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1.
Nat Commun ; 11(1): 1620, 2020 03 27.
Artigo em Inglês | MEDLINE | ID: mdl-32221306

RESUMO

Since 2002, beta coronaviruses (CoV) have caused three zoonotic outbreaks, SARS-CoV in 2002-2003, MERS-CoV in 2012, and the newly emerged SARS-CoV-2 in late 2019. However, little is currently known about the biology of SARS-CoV-2. Here, using SARS-CoV-2 S protein pseudovirus system, we confirm that human angiotensin converting enzyme 2 (hACE2) is the receptor for SARS-CoV-2, find that SARS-CoV-2 enters 293/hACE2 cells mainly through endocytosis, that PIKfyve, TPC2, and cathepsin L are critical for entry, and that SARS-CoV-2 S protein is less stable than SARS-CoV S. Polyclonal anti-SARS S1 antibodies T62 inhibit entry of SARS-CoV S but not SARS-CoV-2 S pseudovirions. Further studies using recovered SARS and COVID-19 patients' sera show limited cross-neutralization, suggesting that recovery from one infection might not protect against the other. Our results present potential targets for development of drugs and vaccines for SARS-CoV-2.


Assuntos
Anticorpos Antivirais/imunologia , Betacoronavirus/fisiologia , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Internalização do Vírus , Betacoronavirus/química , Betacoronavirus/imunologia , Canais de Cálcio/metabolismo , Catepsina L/metabolismo , Catepsinas/antagonistas & inibidores , Catepsinas/metabolismo , Fusão Celular , Infecções por Coronavirus/imunologia , Reações Cruzadas , Endocitose , Células Gigantes/fisiologia , Células HEK293 , Humanos , Testes de Neutralização , Pandemias , Peptidil Dipeptidase A/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Pneumonia Viral/imunologia , Domínios Proteicos , Multimerização Proteica , Receptores Virais/metabolismo , Vírus da SARS/imunologia , Síndrome Respiratória Aguda Grave/imunologia , Glicoproteína da Espícula de Coronavírus/química , Tripsina/metabolismo
2.
PLoS One ; 14(12): e0222055, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31856175

RESUMO

Cruzain, a cysteine protease of Trypanosoma cruzi, is a validated target for the treatment of Chagas disease. Due to its high similarity in three-dimensional structure with human cathepsins and their sequence identity above 70% in the active site regions, identifying potent but selective cruzain inhibitors with low side effects on the host organism represents a significant challenge. Here a panel of nitrile ligands with varying potencies against cathepsin K, cathepsin L and cruzain, are studied by molecular dynamics simulations as both non-covalent and covalent complexes. Principal component analysis (PCA), identifies and quantifies patterns of ligand-induced conformational selection that enable the construction of a decision tree which can predict with high confidence a low-nanomolar inhibitor of each of three proteins, and determine the selectivity for one against others.


Assuntos
Inibidores de Cisteína Proteinase/metabolismo , Inibidores de Cisteína Proteinase/farmacologia , Proteínas de Protozoários/antagonistas & inibidores , Animais , Domínio Catalítico , Catepsina L/metabolismo , Catepsinas/antagonistas & inibidores , Catepsinas/metabolismo , Doença de Chagas , Cisteína Endopeptidases/metabolismo , Cisteína Proteases/metabolismo , Humanos , Ligantes , Simulação de Dinâmica Molecular , Análise de Componente Principal/métodos , Ligação Proteica , Proteínas de Protozoários/metabolismo , Relação Estrutura-Atividade , Trypanosoma cruzi/metabolismo
3.
Int J Mol Sci ; 20(19)2019 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-31569504

RESUMO

Cysteine cathepsins are critical components of the adaptive immune system involved in the generation of epitopes for presentation on human leukocyte antigen (HLA) molecules and have been implicated in degradation of autoantigens. Immunoglobulin variable regions with somatic mutations and random complementarity region 3 amino acid composition are inherently immunogenic. T cell reactivity towards immunoglobulin variable regions has been investigated in relation to specific diseases, as well as reactivity to therapeutic monoclonal antibodies. Yet, how the immunoglobulins, or the B cell receptors, are processed in endolysosomal compartments of professional antigen presenting cells has not been described in detail. Here we present in silico and in vitro experimental evidence suggesting that cysteine cathepsins S, L and B may have important roles in generating peptides fitting HLA class II molecules, capable of being presented to T cells, from monoclonal antibodies as well as from central nervous system proteins including a well described autoantigen. By combining neural net models with in vitro proteomics experiments, we further suggest how such degradation can be predicted, how it fits with available cellular models, and that it is immunoglobulin heavy chain variable family dependent. These findings are relevant for biotherapeutic drug design as well as to understand disease development. We also suggest how these tools can be improved, including improved machine learning methodology.


