A system for enhancing genome-wide coexpression dynamics study.
ABSTRACT: Statistical similarity analysis has been instrumental in elucidation of the voluminous microarray data. Genes with correlated expression profiles tend to be functionally associated. However, the majority of functionally associated genes turn out to be uncorrelated. One conceivable reason is that the expression of a gene can be sensitively dependent on the often-varying cellular state. The intrinsic state change has to be plastically accommodated by gene-regulatory mechanisms. To capture such dynamic coexpression between genes, a concept termed "liquid association" (LA) has been introduced recently. LA offers a scoring system to guide a genome-wide search for critical cellular players that may interfere with the coexpression of a pair of genes, thereby weakening their overall correlation. Although the LA method works in many cases, a direct extension to more than two genes is hindered by the "curse of dimensionality." Here we introduce a strategy of finding an informative 2D projection to generalize LA for multiple genes. A web site is constructed that performs on-line LA computation for any user-specified group of genes. We apply this scoring system to study yeast protein complexes by using the Saccharomyces cerevisiae protein complexes database of the Munich Information Center for Protein Sequences. Human genes are also investigated by profiling of 60 cancer cell lines of the National Cancer Institute. In particular, our system links the expression of the Alzheimer's disease hallmark gene APP (amyloid-beta precursor protein) to the beta-site-cleaving enzymes BACE and BACE2, the gamma-site-cleaving enzymes presenilin 1 and 2, apolipoprotein E, and other Alzheimer's disease-related genes.
Project description:Amyloid-? protein (A?) is the main component of neuritic plaques, the pathological hallmark of Alzheimer's disease (AD). ?-site APP cleaving enzyme 1 (BACE1) is a major ?-secretase contributing to A? generation. ?-site APP cleaving enzyme 2 (BACE2), the homolog of BACE1, is not only a ?-secretase but also a conditional ?-secretase. Previous studies showed that regulator of calcineurin 1 (RCAN1) is markedly increased by AD and promotes BACE1 expression. However, the role of RCAN1 in BACE2 regulation remains elusive. Here, we showed that RCAN1 increases BACE2 protein levels. Moreover, RCAN1 inhibits the turnover of BACE2 protein. Furthermore, RCAN1 attenuates proteasome-mediated BACE2 degradation, but not lysosome-mediated BACE2 degradation. Taken together, our work indicates that RCAN1 inhibits BACE2 turnover by attenuating proteasome-mediated BACE2 degradation. It advances our understanding of BACE2 regulation and provides a potential mechanism of BACE2 dysregulation in AD.
Project description:BACKGROUND:Alzheimer's disease is the most common neurodegenerative disease in the elderly. Amyloid-? protein (A?) is the major component of neuritic plaques which are the hallmark of AD pathology. ?-site APP cleaving enzyme 1 (BACE1) is the major ?-secretase contributing to A? generation. ?-site APP-cleaving enzyme 2 (BACE2), the homolog of BACE1, might play a complex role in the pathogenesis of Alzheimer's disease as it is not only a ?-secretase but also a conditional ?-secretase. Dysregulation of BACE2 is observed in Alzheimer's disease. However, the regulation of BACE2 is less studied compared with BACE1, including its degradation pathways. In this study, we investigated the turnover rates and degradation pathways of BACE2 in both neuronal cells and non-neuronal cells. RESULTS:Both lysosomal inhibition and proteasomal inhibition cause a time- and dose-dependent increase of transiently overexpressed BACE2 in HEK293 cells. The half-life of transiently overexpressed BACE2 protein is approximately 6?h. Moreover, the half-life of endogenous BACE2 protein is approximately 4?h in both HEK293 cells and mouse primary cortical neurons. Furthermore, both lysosomal inhibition and proteasomal inhibition markedly increases endogenous BACE2 in HEK293 cells and mouse primary cortical neurons. CONCLUSIONS:This study demonstrates that BACE2 is degraded by both the proteasome and lysosome pathways in both neuronal and non-neuronal cells at endogenous level and in transient overexpression system. It indicates that BACE2 dysregulation might be mediated by the proteasomal and lysosomal impairment in Alzheimer's disease. This study advances our understanding of the regulation of BACE2 and provides a potential mechanism of its dysregulation in Alzheimer's disease.
