共查询到20条相似文献,搜索用时 15 毫秒
1.
Edmar Zanoteli Diantha van de Vlekkert Erik J. Bonten Huimin Hu Linda Mann Elida M. Gomero A. John Harris Giulio Ghersi Alessandra d'Azzo 《生物化学与生物物理学报:疾病的分子基础》2010,1802(7-8):659-672
Neuraminidase 1 (NEU1) regulates the catabolism of sialoglycoconjugates in lysosomes. Congenital NEU1 deficiency in children is the basis of sialidosis, a severe neurosomatic disorder in which patients experience a broad spectrum of clinical manifestations varying in the age of onset and severity. Osteoskeletal deformities and muscle hypotonia have been described in patients with sialidosis. Here we present the first comprehensive analysis of the skeletal muscle pathology associated with loss of Neu1 function in mice. In this animal model, skeletal muscles showed an expansion of the epimysial and perimysial spaces, associated with proliferation of fibroblast-like cells and abnormal deposition of collagens. Muscle fibers located adjacent to the expanded connective tissue underwent extensive invagination of their sarcolemma, which resulted in the infiltration of the fibers by fibroblast-like cells and extracellular matrix, and in their progressive cytosolic fragmentation. Both the expanded connective tissue and the juxtaposed infiltrated muscle fibers were strongly positive for lysosomal markers and displayed increased proteolytic activity of lysosomal cathepsins and metalloproteinases. These combined features could lead to abnormal remodeling of the extracellular matrix that could be responsible for sarcolemmal invagination and progressive muscle fiber degeneration, ultimately resulting in an overt atrophic phenotype. This unique pattern of muscle damage, which has never been described in any myopathy, might explain the neuromuscular manifestations reported in patients with the type II severe form of sialidosis. More broadly, these findings point to a potential role of NEU1 in cell proliferation and extracellular matrix remodeling. 相似文献
2.
Erik J. Bonten Gouri Yogalingam Huimin Hu Elida Gomero Diantha van de Vlekkert Alessandra d'Azzo 《生物化学与生物物理学报:疾病的分子基础》2013,1832(10):1784-1792
The lysosomal storage disease sialidosis is caused by a primary deficiency of the sialidase N-acetyl-α-neuraminidase-1 (NEU1). Patients with type I sialidosis develop an attenuated, non-neuropathic form of the disease also named cherry red spot myoclonus syndrome, with symptoms arising during juvenile/ adult age. NEU1 requires binding to its chaperone, protective protein/cathepsin A (PPCA), for lysosomal compartmentalization, stability and catalytic activation. We have generated a new mouse model of type I sialidosis that ubiquitously expresses a NEU1 variant carrying a V54M amino acid substitution identified in an adult patient with type I sialidosis. Mutant mice developed signs of lysosomal disease after 1 year of age, predominantly in the kidney, albeit low residual NEU1 activity was detected in most organs and cell types. We demonstrate that the activity of the mutant enzyme could be effectively increased in all systemic tissues by chaperone-mediated gene therapy with a liver-tropic recombinant AAV2/8 vector expressing PPCA. This resulted in clear amelioration of the disease phenotype. These results suggest that at least some of the NEU1 mutations associated with type I sialidosis may respond to PPCA-chaperone-mediated gene therapy. 相似文献
3.
Oheda Y Kotani M Murata M Sakuraba H Kadota Y Tatano Y Kuwahara J Itoh K 《Glycobiology》2006,16(4):271-280
Sialidosis and galactosialidosis are lysosomal storage diseases caused by the genetic defects of lysosomal sialidase (neuraminidase-1; NEU1) and lysosomal protective protein/cathepsin A (PPCA), respectively, associated with a NEU1 deficiency, excessive accumulation of sialylglycoconjugates, and development of progressive neurosomatic manifestations; in addition, the latter disorder is accompanied by simultaneous deficiencies of beta-galactosidase and cathepsin A. We demonstrated that a few soluble N-glycosylated proteins carrying sialyloligosaccharides sensitive to glycopeptidase F (GPF) can be specifically detected in cultured fibroblasts from sialidosis and galactosialidosis cases by blotting with a Maackia amurensis (MAM) lectin. We also examined the therapeutic effects of normal gene transfer and enzyme replacement by evaluating the decreases in sialylglycoconjugates accumulated in fibroblasts with these NEU1 deficiencies. The specific N-glycosylated proteins detected on MAM lectin blotting as well as the granular lysosomal fluorescence due to an avidin-FITC/biotinylated MAM lectin conjugate in sialidosis and galactosialidosis fibroblasts disappeared in parallel with the restoration of the intracellular NEU1 activity after transfection of the recombinant NEU1 fused to HA tag sequence and the wild-type PPCA cDNA as well as administration of the recombinant PPCA precursor protein. The detection method for the abnormal sialylglycoproteins in cultured cells involving MAM lectin was demonstrated to be useful not only for biochemical and diagnostic analyses of NEU1 deficiencies but also for therapeutic evaluation of these conditions. 相似文献
4.