Assuntos
Catepsinas/química , Catepsinas/metabolismo , Cisteína/metabolismo , Imunoglobulina G/química , Imunoglobulina G/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Imunoglobulina G/genética , Cadeias Pesadas de Imunoglobulinas/química , Cadeias Pesadas de Imunoglobulinas/genética , Cadeias Pesadas de Imunoglobulinas/metabolismo , Conformação Molecular , Ligação Proteica , Proteólise , Reprodutibilidade dos Testes , Relação Estrutura-Atividade
4.
Cell Physiol Biochem ; 53(3): 550-572, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31529928

RESUMO

BACKGROUND/AIMS: Atherosclerosis underlies the majority of cardiovascular events, consequent to non-resolving inflammation. Considerable evidence implicates autophagy dysfunction at the core of this inflammatory condition, but the basis of this dysfunction is not fully understood. METHODS: Using an in vitro model of lipid-laden macrophages, activity-based probes and high-throughput techniques, we studied the role of the cysteine proteases cathepsins in autophagy. RESULTS: We showed that cathepsin activity is suppressed by oxidized lipids and that cathepsin has an indispensable role in the autophagy-lysosomal degradation pathway. Accordingly, loss of cathepsin function resulted in autophagy derangement. Shotgun proteomics confirmed autophagy dysfunction and unveiled a pivotal role of cathepsin L in a putative cathepsin degradation network. At the physiological level, cathepsin inhibition resulted in mitochondrial stress, which translated into impaired oxidative metabolism, excessive production of reactive oxygen species and activation of the cellular stress response, driven by ATF4-CHOP transcription factors. In addition, transcriptomic analysis of these cells uncovered some genetic similarities with the inflammatory macrophage phenotype (a.k.a M1 macrophages) and increased expression of inflammatory cytokines. CONCLUSION: Our data highlight the importance of cathepsins for mitochondrial quality control mechanisms and amelioration of vascular inflammation.


Assuntos
Anti-Inflamatórios/farmacologia , Catepsina B/metabolismo , Catepsina L/metabolismo , Catepsinas/metabolismo , Macrófagos/metabolismo , Animais , Autofagia/efeitos dos fármacos , Células da Medula Óssea/citologia , Catepsina B/antagonistas & inibidores , Catepsina L/antagonistas & inibidores , Células Cultivadas , Colesterol/metabolismo , Humanos , Macrófagos/efeitos dos fármacos , Masculino , Espectrometria de Massas , Camundongos , Camundongos Knockout , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Dinâmica Mitocondrial , Estresse Oxidativo/efeitos dos fármacos , Proteômica/métodos , Células RAW 264.7 , Espécies Reativas de Oxigênio/metabolismo
5.
Biochemistry (Mosc) ; 84(7): 746-761, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31509726

RESUMO

Cysteine cathepsins are proteolytic enzymes involved in protein degradation in lysosomes and endosomes. Cysteine cathepsins have been also found in the tumor microenvironment during carcinogenesis, where they are implicated in proliferation, invasion and metastasis of tumor cells through the degradation of extracellular matrix, suppression of cell-cell interactions, and promotion of angiogenesis. In this regard, cathepsins can have a diagnostic value and represent promising targets for antitumor drugs aimed at inhibition of these proteases. Moreover, cysteine cathepsins can be used as activators of novel targeted therapeutic agents. This review summarizes recent discovered roles of cysteine cathepsins in carcinogenesis and discusses new trends in cancer therapy and diagnostics using cysteine cathepsins as markers, targets, or activators.


