate histamine-induced Egr-1 expression. Our data provide the first evidence that histamine regulates expression of Egr-1 in mammalian cells and demonstrate a novel role of PKCdelta in up-regulation of Egr-1 expression. The present study reveals the following regulatory mechanism: histamine up-regulates Egr-1 expression in primary HAECs via the H1 receptor and the PKCdelta-dependent ERK activation pathway. Our data also imply that CREB, a downstream component of the ERK pathway, regulates Egr-1 expression in HAECs. Importantly, these results suggest a central role of Egr-1 in histamine-induced gene expression and in histamine-induced vascular disease.
Journal of Biological Chemistry 09/2008; 283(40):26928-36. · 4.77 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Overwhelming evidence supports the amyloid hypothesis of Alzheimer's disease that stipulates that the relative level of the 42 amino acid beta-amyloid peptide (Abeta(42)) in relationship to Abeta(40) is critical to the pathogenesis of the disease. While it is clear that the multi-subunit gamma secretase is responsible for cleavage of the amyloid precursor protein (APP) into Abeta(42) and Abeta(40), the exact molecular mechanisms regulating the production of the various Abeta species remain elusive. To elucidate the underlying mechanisms, we replaced individual amino acid residues from positions 43 to 52 of Abeta with phenylalanine to examine the effects on the production of Abeta(40) and Abeta(42). All mutants, except for V50F, resulted in a decrease in total Abeta with a more prominent reduction in Abeta for residues 45, 48, and 51, following an every three residue repetition pattern. In addition, the mutations with the strongest reductions in total Abeta had the largest increases in the ratio of Abeta(42)/Abeta(40). Curiously, the T43F, V44F, and T48F mutations caused a striking decrease in the accumulation of membrane bound Abeta(46), albeit by a different mechanism. Our data suggest that initial cleavage of APP at the epsilon site is crucial in the generation of Abeta. The implicated sequential cleavage and an alpha-helical model may lead to a better understanding of the gamma-secretase-mediated APP processing and may also provide useful information for therapy and drug design aimed at altering Abeta production.
Journal of Neurochemistry 09/2008; 107(3):722-33. · 4.06 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Presenilin-associated protein (PSAP) was originally identified as a PS1-associated, PDZ domain protein. In a subsequent study, PSAP was found to be a mitochondrial apoptotic molecule. In this study, we cloned the PSAP gene and found that it is composed of 12 exons and localizes on chromosome 6. To better understand the structure and function of PSAP, we have generated a series of antibodies that recognize different regions of PSAP. Using these antibodies, we found that PSAP is expressed in four isoforms as a result of differential splicing of exon 8 in addition to the use of either the first or the second ATG codon as the start codon. We also found that all these isoforms are localized in the mitochondria and are pro-apoptotic. Furthermore, our data revealed that the PDZ domain and N-terminal fragment are required for the pro-apoptotic activity of PSAP.
Biochimica et Biophysica Acta 05/2008; 1780(4):696-708. · 4.66 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Prostate cancer cell migration is an essential event both in the progression of prostate cancer and in the steps leading to metastasis. We report here that lysophosphatidic acid (LPA), a potent bioactive phospholipid, induces prostate cancer PC3 cell migration via the activation of the LPA(1) receptor, which is linked to a PTX-sensitive activation mechanism of the mitogen-activated protein kinases (MAPK). Our results demonstrate that parallel activation of ERK1/2 and p38, but not JNK, is responsible for LPA-stimulated PC3 cell migration. Furthermore, using small interfering RNA (siRNA) technology, and overexpressing dominant-negative mutants of p38 MAPK isotypes of alpha, beta, gamma and delta, we have identified that the activation of ERK2 (p42) and p38alpha, but not of ERK1 and the other isoforms of p38 MAPK, is required for LPA-induced migration. Our study provides the first evidence for a functional role of p42 and p38alpha in LPA-induced mammalian cell migration, and also demonstrates, for the first time, that the receptor LPA(1) mediates prostate cancer cell migration. The results of the present study suggest that LPA, the receptor LPA(1), ERK2 and p38alpha are important regulators for prostate cancer cell invasion and thus could play a significant role in the development of metastasis.