Neuraminidase 1 is a negative regulator of lysosomal exocytosis 总被引:1,自引:0,他引:1
Yogalingam G Bonten EJ van de Vlekkert D Hu H Moshiach S Connell SA d'Azzo A 《Developmental cell》2008,15(1):74-86
Lysosomal exocytosis is a Ca2+-regulated mechanism that involves proteins responsible for cytoskeletal attachment and fusion of lysosomes with the plasma membrane. However, whether luminal lysosomal enzymes contribute to this process remains unknown. Here we show that neuraminidase NEU1 negatively regulates lysosomal exocytosis in hematopoietic cells by processing the sialic acids on the lysosomal membrane protein LAMP-1. In macrophages from NEU1-deficient mice, a model of the disease sialidosis, and in patients' fibroblasts, oversialylated LAMP-1 enhances lysosomal exocytosis. Silencing of LAMP-1 reverts this phenotype by interfering with the docking of lysosomes at the plasma membrane. In neu1-/- mice the excessive exocytosis of serine proteases in the bone niche leads to inactivation of extracellular serpins, premature degradation of VCAM-1, and loss of bone marrow retention. Our findings uncover an unexpected mechanism influencing lysosomal exocytosis and argue that exacerbations of this process form the basis for certain genetic diseases. 相似文献
5.
Dario Bonardi Viola Ravasio Giuseppe Borsani Alessandra d'Azzo Roberto Bresciani Eugenio Monti Edoardo Giacopuzzi 《PloS one》2014,9(8)
The NEU1 gene is the first identified member of the human sialidases, glycohydrolitic enzymes that remove the terminal sialic acid from oligosaccharide chains. Mutations in NEU1 gene are causative of sialidosis (MIM 256550), a severe lysosomal storage disorder showing autosomal recessive mode of inheritance. Sialidosis has been classified into two subtypes: sialidosis type I, a normomorphic, late-onset form, and sialidosis type II, a more severe neonatal or early-onset form. A total of 50 causative mutations are reported in HGMD database, most of which are missense variants. To further characterize the NEU1 gene and identify new functionally relevant protein isoforms, we decided to study its genetic variability in the human population using the data generated by two large sequencing projects: the 1000 Genomes Project (1000G) and the NHLBI GO Exome Sequencing Project (ESP). Together these two datasets comprise a cohort of 7595 sequenced individuals, making it possible to identify rare variants and dissect population specific ones. By integrating this approach with biochemical and cellular studies, we were able to identify new rare missense and frameshift alleles in NEU1 gene. Among the 9 candidate variants tested, only two resulted in significantly lower levels of sialidase activity (p<0.05), namely c.650T>C and c.700G>A. These two mutations give rise to the amino acid substitutions p.V217A and p.D234N, respectively. NEU1 variants including either of these two amino acid changes have 44% and 25% residual sialidase activity when compared to the wild-type enzyme, reduced protein levels and altered subcellular localization. Thus they may represent new, putative pathological mutations resulting in sialidosis type I. The in silico approach used in this study has enabled the identification of previously unknown NEU1 functional alleles that are widespread in the population and could be tested in future functional studies. 相似文献
6.