Assuntos
Catepsinas/metabolismo , Cisteína/metabolismo , Neoplasias/diagnóstico , Neoplasias/tratamento farmacológico , Animais , Biomarcadores Tumorais , Carcinogênese/metabolismo , Catepsinas/antagonistas & inibidores , Catepsinas/classificação , Resistencia a Medicamentos Antineoplásicos , Matriz Extracelular/metabolismo , Transferência Ressonante de Energia de Fluorescência , Humanos , Imunoconjugados/metabolismo , Camundongos , Terapia de Alvo Molecular , Nanopartículas/metabolismo , Neoplasias/metabolismo , Proteólise , Microambiente Tumoral
6.
Ecotoxicol Environ Saf ; 183: 109512, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31398584

RESUMO

Azadirachtin, a botanical insecticide with high potential, has been widely used in pest control. Azadirachtin has shown strong biological activity against Bactrocera dorsalis in toxicological reports, but its mechanism remains unclear. This study finds that azadirachtin A inhibits the growth and development of Bactrocera dorsalis larvae. The larval weights and body sizes of the azadirachtin-treated group were significantly less than those of the control group in a concentration-dependent manner. Further, pathological sections revealed that azadirachtin destroyed the midgut cell structure and intestinal walls, while TUNEL staining showed that azadirachtin could induce apoptosis of midgut cells, and Western blot analysis indicated that Bcl-XL expression was inhibited and cytochrome c (CytC) released into the cytoplasm. The results also imply azadirachtin-induced structural alterations in the Bactrocera dorsalis larvae midgut by activation of apoptosis. RNA-seq analysis of midgut cells found that 482 and 708 unique genes were upregulated and downregulated, respectively. These differentially expressed genes (DEGs) were enriched in apoptotic and lysosomal signaling pathways and included 26 genes of the cathepsin family. qRT-PCR verified the expression patterns of some DEGs, indicating that Cathepsin F was upregulated by 278.47-fold and that Cathepsin L and Cathepsin D were upregulated by 28.06- and 8.97-fold, respectively. Finally, association analysis between DEGs and DEMs (differentially expressed metabolites) revealed that azadirachtin significantly reduced the digestion and absorption of carbohydrates, proteins, fats, vitamins and minerals in the midgut. In conclusion, azadirachtin induces the release of cathepsin from lysosomes, causing apoptosis in the midgut. Ultimately, this leads to reduced digestion and absorption of nutrient metabolites in the midgut and inhibition of the growth and development of Bactrocera dorsalis larvae.


Assuntos
Catepsinas/metabolismo , Inseticidas/toxicidade , Intestinos/efeitos dos fármacos , Larva/efeitos dos fármacos , Limoninas/toxicidade , Tephritidae/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Intestinos/patologia , Larva/crescimento & desenvolvimento , Larva/metabolismo , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Lisossomos/patologia , Transdução de Sinais , Tephritidae/metabolismo
7.
Int J Mol Sci ; 20(16)2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31434216

RESUMO

Adipose tissue is a major endocrine organ capable of secreting adipokines with a role in whole-body metabolism. Changes in the secretome profile during the development of obesity is suspected to contribute to the risk of health complications such as those associated with weight regain after weight loss. However, the number of studies on weight regain is limited and secretome changes during weight regain have hardly been investigated. In an attempt to generate leads for in vivo studies, we have subjected human Simpson Golabi Behmel Syndrome adipocytes to glucose restriction (GR) followed by refeeding (RF) as an in vitro surrogate for weight regain after weight loss. Using LC-MS/MS, we compared the secreted protein profile after GR plus RF with that of normal feeding (NF) to assess the consequences of GR plus RF. We identified 338 secreted proteins of which 49 were described for the first time as being secreted by adipocytes. In addition, comparison between NF and GR plus RF showed 39 differentially secreted proteins. Functional classification revealed GR plus RF-induced changes of enzymes for extracellular matrix modification, complement system factors, cathepsins, and several proteins related to Alzheimer's disease. These observations can be used as clues to investigate metabolic consequences of weight regain, weight cycling or intermittent fasting.


Assuntos
Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Catepsinas/metabolismo , Glucose/farmacologia , Adipocinas/metabolismo , Células Cultivadas , Cromatografia Líquida , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Humanos , Espectrometria de Massas em Tandem
8.
Int J Mol Sci ; 20(17)2019 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-31461977

