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1.
Nat Commun ; 15(1): 1976, 2024 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-38438396

RESUMO

Hemorrhagic toxin (TcsH) is a major virulence factor produced by Paeniclostridium sordellii, which is a non-negligible threat to women undergoing childbirth or abortions. Recently, Transmembrane Serine Protease 2 (TMPRSS2) was identified as a host receptor of TcsH. Here, we show the cryo-EM structures of the TcsH-TMPRSS2 complex and uncover that TcsH binds to the serine protease domain (SPD) of TMPRSS2 through the CROP unit-VI. This receptor binding mode is unique among LCTs. Five top surface loops of TMPRSS2SPD, which also determine the protease substrate specificity, constitute the structural determinants recognized by TcsH. The binding of TcsH inhibits the proteolytic activity of TMPRSS2, whereas its implication in disease manifestations remains unclear. We further show that mutations selectively disrupting TMPRSS2-binding reduce TcsH toxicity in the intestinal epithelium of the female mice. These findings together shed light on the distinct molecular basis of TcsH-TMPRSS2 interactions, which expands our knowledge of host recognition mechanisms employed by LCTs and provides novel targets for developing therapeutics against P. sordellii infections.


Assuntos
Serina Proteases , Toxinas Biológicas , Gravidez , Feminino , Humanos , Animais , Camundongos , Serina Proteases/genética , Serina , Fatores de Virulência/genética , Clostridiales , Serina Endopeptidases/genética
2.
Zh Nevrol Psikhiatr Im S S Korsakova ; 124(3. Vyp. 2): 42-48, 2024.
Artigo em Russo | MEDLINE | ID: mdl-38512094

RESUMO

The review presents the main physiological functions of thrombin. The procoagulant and anticoagulant activities of the key serine protease are discussed in both physiological and pathological conditions of hemostasis. The involvement of thrombin in atherogenesis, as well as its role as a mediator of vascular dysfunction and inflammation in both the peripheral and central nervous system, is highlighted. A pronounced imbalance between the pro- and anticoagulant systems leads to an increase in thrombin formation and creates conditions for the development of thrombosis. Tests that allow direct or indirect assessment of thrombin's functional activity are presented. The potential applications of direct thrombin inhibitors and direct blockers of thrombin PAR receptors in vascular neurology are also considered.


Assuntos
Neurologia , Trombina , Humanos , Serina Endopeptidases , Anticoagulantes , Sistema Nervoso Central
3.
Nat Commun ; 15(1): 2064, 2024 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-38453899

RESUMO

FAM111A, a serine protease, plays roles in DNA replication and antiviral defense. Missense mutations in the catalytic domain cause hyper-autocleavage and are associated with genetic disorders with developmental defects. Despite the enzyme's biological significance, the molecular architecture of the FAM111A serine protease domain (SPD) is unknown. Here, we show that FAM111A is a dimerization-dependent protease containing a narrow, recessed active site that cleaves substrates with a chymotrypsin-like specificity. X-ray crystal structures and mutagenesis studies reveal that FAM111A dimerizes via the N-terminal helix within the SPD. This dimerization induces an activation cascade from the dimerization sensor loop to the oxyanion hole through disorder-to-order transitions. Dimerization is essential for proteolytic activity in vitro and for facilitating DNA replication at DNA-protein crosslink obstacles in cells, while it is dispensable for autocleavage. These findings underscore the role of dimerization in FAM111A's function and highlight the distinction in its dimerization dependency between substrate cleavage and autocleavage.


Assuntos
Serina Endopeptidases , Serina Proteases , Dimerização , Serina Endopeptidases/metabolismo , Proteólise , Replicação do DNA , Serina
4.
FASEB J ; 38(5): e23531, 2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38466220