Dongning Wang Slava Zaitsev Garry Taylor Alessandra d'Azzo Erik Bonten 《Biochimica et Biophysica Acta (BBA)/General Subjects》2009
Background
Neuraminidase-1 (NEU1) catabolizes the hydrolysis of sialic acids from sialo-glycoconjugates. NEU1 depends on its interaction with the protective protein/cathepsin A (PPCA) for lysosomal compartmentalization and catalytic activation. Murine NEU1 contains 4 N-glycosylation sites, 3 of which are conserved in the human enzyme. The expression of NEU1 gives rise to differentially glycosylated proteins.Methods
We generated single-point mutations in mouse NEU1 at each of the 4 N-glycosylation sites. Mutant enzymes were expressed in NEU1-deficient cells in the presence and absence of PPCA.Results
All 4 N-glycosylation variants were targeted to the lysosomal/endosomal compartment. All N-glycans, with the exception of the most C-terminal glycan, were important for maintaining stability or catalytic activity. The loss of catalytic activity caused by the deletion of the second N-glycan was rescued by increasing PPCA expression. Similar results were obtained with a human NEU1 N-glycosylation mutant identified in a sialidosis patient. The N-terminal N-glycan of NEU1 is indispensable for its function, whereas the C-terminal N-glycan appears to be non-essential. The omission of the second N-glycan can be compensated for by upregulating the expression of PPCA.General significance
These findings could be relevant for the design of target therapies for patients carrying specific NEU1 mutations. 相似文献7.
8.
Takamichi Yamada Shoji Tsuji Toshio Ariga Tadashi Miyatake 《Biochimica et Biophysica Acta (BBA)/General Subjects》1983,755(1):106-111
We analyzed the subcellular localization of sialidases in human lymphocytes from a patient with adult type sialidosis with partial β-galactosidase deficiency and normal controls. Sialidase activities were measured with α,2 → 3 NeuAc-lactitol, 4-methylumbelliferyl-NeuAc and GM3 ganglioside as substrates. Sialidases in the lysosomes were sonication-labile and hydrolyzed mainly hydrophilic substrates such as NeuAc-lactitol and 4-methylumbelliferyl-NeuAc, but hydrolyzed subsidiarily GM3 ganglioside. On the other hand, sialidases in the plasma membrane were sonication-stable and hydrolyzed both hydrophilic substrates and GM3 ganglioside. In sialidosis with partial β-galactosidase deficiency, the sialidases of the lysosomes showed 3–5% activity toward hydrophilic substrates and 25% activity toward GM3 ganglioside as compared with sialidase activities of the controls. However, there are no differences in the activities of the sialidases in the plasma membrane. These results demonstrate that the essential defect in this disease is the deficiency of a lysosomal sialidase. 相似文献
9.
Heterodimerization of the Sialidase NEU1 with the Chaperone Protective Protein/Cathepsin A Prevents Its Premature Oligomerization 总被引:1,自引:0,他引:1
Erik J. Bonten Yvan Campos Viateslav Zaitsev Amanda Nourse Brett Waddell William Lewis Garry Taylor Alessandra d'Azzo 《The Journal of biological chemistry》2009,284(41):28430-28441
Lysosomal neuraminidase-1 (NEU1) forms a multienzyme complex with β-galactosidase and protective protein/cathepsin A (PPCA). Because of its association with PPCA, which acts as a molecular chaperone, NEU1 is transported to the lysosomal compartment, catalytically activated, and stabilized. However, the mode(s) of association between these two proteins both en route to the lysosome and in the multienzyme complex has remained elusive. Here, we have analyzed the hydrodynamic properties of PPCA, NEU1, and a complex of the two proteins and identified multiple binding sites on both proteins. One of these sites on NEU1 that is involved in binding to PPCA can also bind to other NEU1 molecules, albeit with lower affinity. Therefore, in the absence of PPCA, as in the lysosomal storage disease galactosialidosis, NEU1 self-associates into chain-like oligomers. Binding of PPCA can reverse self-association of NEU1 by causing the disassembly of NEU1-oligomers and the formation of a PPCA-NEU1 heterodimeric complex. The identification of binding sites between the two proteins allowed us to create innovative structural models of the NEU1 oligomer and the PPCA-NEU1 heterodimeric complex. The proposed mechanism of interaction between NEU1 and its accessory protein PPCA provides a rationale for the secondary deficiency of NEU1 in galactosialidosis.Mammalian neuraminidases have been classified as lysosomal (NEU1),4 cytosolic (NEU2), plasma membrane (NEU3), and mitochondria/lysosomal (NEU4) based on their subcellular distributions, pH optimum, kinetic properties, responses to ions and detergents, and substrate specificities (1–3). Of the four sialidases, only NEU1 is ubiquitously expressed at different levels in various tissues and cell types (4–7). The importance of these proteins in normal cellular physiology is illustrated by the numerous metabolic processes that they control, including cell proliferation and differentiation, cell