RESUMO

Vascular calcification can be enhanced by hyperglycemia. Elastin loss in tunica media promotes the osteogenic transformation of smooth muscle cells (SMCs) and involves arterial medial calcification (AMC) that is associated with a high incidence of cardiovascular risk in patients with type 2 diabetes. Here, we tested whether hydrogen sulfide (H2S), an endogenous gaseous mediator, can prevent elastin loss and attenuate calcification induced by high glucose in SMCs. Calcification was induced by high glucose (4500 mg/L) in human aortic SMCs (HASMCs) under the condition of calcifying medium containing 10 mM ß-glycerophosphate (ß-GP). The experiments showed that NaHS (an H2S donor, 100 µM) mitigated the calcification of HASMCs treated with high glucose by decreasing calcium and phosphorus levels, calcium deposition and ALP activity and inhibited osteogenic transformation by increasing SMα-actin and SM22α, two phenotypic markers of smooth muscle cells, and decreasing core binding factor α-1 (Cbfα-1), a key factor in bone formation, protein expressions in HASMCs. Moreover, NaHS administration inhibited the activation of Stat3, cathepsin S (CAS) activity and its expression, but increased the level of elastin protein. Pharmacological inhibition or gene silencing Stat3 not only reversed elastin loss, but also attenuated CAS expression. Inhibition of CAS alleviated, while CAS overexpression exacerbated, elastin loss. Interestingly, overexpression of wild type (WT)-Stat3, but not its mutant C259S, elevated CAS protein expression and reduced elastin level. Moreover, NaHS induced S-sulfhydration in WT, but not in the C259S Stat3. These data suggest that H2S may directly regulate Cys259 residue in Stat3 and then impair its signaling function. Our data indicate that H2S may attenuate vascular calcification by upregulating elastin level through the inhibition of Stat3/CAS signaling.


Assuntos
Catepsinas/metabolismo , Elastina/metabolismo , Sulfeto de Hidrogênio/metabolismo , Miócitos de Músculo Liso/metabolismo , Fator de Transcrição STAT3/metabolismo , Calcificação Vascular/metabolismo , Cálcio/metabolismo , Células Cultivadas , Glucose/metabolismo , Humanos , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/efeitos dos fármacos , Fósforo/metabolismo , Transdução de Sinais , Sulfetos/farmacologia
9.
Int J Mol Sci ; 20(14)2019 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-31340550

RESUMO

Cysteine cathepsins are lysosomal enzymes belonging to the papain family. Their expression is misregulated in a wide variety of tumors, and ample data prove their involvement in cancer progression, angiogenesis, metastasis, and in the occurrence of drug resistance. However, while their overexpression is usually associated with highly aggressive tumor phenotypes, their mechanistic role in cancer progression is still to be determined to develop new therapeutic strategies. In this review, we highlight the literature related to the role of the cysteine cathepsins in cancer biology, with particular emphasis on their input into tumor biology.


Assuntos
Catepsinas/genética , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica , Lisossomos/enzimologia , Neoplasias/genética , Neovascularização Patológica/genética , Animais , Antineoplásicos/uso terapêutico , Catepsinas/química , Catepsinas/classificação , Catepsinas/metabolismo , Morte Celular/efeitos dos fármacos , Morte Celular/genética , Linhagem Celular Tumoral , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Humanos , Metástase Linfática , Lisossomos/efeitos dos fármacos , Modelos Moleculares , Neoplasias/tratamento farmacológico , Neoplasias/enzimologia , Neoplasias/patologia , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/enzimologia , Neovascularização Patológica/patologia , Conformação Proteica , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Transdução de Sinais
10.
Biochimie ; 166: 77-83, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31181234

RESUMO

The cysteine protease legumain (asparaginyl endopeptidase, AEP) plays important roles in normal physiology but is also associated with several disorders, such as atherosclerosis, osteoporosis, cancer and neurodegenerative diseases. The functional roles of legumain have mainly been associated with the presence in lysosomes where legumain is active and mediates processing of multiple proteins, such as the conversion of single to double chain forms of cysteine cathepsins. However, in recent years, a number of studies point to extracellular roles of legumain in addition to the pivotal roles in the lysosomes. In this review, recent knowledge on novel extracellular functions of this protease will be addressed and new discoveries in relation to the diseases mentioned above will be presented.


Assuntos
Aterosclerose/enzimologia , Cisteína Endopeptidases/metabolismo , Lisossomos/enzimologia , Neoplasias/enzimologia , Doenças Neurodegenerativas/enzimologia , Osteoporose/enzimologia , Animais , Biomarcadores/metabolismo , Catepsinas/metabolismo , Humanos , Camundongos
11.
Immunogenetics ; 71(5-6): 421-432, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31089760