Biochimica et Biophysica Acta 08/2007; 1771(7):883-92. · 4.66 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: It has been hypothesized that different C-terminus of beta-amyloid peptide (Abeta) may be generated by different gamma-secretase activities. Recently, we have identified a new zeta-cleavage site at Abeta46, leading to an important finding that the C-terminus of Abeta is produced by a series of sequential cleavages. This finding prompted us to examine the effects of the known gamma-secretase inhibitors on different steps of the gamma-secretase-mediated sequential cleavages and specifically their effects on the formation and turnover of the intermediate Abeta(46). Our results demonstrate that some of the known inhibitors, such as L-685,458 and III-31C as well as inhibitors IV and V, inhibit the formation of secreted Abeta(40/42) by inhibiting the formation of the intermediate Abeta(46). However, most of the other inhibitors show no inhibitory effect on the formation of the intermediate Abeta(46), but rather inhibit the turnover of Abeta(46), resulting in its accumulation. In addition, the non-steroidal anti-inflammatory drugs (NSAIDs) ibuprofen and sulindac sulfide have no effect on the formation and turnover of Abeta(46), but rather modulate the ratio of secreted Abeta at a step after the formation of Abeta(40) and Abeta(42). Thus, our data strongly suggest that the multi-sequential intramembrane cleavages of amyloid precursor protein C (APP) are likely catalyzed by the same gamma-secretase.
Journal of Neurochemistry 04/2007; 100(5):1234-46. · 4.06 Impact Factor
-
Alzheimers & Dementia - ALZHEIMERS DEMENT. 01/2007; 3(3).
-
[show abstract]
[hide abstract]
ABSTRACT: Lysophosphatidic acid (LPA), one component of oxidized low-density lipoprotein, is a potent bioactive phospholipid. Early growth response gene-1 (Egr-1), an important transcription factor, regulates expression of an array of genes involved in vascular diseases. Whether and how LPA regulates the transcriptional machinery of Egr-1 gene is unknown and is addressed in this study.
We found that LPA markedly induces Egr-1 mRNA and protein in aortic smooth muscle cells (SMCs). RNA stability and nuclear run-on assays reveal that LPA-induced Egr-1 gene expression is controlled at the transcriptional level. Reporter gene analyses have shown that the -141 to +20 nt region of the Egr-1 promoter contains regulatory elements. Electrophoretic mobility shift assays reveal that the DNA-binding activities of both CREB and SRF to the CRE and SRE motifs of the Egr-1 promoter are markedly elevated in response to LPA. The increased binding activity depends on the phosphorylation of CREB and SRF. Luciferase assays of a series of deleted or mutated Egr-1 promoter-reporter gene constructs, along with dominant negative CREB transfection analysis revealed that the 2 CRE sites and the 2 proximal SRE sites in the Egr-1 promoter are required for maximal LPA-induced Egr-1 gene expression.
Our data reveal that LPA regulates Egr-1 expression via transcription factors CREB and SRF. These results establish a novel role for CREB in mediating LPA-induced gene expression. Our results imply that elevated LPA levels may, through activation of Egr-1, which regulates an array of atherogenic genes, exacerbate atheromatous lesions.