Project description:The ?-site APP cleaving enzymes 1 and 2 (BACE1 and BACE2) were initially identified as transmembrane aspartyl proteases cleaving the amyloid precursor protein (APP). BACE1 is a major drug target for Alzheimer's disease because BACE1-mediated cleavage of APP is the first step in the generation of the pathogenic amyloid-? peptides. BACE1, which is highly expressed in the nervous system, is also required for myelination by cleaving neuregulin 1. Several recent proteomic and in vivo studies using BACE1- and BACE2-deficient mice demonstrate a much wider range of physiological substrates and functions for both proteases within and outside of the nervous system. For BACE1 this includes axon guidance, neurogenesis, muscle spindle formation, and neuronal network functions, whereas BACE2 was shown to be involved in pigmentation and pancreatic ?-cell function. This review highlights the recent progress in understanding cell biology, substrates, and functions of BACE proteases and discusses the therapeutic options and potential mechanism-based liabilities, in particular for BACE inhibitors in Alzheimer's disease. The protease BACE1 is a major drug target in Alzheimer disease. Together with its homolog BACE2, both proteases have an increasing number of functions within and outside of the nervous system. This review highlights recent progress in understanding cell biology, substrates, and functions of BACE proteases and discusses the therapeutic options and potential mechanism-based liabilities, in particular for BACE inhibitors in Alzheimer disease.
Project description:Deposition of amyloid-? protein (A?) to form neuritic plaques is the characteristic neuropathology of Alzheimer's disease (AD). A? is generated from amyloid precursor protein (APP) by ?- and ?-secretase cleavages. BACE1 is the ?-secretase and its inhibition induces severe side effects, whereas its homolog BACE2 normally suppresses A? by cleaving APP/A? at the ?-site (Phe20) within the A? domain. Here, we report that BACE2 also processes APP at the ? site, and the juxtamembrane helix (JH) of APP inhibits its ?-secretase activity, enabling BACE2 to cleave nascent APP and aggravate AD symptoms. JH-disrupting mutations and clusterin binding to JH triggered BACE2-mediated ?-cleavage. Both BACE2 and clusterin were elevated in aged mouse brains, and enhanced ?-cleavage during aging. Therefore, BACE2 contributes to AD pathogenesis as a conditional ?-secretase and could be a preventive and therapeutic target for AD without the side effects of BACE1 inhibition.
Project description:?-Site APP-cleaving enzyme 1 (BACE1) inhibition is considered one of the most promising therapeutic strategies for Alzheimer's disease, but current BACE1 inhibitors also block BACE2. As the localization and function of BACE2 in the brain remain unknown, it is difficult to predict whether relevant side effects can be caused by off-target inhibition of BACE2 and whether it is important to generate BACE1-specific inhibitors. Here, we show that BACE2 is expressed in discrete subsets of neurons and glia throughout the adult mouse brain. We uncover four new substrates processed by BACE2 in cultured glia: vascular cell adhesion molecule 1, delta and notch-like epidermal growth factor-related receptor, fibroblast growth factor receptor 1, and plexin domain containing 2. Although these substrates were not prominently cleaved by BACE2 in healthy adult mice, proinflammatory TNF induced a drastic increase in BACE2-mediated shedding of vascular cell adhesion molecule 1 in CSF. Thus, although under steady-state conditions the effect of BACE2 cross-inhibition by BACE1-directed inhibitors is rather subtle, it is important to consider that side effects might become apparent under physiopathological conditions that induce TNF expression.
Project description:?-Secretase, the rate-limiting enzymatic activity in the production of the amyloid-? (A?) peptide, is a major target of Alzheimer's disease (AD) therapeutics. There are two forms of the enzyme: ?-site A? precursor protein cleaving enzyme (BACE) 1 and BACE2. Although BACE1 increases in late-stage AD, little is known about BACE2. We conducted a detailed examination of BACE2 in patients with preclinical to late-stage AD, including amnestic mild cognitive impairment, and age-matched controls, cases of frontotemporal dementia, and Down's syndrome. BACE2 protein and enzymatic activity increased as early as preclinical AD and were found in neurons and astrocytes. Although the levels of total BACE2 mRNA were unchanged, the mRNA for BACE2 splice form C (missing exon 7) increased in parallel with BACE2 protein and activity. BACE1 and BACE2 were strongly correlated with each other at all levels, suggesting that their regulatory mechanisms may be largely shared. BACE2 was also elevated in frontotemporal dementia but not in Down's syndrome, even in patients with substantial A? deposition. Thus, expression of both forms of ?-secretase are linked and may play a combined role in human neurologic disease. A better understanding of the normal functions of BACE1 and BACE2, and how these change in different disease states, is essential for the future development of AD therapeutics.