RESUMO

Cathepsins are key mammalian proteases that play an important role in the immune response. Several studies have revealed the versatile and critical functions of cathepsins. Here, we obtained ten kinds of cathepsin homologs and identified seven homologs with complete coding sequences. Phylogenetic analysis verified their identities and supported the classification of cathepsins into seven families, which is similar to other vertebrates. Tissue-specific expression analysis showed that all lamprey cathepsins (L-cathepsins) are present in the supraneural body (SB), kidney, gill, intestine, brain, heart, and liver, but their relative abundance varied among tissues. Additionally, we focused on the lamprey cathepsin L (L-cathepsin L) and used recombinant L-cathepsin L protein (rL-cathepsin L) to prepare anti rL-cathepsin L polyclonal antibodies, which were used to detect its distribution in lamprey tissues. The L-cathepsin L protein was primarily detected in the SB, kidney, gill, intestine, brain, and liver via western blot and immunohistochemistry assays. Importantly, quantitative real-time PCR (RT-PCR) revealed that the expression level of L-cathepsins mRNA significantly increased after exposure to three different stimuli (poly I:C, Staphylococcus aureus (S.a) and Vibro anguilarum (V.an)). This suggested that L-cathepsins may participate in defense processes. These results revealed that L-cathepsins may play key roles in the immune response to exogenous stimuli. The findings provide important information for future studies aiming to understand the molecular mechanisms underlying the immune response to pathogen invasion in lamprey.


Assuntos
Catepsinas/genética , Lampreias/genética , Sequência de Aminoácidos , Animais , Catepsinas/química , Catepsinas/metabolismo , Clonagem Molecular , Evolução Molecular , Expressão Gênica , Genômica/métodos , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Imunomodulação , Lampreias/classificação , Lampreias/imunologia , Família Multigênica , Filogenia , Análise de Sequência de DNA
12.
Zhonghua Jie He He Hu Xi Za Zhi ; 42(5): 372-377, 2019 May 12.
Artigo em Chinês | MEDLINE | ID: mdl-31137114

RESUMO

Objective: To explore the value of cathepsin S in the bronchoalveolar lavage fluid (BALF) of patients with chronic obstructive pulmonary disease (COPD) in the evaluation of pulmonary function and CT phenotypes. Method: From April 2014 to April 2017, 46 patients with stable COPD were enrolled, and 29 healthy volunteers served as the control group. The patients were divided into 4 subgroups: GOLD Ⅰ(n=12), GOLD Ⅱ(n=6), GOLD Ⅲ(n=14), GOLD Ⅳ(n=14). The levels of cathepsin S and IFN-γ in BALF were determined by enzyme-linked immunosorbent assay (ELISA). The percentage ratio of low attenuation area to total lung area (LAA%), two times the ratio of airway wall thickness to outer diameter(2T/D), and the ratio of wall area to total cross-sectional area (WA) were measured by HRCT. Results: There were significant differences in the levels of cathepsin S in BALF between the groups (F=6.639, P=0.000). BALF cathepsin S levels were as follows: GOLD Ⅳ grou P>GOLD Ⅲ grou P>GOLD Ⅱ grou P>GOLD group Ⅰ >healthy control group (P value were all<0.05); LAA grade 3>LAA grade 2>LAA grade 1>LAA grade 0 (P value were all<0.05). Correlation analysis showed that BALF cathepsin S levels were correlated negatively with FEV(1)/FVC, FEV(1)% predicted, and DLCO% (r value was -0.065、-0.576、-0.392, respectively, P value were all<0.05), and but positively with RV/TLC%, LAA%, 2T/D, WA and IFN-γ(r value was 0.695, 0.497, 0.142, 0.309, 0.148, respectively, P value were all<0.05). Conclusion: The levels of cathepsin S were associated with the degree of airflow limitation and emphysema phenotype in COPD.


Assuntos
Líquido da Lavagem Broncoalveolar/imunologia , Catepsinas/metabolismo , Pulmão/diagnóstico por imagem , Doença Pulmonar Obstrutiva Crônica/metabolismo , Tomografia Computadorizada por Raios X/métodos , Biomarcadores , Catepsinas/imunologia , Ensaio de Imunoadsorção Enzimática , Humanos , Fenótipo , Doença Pulmonar Obstrutiva Crônica/imunologia , Testes de Função Respiratória
13.
Cells ; 8(3)2019 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-30897858

RESUMO

: For a long time, cysteine cathepsins were considered primarily as proteases crucial for nonspecific bulk proteolysis in the endolysosomal system. However, this view has dramatically changed, and cathepsins are now considered key players in many important physiological processes, including in diseases like cancer, rheumatoid arthritis, and various inflammatory diseases. Cathepsins are emerging as important players in the extracellular space, and the paradigm is shifting from the degrading enzymes to the enzymes that can also specifically modify extracellular proteins. In pathological conditions, the activity of cathepsins is often dysregulated, resulting in their overexpression and secretion into the extracellular space. This is typically observed in cancer and inflammation, and cathepsins are therefore considered valuable diagnostic and therapeutic targets. In particular, the investigation of limited proteolysis by cathepsins in the extracellular space is opening numerous possibilities for future break-through discoveries. In this review, we highlight the most important findings that establish cysteine cathepsins as important players in the extracellular space and discuss their roles that reach beyond processing and degradation of extracellular matrix (ECM) components. In addition, we discuss the recent developments in cathepsin research and the new possibilities that are opening in translational medicine.