RESUMO

Inhaled aeroallergens can directly activate airway epithelial cells (AECs). Exposure to cockroach allergens is a strong risk factor for asthma. Cockroach allergens mediate some of their effects through their serine protease activity; protease activity is also a major contributor to allergenicity. The Th2 cytokine interleukin-13 (IL-13) induces upregulation of the eosinophil chemotactic factor CCL26. CCL26 induces eosinophil migration in allergic inflammation. In this work, we studied the effect of cockroach proteases on IL-13-induced effects. Immersed cultures of the human bronchial epithelial cell line BEAS-2B and air-liquid interface (ALI) cultures of primary normal human bronchial epithelial (NHBE) cells were stimulated with IL-13, Blattella Germanica cockroach extract (CE), or both. IL-13-induced genes were analyzed with qRT-PCR. IL-13 induced upregulation of CCL26, periostin, and IL-13Rα2 in bronchial epithelial cells which were decreased by CE. CE was heat-inactivated (HICE) or pre-incubated with protease inhibitors. HICE and CE preincubated with serine protease inhibitors did not prevent IL-13-induced CCL26 upregulation. CE-degraded IL-13 and specific cleavage sites were identified. CE also decreased IL-4-induced CCL26 upregulation and degraded IL-4. Other serine proteases such as bovine trypsin and house dust mite (HDM) serine proteases did not have the same effects on IL-13-induced CCL26. We conclude that CE serine proteases antagonize IL-13-induced effects in AECs, and this CE effect is mediated primarily through proteolytic cleavage of IL-13. IL-13 cleavage by cockroach serine proteases may modulate CCL26-mediated effects in allergic airway inflammation by interfering directly with the pro-inflammatory effects of IL-13 in vivo.


Assuntos
Blattellidae , Humanos , Animais , Bovinos , Interleucina-13 , Interleucina-4 , Serina Proteases , Serina Endopeptidases , Inflamação , Quimiocina CCL26
5.
FASEB J ; 38(6): e23566, 2024 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-38526868

RESUMO

Trypanosoma cruzi is the causative agent of Chagas disease, a chronic pathology that affects the heart and/or digestive system. This parasite invades and multiplies in virtually all nucleated cells, using a variety of host cell receptors for infection. T. cruzi has a gene that encodes an ecotin-like inhibitor of serine peptidases, ISP2. We generated ISP2-null mutants (Δisp2) in T. cruzi Dm28c using CRISPR/Cas9. Epimastigotes of Δisp2 grew normally in vitro but were more susceptible to lysis by human serum compared to parental and ISP2 add-back lines. Tissue culture trypomastigotes of Δisp2 were more infective to human muscle cells in vitro, which was reverted by the serine peptidase inhibitors aprotinin and camostat, suggesting that host cell epitheliasin/TMPRSS2 is the target of ISP2. Pretreatment of host cells with an antagonist to the protease-activated receptor 2 (PAR2) or an inhibitor of Toll-like receptor 4 (TLR4) selectively counteracted the increased cell invasion by Δisp2, but did not affect invasion by parental and add-back lines. The same was observed following targeted gene silencing of PAR2, TLR4 or TMPRSS2 in host cells by siRNA. Furthermore, Δisp2 caused increased tissue edema in a BALB/c mouse footpad infection model after 3 h differently to that observed following infection with parental and add-back lines. We propose that ISP2 contributes to protect T. cruzi from the anti-microbial effects of human serum and to prevent triggering of PAR2 and TLR4 in host cells, resulting in the modulation of host cell invasion and contributing to decrease inflammation during acute infection.


Assuntos
Doença de Chagas , Trypanosoma cruzi , Animais , Camundongos , Humanos , Receptor 4 Toll-Like/genética , Receptor PAR-2/genética , Doença de Chagas/genética , Doença de Chagas/parasitologia , Antivirais/farmacologia , Inibidores de Serino Proteinase/farmacologia , Inflamação , Serina , Serina Endopeptidases/genética
6.
Placenta ; 148: 77-83, 2024 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-38417305