Project description:Amyloids are often associated with pathologic processes such as in Alzheimer's disease (AD), but can also underlie physiological processes such as pigmentation. Formation of pathological and functional amyloidogenic substrates can require precursor processing by proteases, as exemplified by the generation of A? peptide from amyloid precursor protein (APP) by beta-site APP cleaving enzyme (BACE)1 and ?-secretase. Proteolytic processing of the pigment cell-specific Melanocyte Protein (PMEL) is also required to form functional amyloid fibrils during melanogenesis, but the enzymes involved are incompletely characterized. Here we show that the BACE1 homologue BACE2 processes PMEL to generate functional amyloids. BACE2 is highly expressed in pigment cells and Bace2(-/-) but not Bace1(-/-) mice display coat color defects, implying a specific role for BACE2 during melanogenesis. By using biochemical and morphological analyses, combined with RNA silencing, pharmacologic inhibition, and BACE2 overexpression in a human melanocytic cell line, we show that BACE2 cleaves the integral membrane form of PMEL within the juxtamembrane domain, releasing the PMEL luminal domain into endosomal precursors for the formation of amyloid fibrils and downstream melanosome morphogenesis. These studies identify an amyloidogenic substrate of BACE2, reveal an important physiological role for BACE2 in pigmentation, and highlight analogies in the generation of PMEL-derived functional amyloids and APP-derived pathological amyloids.
Project description:Melanocytes of the hair follicle produce melanin and are essential in determining the differences in hair color. Pigment cell-specific MELanocyte Protein (PMEL17) plays a crucial role in melanogenesis. One of the critical steps is the amyloid-like functional oligomerization of PMEL17. Beta Site APP Cleaving Enzyme-2 (BACE2) and ?-secretase have been shown to be key players in generating the proteolytic fragments of PMEL17. The ?-secretase (BACE1) is responsible for the generation of amyloid-? (A?) fragments in the brain and is therefore proposed as a therapeutic target for Alzheimer's disease (AD). Currently BACE1 inhibitors, most of which lack selectivity over BACE2, have demonstrated efficacious reduction of amyloid-? peptides in animals and the CSF of humans. BACE2 knock-out mice have a deficiency in PMEL17 proteolytic processing leading to impaired melanin storage and hair depigmentation. Here, we confirm BACE2-mediated inhibition of PMEL17 proteolytic processing in vitro in mouse and human melanocytes. Furthermore, we show that wildtype as well as bace2(+/-) and bace2(-/-) mice treated with a potent dual BACE1/BACE2 inhibitor NB-360 display dose-dependent appearance of irreversibly depigmented hair. Retinal pigmented epithelium showed no morphological changes. Our data demonstrates that BACE2 as well as additional BACE1 inhibition affects melanosome maturation and induces hair depigmentation in mice.
Project description:It was recently suggested that beta-site amyloid precursor protein (APP)-cleaving enzyme 2 (BACE2) functions as an amyloid beta (A?)-degrading enzyme; in addition to its better understood role as an APP secretase. Due to this finding we sought to understand the possible genetic risk contributed by the BACE2 locus to the development of late-onset Alzheimer's disease (AD). In this study, we report that common single nucleotide polymorphism (SNP) variation in BACE2 is associated with altered AD risk in apolipoprotein E gene (APOE) epsilon 4 variant (?4) non-carriers. In addition, in ?4 non-carriers diagnosed with AD or mild cognitive impairment (MCI), SNPs within the BACE2 locus are associated with cerebrospinal fluid (CSF) levels of A?1-42. Further, SNP variants in BACE2 are also associated with BACE2 RNA expression levels suggesting a potential mechanism for the CSF A?1-42 findings. Lastly, overexpression of BACE2 in vitro resulted in decreased A?1-40 and A?1-42 fragments in a cell line model of A? production. These findings suggest that genetic variation at the BACE2 locus modifies AD risk for those individuals who don't carry the ?4 variant of APOE. Further, our data indicate that the biological mechanism associated with this altered risk is linked to amyloid generation or clearance possibly through BACE2 expression changes.
Project description:Gliomas are the most common primary malignant tumours of the central nervous system, and new molecular biomarkers are urgently needed for diagnosis and targeted therapy. Here, we report that increased beta-site APP-cleaving enzyme 2 (BACE2) expression is associated with increases in the grade of human glioma, the incidence of the mesenchymal molecular glioblastoma multiforme subtype and the likelihood of poor prognoses for patients. BACE2 knockdown suppressed cell invasion, cell migration and tumour growth both in vitro and in vivo, while BACE2 overexpression promoted the mesenchymal transition and cell proliferation. Furthermore, TGF?1 stimulated BACE2 expression through Smad-dependent signalling, which modulated TNF-?-induced NF-?B activity through the PP1A/IKK pathway to promote tumorigenesis in both U87MG and U251 cells. Our study indicated that BACE2 plays a significant role in glioma development. Therefore, BACE2 is a potential therapeutic target for human gliomas due to its function and ability to be regulated.