adhesion, membrane fusion and fluidity, immunocyte function, and receptor modification (8–21).NEU1 initiates the intralysosomal hydrolysis of sialo-oligosaccharides, -glycolipids, and -glycoproteins by removing their terminal sialic acid residues. In human and murine tissues, NEU1 forms a complex with at least two other proteins, β-galactosidase and the protective protein/cathepsin A (PPCA) (22). By virtue of their association with PPCA, NEU1 and β-galactosidase acquire their active and stable conformation in lysosomes. However, PPCA appears to function as a crucial chaperone/transport protein for NEU1. Because NEU1 is poorly mannose 6-phosphorylated, it depends on PPCA for correct compartmentalization and catalytic activation in lysosomes (23–25). Only a small amount of PPCA and β-galactosidase activities is found in the NEU1-PPCA-β-galactosidase complex, which instead contains all of the NEU1 catalytic activity (24–27). By understanding how and when NEU1 and PPCA interact, how they regulate each other in different cell types, and what determinants control their association, we may gain important insight into their significance in physiologic and pathologic conditions.The absence of NEU1 is associated with two neurodegenerative diseases that involve glycoprotein metabolism; sialidosis, which is caused by structural lesions in the lysosomal NEU1 locus (28), and galactosialidosis (GS), a combined deficiency of NEU1 and β-galactosidase which is caused by the absence of PPCA (22). Patients with sialidosis and those with GS have similar clinical and biochemical features, and both diseases are characterized by multiple phenotypes that are classified according to the age of onset and severity of the symptoms.Previously, we generated two animal models of primary or secondary NEU1 deficiency, Neu1−/− mice and Ppca−/− mice. Both mouse models have a profound loss of Neu1 activity in multiple tissues and develop clinical, biochemical, and pathologic manifestations resembling those seen in patients with severe sialidosis and GS (29–31). Neu1−/− mice are phenotypically similar but not identical to Ppca−/− mice and, like children with the disease, exhibit a time-dependent splenomegaly associated with extramedullary hematopoiesis (30, 31). We found that the cause of these phenotypic abnormalities is the gradual loss of retention of hematopoietic progenitors within the bone niche due to exacerbated lysosomal exocytosis of bone marrow cells. The latter process is negatively regulated by NEU1 activity (31).The mode of interaction between PPCA and NEU1 and the mechanism of catalytic activation are not well understood. Here we present biochemical, analytical, and structural analyses of NEU1, PPCA, and the PPCA-NEU1 complex by using purified baculovirus (BV)-expressed wild-type and mutagenized recombinant enzymes and synthetic peptides. 相似文献
10.
New insights on the sialidase protein family revealed by a phylogenetic analysis in metazoa 总被引:1,自引:0,他引:1
Sialidases are glycohydrolytic enzymes present from virus to mammals that remove sialic acid from oligosaccharide chains. Four different sialidase forms are known in vertebrates: the lysosomal NEU1, the cytosolic NEU2 and the membrane-associated NEU3 and NEU4. These enzymes modulate the cell sialic acid content and are involved in several cellular processes and pathological conditions. Molecular defects in NEU1 are responsible for sialidosis, an inherited disease characterized by lysosomal storage disorder and neurodegeneration. The studies on the biology of sialic acids and sialyltransferases, the anabolic counterparts of sialidases, have revealed a complex picture with more than 50 sialic acid variants selectively present in the different branches of the tree of life. The gain/loss of specific sialoconjugates have been proposed as key events in the evolution of deuterostomes and Homo sapiens, as well as in the host-pathogen interactions. To date, less attention has been paid to the evolution of sialidases. Thus we have conducted a survey on the state of the sialidase family in metazoan. Using an in silico approach, we identified and characterized sialidase orthologs from 21 different organisms distributed among the evolutionary tree: Metazoa relative (Monosiga brevicollis), early Deuterostomia, precursor of Chordata and Vertebrata (teleost fishes, amphibians, reptiles, avians and early and recent mammals). We were able to reconstruct the evolution of the sialidase protein family from the ancestral sialidase NEU1 and identify a new form of the enzyme, NEU5, representing an intermediate step in the evolution leading to the modern NEU3, NEU4 and NEU2. Our study provides new insights on the mechanisms that shaped the substrate specificity and other peculiar properties of the modern mammalian sialidases. Moreover, we further confirm findings on the catalytic residues and identified enzyme loop portions that behave as rapidly diverging regions and may be involved in the evolution of specific properties of sialidases. 相似文献
11.