RESUMO

INTRODUCTION: Corin protein and its coding gene variants have been associated with hypertensive disorders of pregnancy (HDP), but the underlying mechanisms are unclear. As a mediator linking fixed genome with the dynamic environment, DNA methylation at the CORIN gene may link corin with HDP but not has been studied. This study aimed to examine whether CORIN promoter methylation and HDP in Chinese pregnant women. METHODS: Based on a cohort of Chinese pregnant women, we designed a nested case-control study including 196 cases with HDP and 200 healthy controls. DNA methylation levels in the CORIN promoter were quantified by pyrosequencing using peripheral blood before 20 gestational weeks. The association between DNA methylation in CORIN promoter and HDP was systemically examined by single CpG association analysis, followed by gene-based analysis. Multiple testing was controlled by the false discovery rate (FDR) method. RESULTS: The single CpG association analysis found that, among the 5 CpG sites assayed, hypermethylation at one CpG site (Chr4:47839945) was significantly associated with HDP (OR = 1.94, raw P = 0.020), but the significance did not survive for multiple testing correction (FDR-P = 0.100). The gene-based association analysis found that DNA methylation of the 5 CpG sites was jointly associated with HDP (raw P = 0.003). In addition to HDP, CORIN promoter methylation was also significantly associated with dynamic blood pressure during pregnancy (raw P < 0.05). DISCUSSION: Hypermethylation in CORIN promoter at early pregnancy was associated with the risk of HDP during late pregnancy in Chinese women. However, further evidence is required to establish the causality between CORIN promoter methylation and HDP.


Assuntos
Hipertensão Induzida pela Gravidez , Pré-Eclâmpsia , Gravidez , Feminino , Humanos , Estudos de Casos e Controles , Pré-Eclâmpsia/genética , Pressão Sanguínea , Metilação de DNA , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo
7.
World J Surg Oncol ; 22(1): 49, 2024 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-38331878

RESUMO

BACKGROUND: TMPRSS2-ERG (T2E) fusion is highly related to aggressive clinical features in prostate cancer (PC), which guides individual therapy. However, current fusion prediction tools lacked enough accuracy and biomarkers were unable to be applied to individuals across different platforms due to their quantitative nature. This study aims to identify a transcriptome signature to detect the T2E fusion status of PC at the individual level. METHODS: Based on 272 high-throughput mRNA expression profiles from the Sboner dataset, we developed a rank-based algorithm to identify a qualitative signature to detect T2E fusion in PC. The signature was validated in 1223 samples from three external datasets (Setlur, Clarissa, and TCGA). RESULTS: A signature, composed of five mRNAs coupled to ERG (five ERG-mRNA pairs, 5-ERG-mRPs), was developed to distinguish T2E fusion status in PC. 5-ERG-mRPs reached 84.56% accuracy in Sboner dataset, which was verified in Setlur dataset (n = 455, accuracy = 82.20%) and Clarissa dataset (n = 118, accuracy = 81.36%). Besides, for 495 samples from TCGA, two subtypes classified by 5-ERG-mRPs showed a higher level of significance in various T2E fusion features than subtypes obtained through current fusion prediction tools, such as STAR-Fusion. CONCLUSIONS: Overall, 5-ERG-mRPs can robustly detect T2E fusion in PC at the individual level, which can be used on any gene measurement platform without specific normalization procedures. Hence, 5-ERG-mRPs may serve as an auxiliary tool for PC patient management.


Assuntos
Neoplasias da Próstata , Transcriptoma , Masculino , Humanos , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Proteínas de Fusão Oncogênica/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , RNA Mensageiro/genética , Regulador Transcricional ERG/genética , Regulador Transcricional ERG/metabolismo , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo , Serina Endopeptidases/uso terapêutico
8.
BMC Pediatr ; 24(1): 105, 2024 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-38341535

RESUMO

Several studies have shown association of single nucleotide polymorphisms (SNPs) of hepcidin regulatory pathways genes with impaired iron status. The most common is in the TMPRSS6 gene. In Africa, very few studies have been reported. We aimed to investigate the correlation between the common SNPs in the transmembrane protease, serine 6 (TMPRSS6) gene and iron indicators in a sample of Egyptian children for identifying the suitable candidate for iron supplementation.Patients and methods One hundred and sixty children aged 5-13 years were included & classified into iron deficient, iron deficient anemia and normal healthy controls. All were subjected to assessment of serum iron, serum ferritin, total iron binding capacity, complete blood count, reticulocyte count, serum soluble transferrin receptor and serum hepcidin. Molecular study of TMPRSS6 genotyping polymorphisms (rs4820268, rs855791 and rs11704654) were also evaluated.Results There was an association of iron deficiency with AG of rs855791 SNP, (P = 0.01). The minor allele frequency for included children were 0.43, 0.45 & 0.17 for rs4820268, rs855791 & rs11704654 respectively. Genotype GG of rs4820268 expressed the highest hepcidin gene expression fold, the lowest serum ferroportin & iron store compared to AA and AG genotypes (p = 0.05, p = 0.05, p = 0.03 respectively). GG of rs855791 had lower serum ferritin than AA (p = 0.04), lowest iron store & highest serum hepcidin compared to AA and AG genotypes (p = 0.04, p = 0.01 respectively). Children having CC of rs11704654 had lower level of hemoglobin, serum ferritin and serum hepcidin compared with CT genotype (p = 0.01, p = 0.01, p = 0.02) respectively.Conclusion Possible contribution of SNPs (rs855791, rs4820268 and rs11704654) to low iron status.