Assuntos
Catepsinas/metabolismo , Microambiente Celular , Cisteína/metabolismo , Matriz Extracelular/metabolismo , Humanos , Modelos Moleculares , Pesquisa Médica Translacional
14.
Biochem Soc Trans ; 47(2): 615-623, 2019 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-30902923

RESUMO

The mass recruitment to the midgut contents of lysosomal proteolytic enzymes occurred in insects under three major selective pressures. Hemipteran (true bugs, aphids, and cicadas) ancestors lost their serine peptidases (SP) on adapting to feed on protein-free plant sap. When they returned to protein diets, their cathepsins L and B were recruited to replace their lost SP. Among beetles of the series Cucujiformia, cathepsins L were recruited to hydrolyze ingested plant inhibitors that affect their major SP and/or to deal with special seed proteins, such as prolamins. Larval flies have a very acid middle midgut and use cathepsin D to digest bacteria from their infected food. All the recruited enzymes originated from duplicated genes. The recruited digestive enzymes differ from their lysosomal counterparts in critical regions of their amino acid sequences that resulted in changes in substrate specificities and other kinetic properties. The discharge of digestive cathepsins in the midgut contents, instead of lysosomes, seems to be a consequence of their overexpression or the existence of new targeting signals. Their activation at the midgut contents occurs by an autoactivation mechanism or with the help of other enzymes or by a combination of both. The targeting to lysosomes of the insect lysosomal enzymes does not follow the mammalian mannose 6-phosphate route, but an incompletely known mechanism.


Assuntos
Sistema Digestório/enzimologia , Enzimas/metabolismo , Proteínas de Insetos/metabolismo , Lisossomos/enzimologia , Animais , Catepsinas/metabolismo , Insetos
15.
J Vet Med Sci ; 81(4): 522-531, 2019 Apr 16.
Artigo em Inglês | MEDLINE | ID: mdl-30726795

RESUMO

The basement membrane surrounding cardiomyocytes is mainly composed of α1 and α2 chain of type IV collagen. Arresten and canstatin are fragments of non-collagenous C-terminal domain of α1 and α2 chain, respectively. We previously reported that the expression of canstatin was decreased in infarcted area 2 weeks after myocardial infarction in rats. In the present study, we investigated the regulatory mechanism for expression of arresten and canstatin. Myocardial infarction model rats were produced by ligating left anterior descending artery. Western blotting and immunohistochemical staining were performed to determine the protein expression and distribution. Arresten and canstatin were highly expressed in the heart. One day and three days after myocardial infarction, the expression of arresten and canstatin in infarcted area was lower than that in non-infarcted area. The expression of cathepsin S, which is known to degrade arresten and canstatin, was increased in the infarcted area. A knockdown of cathepsin S gene using small interference RNA suppressed the decline of arresten and canstatin in the infarcted area 3 days after myocardial infarction. This study for the first time revealed that arresten and canstatin are immediately degraded by cathepsin S in the infarcted area after myocardial infarction. These findings present a novel fundamental insight into the pathogenesis of myocardial infarction through the turnover of basement membrane-derived endogenous factors.


Assuntos
Arrestina/metabolismo , Catepsinas/metabolismo , Colágeno Tipo IV/metabolismo , Infarto do Miocárdio/fisiopatologia , Animais , Western Blotting , Catepsinas/genética , Coração/fisiopatologia , Imuno-Histoquímica , Masculino , Infarto do Miocárdio/metabolismo , Miocárdio/metabolismo , Interferência de RNA , Ratos Wistar
16.
Arch Biochem Biophys ; 670: 32-42, 2019 07 30.
Artigo em Inglês | MEDLINE | ID: mdl-30807742