Liang F Seyrantepe V Landry K Ahmad R Ahmad A Stamatos NM Pshezhetsky AV 《The Journal of biological chemistry》2006,281(37):27526-27538
12.
Molecular mechanisms of pathogenesis in a glycosphingolipid and a glycoprotein storage disease 总被引:1,自引:0,他引:1
The lysosomal system comprises a specialized network of organelles crucial for the sorting, digestion, recycling and secretion of cellular components. With their content of hydrolytic enzymes, lysosomes regulate the degradation of a multitude of substrates that reach these organelles via the biosynthetic or the endocytic route. Gene defects that affect one or more of these hydrolases lead to LSDs (lysosomal storage diseases). This underscores the apparent lack of redundancy of these enzymes and the importance of the lysosomal system in cell and tissue homoeostasis. Some of the lysosomal enzymes may form multiprotein complexes, which usually work synergistically on substrates and, in this configuration, may respond more efficiently to changes in substrate load and composition. A well-characterized lysosomal multienzyme complex is the one comprising the glycosidases β-gal (β-galactosidase) and NEU1 (neuramidase-1), and of the serine carboxypeptidase PPCA (protective protein/cathepsin A). Three neurodegenerative LSDs are caused by either single or combined deficiency of these lysosomal enzymes. Sialidosis (NEU1 deficiency) and galactosialidosis (combined NEU1 and β-gal deficiency, secondary to a primary defect of PPCA) belong to the glycoprotein storage diseases, whereas GM1-gangliosidosis (β-gal deficiency) is a glycosphingolipid storage disease. Identification of novel molecular pathways that are deregulated because of loss of enzyme activity and/or accumulation of specific metabolites in various cell types has shed light on mechanisms of disease pathogenesis and may pave the way for future development of new therapies for these LSDs. 相似文献
13.
14.
Lukong KE Seyrantepe V Landry K Trudel S Ahmad A Gahl WA Lefrancois S Morales CR Pshezhetsky AV 《The Journal of biological chemistry》2001,276(49):46172-46181
Sialidase (neuraminidase), encoded by the neu-1 gene in the major histocompatibility complex locus catalyzes the intralysosomal degradation of sialylated glycoconjugates. Inherited deficiency of sialidase results in sialidosis or galactosialidosis, both severe metabolic disorders associated with lysosomal storage of oligosaccharides and glycopeptides. Sialidase also plays an important role in cellular signaling and is specifically required for the production of cytokine interleukin-4 by activated T lymphocytes. In these cells, neu-1-encoded sialidase activity is increased on the cell surface, suggesting that a specific mechanism regulates sorting of this enzyme to the plasma membrane. We investigated that mechanism by first showing that sialidase contains the internalization signal found in lysosomal membrane proteins targeted to endosomes via clathrin-coated pits. The signal consists of a C-terminal tetrapeptide (412)YGTL(415), with Tyr(412) and Leu(415) essential for endocytosis of the enzyme. We further demonstrated that redistribution of sialidase from lysosomes to the cell surface of activated lymphocytes is accompanied by increased reactivity of the enzyme with anti-phosphotyrosine antibodies. We speculate that phosphorylation of Tyr(412) results in inhibition of sialidase internalization in activated lymphocytes. 相似文献
15.