Assuntos
Anemia Ferropriva , Ferro , Criança , Humanos , Hepcidinas/genética , Hepcidinas/metabolismo , Projetos Piloto , Serina/genética , Peptídeo Hidrolases/genética , Peptídeo Hidrolases/metabolismo , Egito , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo , Polimorfismo de Nucleotídeo Único , Ferritinas , Anemia Ferropriva/genética , Proteínas de Membrana/genética
9.
Theranostics ; 14(4): 1390-1429, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38389851

RESUMO

Rationale: Tripeptidyl peptidase II (TPP2) has been proven to be related to human immune and neurological diseases. It is generally considered as a cytosolic protein which forms the largest known protease complex in eukaryotic cells to operate mostly downstream of proteasomes for degradation of longer peptides. However, this canonical function of TPP2 cannot explain its role in a wide variety of biological and pathogenic processes. The mechanistic interrelationships and hierarchical order of these processes have yet to be clarified. Methods: Animals, cells, plasmids, and viruses established and/or used in this study include: TPP2 knockout mouse line, TPP2 conditional knockout mouse lines (different neural cell type oriented), TRE-TPP2 knockin mouse line on the C57BL/6 background; 293T cells with depletion of TPP2, ATF6, IRE1, PERK, SYVN1, UCHL1, ATG5, CEPT1, or CCTα, respectively; 293T cells stably expressing TPP2, TPP2 S449A, TPP2 S449T, or CCTα-KDEL proteins on the TPP2-depleted background; Plasmids for eukaryotic transient expression of rat CYP19A1-Flag, CYP19A1 S118A-Flag, CYP19A1 S118D-Flag, Sac I ML GFP Strand 11 Long, OMMGFP 1-10, G-CEPIA1er, GCAMP2, CEPIA3mt, ACC-GFP, or SERCA1-GFP; AAV2 carrying the expression cassette of mouse CYP19A1-3 X Flag-T2A-ZsGreen. Techniques used in this study include: Flow cytometry, Immunofluorescence (IF) staining, Immunohistochemical (IHC) staining, Luxol fast blue (LFB) staining, ß-galactosidase staining, Lipid droplet (LD) staining, Calcium (Ca2+) staining, Stimulated emission depletion (STED) imaging, Transmission electron microscopic imaging, Two-photon imaging, Terminal deoxynucleotidyl transferase (TdT) dUTP nick-end Labeling (TUNEL) assay, Bromodeoxyuridine (BrdU) assay, Enzymatic activity assay, Proximity ligation assay (PLA), In vivo electrophysiological recording, Long-term potentiation (LTP) recording, Split-GFP-based mitochondria-associated membrane (MAM) detection, Immunoprecipitation (IP), Cellular fractionation, In situ hybridization, Semi-quantitative RT-PCR, Immunoblot, Mass spectrometry-based lipidomics, metabolomics, proteomics, Primary hippocampal neuron culture and Morris water maze (MWM) test. Results: We found that TPP2, independent of its enzymatic activity, plays a crucial role in maintaining the homeostasis of intracellular Ca2+ and phosphatidylcholine (PC) in the central nervous system (CNS) of mice. In consistence with the critical importance of Ca2+ and PC in the CNS, TPP2 gene ablation causes presenile dementia in female mice, which is closely associated with Ca2+/PC dysregulation-induced endoplasmic reticulum (ER) stress, abnormal autophagic degradation of CYP19A1 (aromatase), and estrogen depletion. This work therefore uncovers a new role of TPP2 in lipogenesis and neurosteroidogenesis which is tightly related to cognitive function of adult female mice. Conclusion: Our study reveals a crucial role of TPP2 in controlling homeostasis of Ca2+ and lipids in CNS, and its deficiency causes sexual dimorphism in dementia. Thus, this study is not only of great significance for elucidating the pathogenesis of dementia and its futural treatment, but also for interpreting the role of TPP2 in other systems and their related disorders.