RESUMO

Lysosomal cysteine cathepsins are a family of proteases that are involved in a myriad of cellular processes from proteolytic degradation in the lysosome to bone resorption. These proteins mature following the cleavage of a pro-domain in the lysosome to become either exo- or endo-peptidases. The cathepsins B, C, L, S and Z have been implicated in NLRP3 inflammasome activation following their activation with ATP, monosodium urate, silica crystals, or bacterial components, among others. These five cathepsins have both compensatory and independent functions in NLRP3 inflammasome activation. There is much evidence in the literature to support the release of cathepsin B following lysosomal membrane degradation which leads to NLRP3 inflammasome activation. This is likely due to a hitherto unidentified role of this protein in the cytoplasm, although other interactions with autophagy proteins and within lysosomes have been proposed. Cathepsin C is involved in the processing of neutrophil IL-1ß through processing of upstream proteases. Cathepsin Z is non-redundantly required for NLRP3 inflammasome activation following nigericin, ATP and monosodium urate activation. Lysosomal cysteine cathepsins are members of a diverse and complementary family, and likely share both overlapping and independent functions in NLRP3 inflammasome activation.


Assuntos
Catepsinas/metabolismo , Cisteína/metabolismo , Inflamassomos/metabolismo , Lisossomos/enzimologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Animais , Humanos
17.
Gynecol Obstet Invest ; 84(4): 396-406, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30759440

RESUMO

BACKGROUND/AIMS: The study aimed to evaluate molecular changes related to trophoblast adhesion in placenta accreta spectrum (PAS) disorders. METHODS: A retrospective analysis of 10 PAS cases in which both the trophoblast adherent site and the non-adherent site were identified was performed in April 2010 and March 2013. Microarray analysis and reverse transcription polymerase chain reaction (RT-PCR) analyses were performed to extract upregulated genes in the adherent site. Gene expression changes were examined by immunohistochemistry. RESULTS: Microarray analysis showed that 157 transcripts were > 3-fold upregulated, including the following: a disintegrin and metalloproteinase-28 (ADAM28), 3.10-fold; cathepsin V (CTSV), 3.73-fold; cathepsin S (CTSS), 3.46-fold; and matrix metalloproteinase-19 (MMP19), 3.41-fold. RT-PCR showed relatively high mRNA expressions. On immunohistochemistry, extravillous trophoblast (EVT) at the non-adherent site showed weak or no CTSV expression, whereas EVT that invaded myometrium at the adherent site showed strong expression (histological score, median [min-max], 115.6 [37.6-153.6] vs. 184.8 [56.4-222.8], p < 0.05). MMP19 showed moderate staining, with no difference between the adherent and non-adherent sites. ADAM28 and CTSS showed weak or no staining. DISCUSSION: This limited study suggests that CTSV may be involved in the pathogenesis of PAS.


Assuntos
Catepsinas/metabolismo , Adesão Celular/genética , Cisteína Endopeptidases/metabolismo , Placenta Acreta/genética , Trofoblastos/metabolismo , Proteínas ADAM/metabolismo , Adulto , Feminino , Humanos , Imuno-Histoquímica , Metaloproteinases da Matriz Secretadas/metabolismo , Miométrio/metabolismo , Placenta/metabolismo , Gravidez , Estudos Retrospectivos
18.
Dev Genes Evol ; 229(1): 35-41, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30756180

RESUMO

Silicatein is the main protein responsible for the formation of spicules, tiny structures that constitute the silica skeleton of marine demosponges (Phylum Porifera). A unique innovation in Porifera that evolved from the cathepsin L family of proteins, it has been reported that two amino acids (S and H) are necessary to form the catalytic triad (SHN) to enable silica condensation. However, a diversity of silicatein sequence variants has since been reported with a variable pattern of presence/absence across sponge groups. Variants containing CHN or C/SQN at the active site appear more common in sponges from the Haplosclerida. Here, we report the expression levels of five silicatein variants through different developmental stages in the haplosclerid Haliclona indistincta. All five silicatein variants were expressed at low levels in the free-swimming larvae, which lack spicules and expression significantly increased at the two developmental phases in which spicules were visible. At these two phases, silicateins of CHN and C/SQN types were much more highly expressed than the SHN type indicating a possible ability of active sites with these alternative amino acids to condense silica and a more complex evolutionary story for spicule formation in marine demosponges than previously understood.