Li Y Cao H Yu H Chen Y Lau K Qu J Thon V Sugiarto G Chen X 《Molecular bioSystems》2011,7(4):1060-1072
Aberrant expression of human sialidases has been shown to associate with various pathological conditions. Despite the effort in the sialidase inhibitor design, less attention has been paid to designing specific inhibitors against human sialidases and characterizing the substrate specificity of different sialidases regarding diverse terminal sialic acid forms and sialyl linkages. This is mainly due to the lack of sialoside probes and efficient screening methods, as well as limited access to human sialidases. A low cellular expression level of the human sialidase NEU2 hampers its functional and inhibitory studies. Here we report the successful cloning and expression of the human sialidase NEU2 in E. coli. About 11 mg of soluble active NEU2 was routinely obtained from 1 L of E. coli cell culture. Substrate specificity studies of the recombinant human NEU2 using twenty p-nitrophenol (pNP)-tagged α2-3- or α2-6-linked sialyl galactosides containing different terminal sialic acid forms including common N-acetylneuraminic acid (Neu5Ac), non-human N-glycolylneuraminic acid (Neu5Gc), 2-keto-3-deoxy-D-glycero-D-galacto-nonulosonic acid (Kdn), or their C5-derivatives in a microtiter plate-based high-throughput colorimetric assay identified a unique structural feature specifically recognized by the human NEU2 but not two bacterial sialidases. The results obtained from substrate specificity studies were used to guide the design of a sialidase inhibitor that was selective against human NEU2. The selectivity of the inhibitor was revealed by the comparison of sialidase crystal structures and inhibitor docking studies. 相似文献
16.
Mammalian Neu3 is a ganglioside specific sialidase. Gangliosides are involved in various physiological events such as cell growth, differentiation and diseases. Significance of Neu3 and gangliosides is still unclear in aquaculture fish species. To gain more insights of fish Neu3 sialidases, molecular cloning and characterization were carried out in tilapia (Oreochromis niloticus). A tilapia genome-wide search for orthologues of human NEU1, NEU2, NEU3 and NEU4 yielded eight putative tilapia sialidases, five of which were neu3-like and designated as neu3a, neu3b, neu3c, neu3d and neu3e. Among five neu3 genes, neu3a, neu3d and neu3e were amplified by PCR from adult fish brain cDNA with consensus sequences of 1227 bp, 1194 bp and 1155 bp, respectively. Multiple alignments showed conserved three Asp-boxes (SXDXGXTW), YRIP and VGPG motifs. The molecular weights for Neu3a, Neu3d and Neu3e were confirmed using immunoblotting analysis as 45.9 kDa, 44.4 kDa and 43.6 kDa, respectively. Lysate from neu3 genes transfected HEK293 cells showed sialidase activity in Neu3a towards ganglioside mix optimally at pH 4.6. Using pure gangliosides as substrates, highest sialidase activity for Neu3a was observed towards GD3 followed by GD1a and GM3, but not GM1. On the other hand, sialidase activities were not observed in Neu3d and Neu3e towards various sialoglycoconjugates. Indirect immunofluorescence showed that tilapia Neu3a and Neu3d are localized at the plasma membrane, while most Neu3e showed a cytosolic localization. RT-PCR analyses for neu3a showed significant expression in the brain, liver, and spleen tissues, while neu3d and neu3e showed different expression patterns. Based on these results, tilapia Neu3 exploration is an important step towards full understanding of a more comprehensive picture of Neu3 sub-family of proteins in fish. 相似文献
17.
Lysosomal and plasma membrane ganglioside GM3 sialidases of cultured human fibroblasts. Differentiation by detergents and inhibitors 总被引:2,自引:0,他引:2
Cultured human fibroblasts contain two sialidases that degrade gangliosides such as GM3: a lysosomal activity that appears identical with the activity towards water-soluble substrates and that is deficient in the genetic lysosomal disorder sialidosis, and another enzyme that seems localized on the external surface of the plasma membrane. In this report we show that both enzymes can be differentiated in the presence of each other by choice of the detergent used for activation, and also by the inhibitory action of some polyanionic compounds such as sulphated glycosaminoglycans. The lysosomal ganglioside GM3 sialidase is greatly stimulated by sodium glycodeoxycholate and, to lesser degrees, by sodium glycocholate and sodium cholate. The ganglioside GM3 sialidase of the plasma membrane is not measurably active under the conditions of the lysosomal enzyme but is specifically activated by the non-ionic detergent Triton X-100. The glycodeoxycholate-stimulated, but not the Triton-activated, ganglioside GM3 sialidase activity was profoundly diminished in cell lines from patients with the lysosomal disorders sialidosis and galactosialidosis; however, both activities were normal in fibroblasts from patients with mucolipidosis IV, previously thought to be a ganglioside sialidase deficiency disorder. Both the lysosomal and the plasma membrane ganglioside GM3 sialidases were inhibited by sialic acids, suramin, dextran sulphate and sulphated glycosaminoglycans. Among the latter, heparin and heparan sulphate showed a much higher inhibitory potency towards the plasma membrane ganglioside GM3 sialidase than towards the lysosomal onw.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
18.