Assuntos
Doença de Alzheimer , Aminopeptidases , Cálcio , Dipeptidil Peptidases e Tripeptidil Peptidases , Serina Endopeptidases , Animais , Feminino , Humanos , Camundongos , Ratos , Aromatase , Cálcio/metabolismo , Sistema Nervoso Central/metabolismo , Homeostase , Lipídeos , Camundongos Endogâmicos C57BL , Camundongos Knockout
10.
Antonie Van Leeuwenhoek ; 117(1): 41, 2024 Feb 24.
Artigo em Inglês | MEDLINE | ID: mdl-38400879

RESUMO

In the search of new enzymatic activities with a possible industrial application, we focused on those microorganisms and their molecular mechanisms that allow them to succeed in the environment, particularly in the proteolytic activity and its central role in the microorganisms' successful permanence. The use of highly active serine proteases for industrial applications is a modern need, especially for the formulation of detergents, protein processing, and hair removal from animal skins. This report provides the isolation and identification of a highly proteolytic fragment derived from DegQ produced by a Pseudomonas fluorescens environmental strain isolated from a frog carcass. Zymograms demonstrate that a 10 kDa protein mainly generates the total proteolytic activity of this strain, which is enhanced by the detergent SDS. Mass spectroscopy analysis revealed that the protein derived a couple of peptides, the ones showing the highest coverage belonging to DegQ. Interestingly, this small protein fragment contains a PDZ domain but no obvious residues indicating that it is a protease. Protein model analysis shows that this fragment corresponds to the main PDZ domain from DegQ, and its unique sequence and structure render a proteolytic peptide. The results presented here indicate that a novel DegQ fragment is sufficient for obtaining high protease activity highlighting that the analysis of environmental microorganisms can render new strains or enzymes with helpful biotechnological characteristics.


Assuntos
Domínios PDZ , Pseudomonas , Animais , Pseudomonas/genética , Pseudomonas/metabolismo , Serina Endopeptidases/química , Serina Endopeptidases/metabolismo , Peptídeos , Serina Proteases
11.
Int J Mol Sci ; 25(3)2024 Jan 23.
Artigo em Inglês | MEDLINE | ID: mdl-38338655

RESUMO

Trypsin-like serine proteases are involved in many important physiological processes like blood coagulation and remodeling of the extracellular matrix. On the other hand, they are also associated with pathological conditions. The urokinase-pwlasminogen activator (uPA), which is involved in tissue remodeling, can increase the metastatic behavior of various cancer types when overexpressed and dysregulated. Another member of this protease class that received attention during the SARS-CoV 2 pandemic is TMPRSS2. It is a transmembrane serine protease, which enables cell entry of the coronavirus by processing its spike protein. A variety of different inhibitors have been published against both proteases. However, the selectivity over other trypsin-like serine proteases remains a major challenge. In the current study, we replaced the arginine moiety at the P1 site of peptidomimetic inhibitors with different bioisosteres. Enzyme inhibition studies revealed that the phenylguanidine moiety in the P1 site led to strong affinity for TMPRSS2, whereas the cyclohexylguanidine derivate potently inhibited uPA. Both inhibitors exhibited high selectivity over other structurally similar and physiologically important proteases.


Assuntos
Peptidomiméticos , Inibidores de Serino Proteinase , Ativador de Plasminogênio Tipo Uroquinase , Ligantes , Peptídeo Hidrolases , Peptidomiméticos/química , Peptidomiméticos/farmacologia , Tripsina , Ativador de Plasminogênio Tipo Uroquinase/metabolismo , Serina Endopeptidases , Inibidores de Serino Proteinase/química , Inibidores de Serino Proteinase/farmacologia
12.
Int J Mol Sci ; 25(4)2024 Feb 19.
Artigo em Inglês | MEDLINE | ID: mdl-38397107