Assuntos
Catepsinas/genética , Regulação da Expressão Gênica no Desenvolvimento , Haliclona/genética , Animais , Catepsinas/química , Catepsinas/metabolismo , Haliclona/crescimento & desenvolvimento , Haliclona/metabolismo
19.
Neuroscience ; 400: 72-84, 2019 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-30625334

RESUMO

Spino-cerebellar ataxia type 7 (SCA7) is a polyglutamine (polyQ) disorder characterized by neurodegeneration of the brain, cerebellum, and retina caused by a polyglutamine expansion in ataxin7. The presence of an expanded polyQ tract in a mutant protein is known to induce protein aggregation, cellular stress, toxicity, and finally cell death. However, the consequences of the presence of mutant ataxin7 in the retina and the mechanisms underlying photoreceptor degeneration remain poorly understood. In this study, we show that in a retinal SCA7 mouse model, polyQ ataxin7 induces stress within the retina and activates Muller cells. Moreover, unfolded protein response and autophagy are activated in SCA7 photoreceptors. We have also shown that the photoreceptor death does not involve a caspase-dependent apoptosis but instead involves apoptosis inducing factor (AIF) and Leukocyte Elastase Inhibitor (LEI/L-DNase II). When these two cell death effectors are downregulated by their siRNA, a significant reduction in photoreceptor death is observed. These results highlight the consequences of polyQ protein expression in the retina and the role of caspase-independent pathways involved in photoreceptor cell death.


Assuntos
Ataxina-7/metabolismo , Morte Celular , Peptídeos/metabolismo , Degeneração Retiniana/metabolismo , Ataxias Espinocerebelares/metabolismo , Animais , Fator de Indução de Apoptose/metabolismo , Ataxina-7/genética , Calpaína/metabolismo , Caspases/metabolismo , Catepsinas/metabolismo , Modelos Animais de Doenças , Endodesoxirribonucleases/metabolismo , Células HEK293 , Humanos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células Fotorreceptoras/metabolismo , Degeneração Retiniana/etiologia , Transdução de Sinais , Ataxias Espinocerebelares/complicações , Estresse Fisiológico
20.
Am J Respir Crit Care Med ; 200(1): 51-62, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-30641028

RESUMO

Rationale: CTSS (cathepsin S) is a cysteine protease that is observed at higher concentrations in BAL fluid and plasma of subjects with chronic obstructive pulmonary disease (COPD). Objectives: To investigate whether CTSS is involved in the pathogenesis of cigarette smoke-induced COPD and determine whether targeting upstream signaling could prevent the disease. Methods: CTSS expression was investigated in animal and human tissue and cell models of COPD. Ctss-/- mice were exposed to long-term cigarette smoke and forced oscillation and expiratory measurements were recorded. Animals were administered chemical modulators of PP2A (protein phosphatase 2A) activity. Measurements and Main Results: Here we observed enhanced CTSS expression and activity in mouse lungs after exposure to cigarette smoke. Ctss-/- mice were resistant to cigarette smoke-induced inflammation, airway hyperresponsiveness, airspace enlargements, and loss of lung function. CTSS expression was negatively regulated by PP2A in human bronchial epithelial cells isolated from healthy nonsmokers and COPD donors and in monocyte-derived macrophages. Modulating PP2A expression or activity, with silencer siRNA or a chemical inhibitor or activator, during acute smoke exposure in mice altered inflammatory responses and CTSS expression and activity in the lung. Enhancement of PP2A activity prevented chronic smoke-induced COPD in mice. Conclusions: Our study indicates that the decrease in PP2A activity that occurs in COPD contributes to elevated CTSS expression in the lungs and results in impaired lung function. Enhancing PP2A activity represents a feasible therapeutic approach to reduce CTSS activity and counter smoke-induced lung disease.


Assuntos
Catepsinas/metabolismo , Fumar Cigarros/metabolismo , Pulmão/metabolismo , Proteína Fosfatase 2/metabolismo , Doença Pulmonar Obstrutiva Crônica/metabolismo , Fumaça/efeitos adversos , Tabaco , Animais , Brônquios/citologia , Estudos de Casos e Controles , Fumar Cigarros/efeitos adversos , Modelos Animais de Doenças , Inibidores Enzimáticos/farmacologia , Inativação Gênica , Humanos , Pulmão/efeitos dos fármacos , Macrófagos Alveolares/metabolismo , Camundongos , Camundongos Knockout , Ácido Okadáico/farmacologia , Proteína Fosfatase 2/antagonistas & inibidores , Doença Pulmonar Obstrutiva Crônica/etiologia , Doença Pulmonar Obstrutiva Crônica/fisiopatologia , Mucosa Respiratória/citologia
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