19.
《Cellular signalling》2014,26(6):1355-1368
Insulin-induced insulin receptor (IR) tyrosine kinase activation and insulin cell survival responses have been reported to be under the regulation of a membrane associated mammalian neuraminidase-1 (Neu1). The molecular mechanism(s) behind this process is unknown. Here, we uncover a novel Neu1 and matrix metalloproteinase-9 (MMP-9) cross-talk in alliance with neuromedin B G-protein coupled receptor (GPCR), which is essential for insulin-induced IR activation and cellular signaling. Neu1, MMP-9 and neuromedin B GPCR form a complex with IRβ subunit on the cell surface. Oseltamivir phosphate (Tamiflu®), anti-Neu1 antibodies, broad range MMP inhibitors piperazine and galardin (GM6001), MMP-9 specific inhibitor (MMP-9i), and GPCR neuromedin B specific antagonist BIM-23127 dose-dependently inhibited Neu1 activity associated with insulin stimulated rat hepatoma cells (HTCs) that overly express human IRs (HTC-IR). Tamiflu, anti-Neu1 antibodies and MMP-9i attenuated phosphorylation of IRβ and insulin receptor substrate-1 (IRS1) associated with insulin-stimulated cells. Olanzapine, an antipsychotic agent associated with insulin resistance, induced Neu3 sialidase activity in WG544 or 1140F01 human sialidosis fibroblast cells genetically defective in Neu1. Neu3 antagonist 2-deoxy-2,3-didehydro-N-acetylneuraminic acid (DANA) and anti-Neu3 antibodies inhibited sialidase activity associated with olanzapine treated murine Neu4 knockout macrophage cells. Olanzapine attenuated phosphorylation of IGF-R and IRS1 associated with insulin-stimulated human wild-type fibroblast cells. Our findings identify a novel insulin receptor-signaling platform that is critically essential for insulin-induced IRβ tyrosine kinase activation and cellular signaling. Olanzapine-induced Neu3 sialidase activity attenuated insulin-induced IGF-R and IRS1 phosphorylation contributing to insulin resistance. 相似文献
20.
N-Glycolylneuraminic acid (Neu5Gc) is a widely expressed sialic acid in mammalian cells. Although humans are genetically deficient in producing Neu5Gc, small amounts are present in human cells in vivo. A dietary origin was suggested by human volunteer studies and by observing that free Neu5Gc is metabolically incorporated into cultured human carcinoma cells by unknown mechanisms. We now show that free Neu5Gc uptake also occurs in other human and mammalian cells. Inhibitors of certain non-clathrin-mediated endocytic pathways reduce Neu5Gc accumulation. Studies with human mutant cells show that the lysosomal sialic acid transporter is required for metabolic incorporation of free Neu5Gc. Incorporation of glycosidically bound Neu5Gc from exogenous glycoconjugates (relevant to human gut epithelial exposure to dietary Neu5Gc) requires the transporter as well as the lysosomal sialidase, which presumably acts to release free Neu5Gc. Thus, exogenous Neu5Gc reaches lysosomes via pinocytic/endocytic pathways and is exported in free form into the cytosol, becoming available for activation and transfer to glycoconjugates. In contrast, N-glycolylmannosamine (ManNGc) apparently traverses the plasma membrane by passive diffusion and becomes available for conversion to Neu5Gc in the cytosol. This mechanism can also explain the metabolic incorporation of chemically synthesized unnatural sialic acids, as reported by others. Finally, to our knowledge, this is the first example of delivery to the cytosol of an extracellular small molecule that cannot cross the plasma membrane, utilizing fluid pinocytosis and a specific lysosomal transporter. The approach could, thus, potentially be generalized to any small molecule that has a specific lysosomal transporter but not a plasma membrane transporter. 相似文献