Arteriosclerosis Thrombosis and Vascular Biology 06/2006; 26(5):1029-35. · 6.37 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: We previously reported the phosphoinositide 3-kinase-dependent activation of the 5'-AMP-activated kinase (AMPK) by peroxynitrite (ONOO-) and hypoxia-reoxygenation in cultured endothelial cells. Here we show the molecular mechanism of activation of this pathway. Exposure of bovine aortic endothelial cells to ONOO- significantly increased the phosphorylation of both Thr172 of AMPK and Ser1179 of endothelial nitric-oxide synthase, a known downstream enzyme of AMPK. In addition, activation of AMPK by ONOO- was accompanied by increased phosphorylation of protein kinase Czeta (PKCzeta) (Thr410/403) and translocation of cytosolic PKCzeta into the membrane. Further, inhibition of PKCzeta abrogated ONOO- -induced AMPK-Thr172 phosphorylation as that of endothelial nitric-oxide synthase. Furthermore, overexpression of a constitutively active PKCzeta mutant enhanced the phosphorylation of AMPK-Thr172, suggesting that PKCzeta is upstream of AMPK activation. In contrast, ONOO- activated PKCzeta in LKB1-deficient HeLa-S3 but affected neither AMPK-Thr172 nor AMPK activity. These data suggest that LKB1 is required for PKCzeta-enhanced AMPK activation. In vitro, recombinant PKCzeta phosphorylated LKB1 at Ser428, resulting in phosphorylation of AMPK at Thr172. Further, direct mutation of Ser428 of LKB1 into alanine, like the kinase-inactive LKB1 mutant, abolished ONOO- -induced AMPK activation. In several cell types originating from human, rat, and mouse, inhibition of PKCzeta significantly attenuated the phosphorylation of both LKB1-Ser428 and AMPK-Thr172 that were enhanced by ONOO-. Taken together, we conclude that PKCzeta can regulate AMPK activity by increasing the Ser428 phosphorylation of LKB1, resulting in association of LKB1 with AMPK and consequent AMPK Thr172 phosphorylation by LKB1.
Journal of Biological Chemistry 04/2006; 281(10):6366-75. · 4.77 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The observations that three major cleavages within the transmembrane domain of APP, namely, the gamma-cleavage, -cleavage, and the newly identified zeta-cleavage, are involved in the generation of secreted Abeta40 and Abeta42 prompted us to determine how the calpain inhibitor III MDL 28170 influences these three cleavages and Abeta formation. With the use of a cell culture system, our data demonstrate that 1) at either high concentrations, or at a low range of concentrations, at early time points, MDL 28170 inhibits the formation of secreted Abeta40 and Abeta42. However, this effect is due to inhibition of the intermediate Abeta46 generation by zeta-cleavage and not due to direct inhibition of the gamma-cleavage that produces Abeta40/42 from Abeta46; 2) at low range of concentrations and at late time points, MDL 28170 causes an increase in secreted Abeta40/42 that likely results from inhibition of degradation of both the initial substrate, CTFbeta, and the final product, Abeta40/42, of gamma-secretase. These data strongly suggest that formation of Abeta46 is a key step in the gamma-secretase mediated generation of Abeta40/42 and provide a new target for the development of Abeta inhibitors. These data also suggest that calpain and related proteases, which are sensitive to MDL 28170, play an important role in the accumulation of secreted Abeta.
The FASEB Journal 03/2006; 20(2):331-3. · 5.71 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: beta-Amyloid precursor protein apparently undergoes at least three major cleavages, gamma-, epsilon-, and the newly identified zeta-cleavage, within its transmembrane domain to produce secreted beta-amyloid protein (Abeta). However, the roles of epsilon- and zeta-cleavages in the formation of secreted Abeta and the relationship among these three cleavages, namely epsilon-, zeta-, and gamma-cleavages, remain elusive. We investigated these issues by attempting to determine the formation and turnover of the intermediate products generated by these cleavages, in the presence or absence of known gamma-secretase inhibitors. By using a differential inhibition strategy, our data demonstrate that Abeta(46) is an intermediate precursor of secreted Abeta. Our co-immunoprecipitation data also reveal that, as an intermediate, Abeta(46) is tightly associated with presenilin in intact cells. Furthermore, we identified a long Abeta species that is most likely the long sought after intermediate product, Abeta(49), generated by epsilon-cleavage, and this Abeta(49) is further processed by zeta- and gamma-cleavages to generate Abeta(46) and ultimately the secreted Abeta(40/42). More interestingly, our data demonstrate that gamma-cleavage not only occurs last but also depends on zeta-cleavage occurring prior to it, indicating that zeta-cleavage is crucial for the formation of secreted Abeta. Thus, we conclude that the C terminus of secreted Abeta is most likely generated by a series of sequential cleavages, namely first epsilon-cleavage which is then followed by zeta- and gamma-cleavages, and that Abeta(46) produced by zeta-cleavage is the precursor of secreted Abeta(40/42).
Journal of Biological Chemistry 12/2005; 280(45):37689-97. · 4.77 Impact Factor