RESUMO

Predicting the potency of inhibitors is key to in silico screening of promising synthetic or natural compounds. Here we describe a predictive workflow that provides calculated inhibitory values, which concord well with empirical data. Calculations of the free interaction energy ΔG with the YASARA plugin FoldX were used to derive inhibition constants Ki from PDB coordinates of protease-inhibitor complexes. At the same time, corresponding KD values were obtained from the PRODIGY server. These results correlated well with the experimental values, particularly for serine proteases. In addition, analyses were performed for inhibitory complexes of cysteine and aspartic proteases, as well as of metalloproteases, whereby the PRODIGY data appeared to be more consistent. Based on our analyses, we calculated theoretical Ki values for trypsin with sunflower trypsin inhibitor (SFTI-1) variants, which yielded the more rigid Pro14 variant, with probably higher potency than the wild-type inhibitor. Moreover, a hirudin variant with an Arg1 and Trp3 is a promising basis for novel thrombin inhibitors with high potency. Further examples from antibody interaction and a cancer-related effector-receptor system demonstrate that our approach is applicable to protein interaction studies beyond the protease field.


Assuntos
Helianthus , Serina Endopeptidases , Inibidores da Tripsina/farmacologia , Tripsina/metabolismo , Helianthus/metabolismo , Peptídeo Hidrolases , Inibidores de Proteases/farmacologia
13.
Science ; 383(6684): eadi3332, 2024 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-38359126

RESUMO

The identification of mechanisms to store glucose carbon in the form of glycogen rather than fat in hepatocytes has important implications for the prevention of nonalcoholic fatty liver disease (NAFLD) and other chronic metabolic diseases. In this work, we show that glycogenesis uses its intermediate metabolite uridine diphosphate glucose (UDPG) to antagonize lipogenesis, thus steering both mouse and human hepatocytes toward storing glucose carbon as glycogen. The underlying mechanism involves transport of UDPG to the Golgi apparatus, where it binds to site-1 protease (S1P) and inhibits S1P-mediated cleavage of sterol regulatory element-binding proteins (SREBPs), thereby inhibiting lipogenesis in hepatocytes. Consistent with this mechanism, UDPG administration is effective at treating NAFLD in a mouse model and human organoids. These findings indicate a potential opportunity to ameliorate disordered fat metabolism in the liver.


Assuntos
Lipogênese , Glicogênio Hepático , Fígado , Hepatopatia Gordurosa não Alcoólica , Pró-Proteína Convertases , Serina Endopeptidases , Uridina Difosfato Glucose , Animais , Humanos , Camundongos , Carbono/metabolismo , Glucose/metabolismo , Hepatócitos/metabolismo , Fígado/metabolismo , Glicogênio Hepático/metabolismo , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/metabolismo , Pró-Proteína Convertases/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo , Uridina Difosfato Glucose/administração & dosagem , Uridina Difosfato Glucose/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Células HEK293
14.
Cells ; 13(3)2024 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-38334616

RESUMO

Fundamental functions of the intestinal epithelium include the digestion of food, absorption of nutrients, and its ability to act as the first barrier against intruding microbes. Campylobacter jejuni is a major zoonotic pathogen accounting for a substantial portion of bacterial foodborne illnesses. The germ colonizes the intestines of birds and is mainly transmitted to humans through the consumption of contaminated poultry meat. In the human gastrointestinal tract, the bacterium triggers campylobacteriosis that can progress to serious secondary disorders, including reactive arthritis, inflammatory bowel disease and Guillain-Barré syndrome. We recently discovered that C. jejuni serine protease HtrA disrupts intestinal epithelial barrier functions via cleavage of the tight and adherens junction components occludin, claudin-8 and E-cadherin. However, it is unknown whether epithelial damage is mediated by the secreted soluble enzyme, by HtrA contained in shed outer-membrane vesicles (OMVs) or by another mechanism that has yet to be identified. In the present study, we investigated whether soluble recombinant HtrA and/or purified OMVs induce junctional damage to polarized intestinal epithelial cells compared to live C. jejuni bacteria. By using electron and confocal immunofluorescence microscopy, we show that HtrA-expressing C. jejuni bacteria trigger efficient junctional cell damage, but not soluble purified HtrA or HtrA-containing OMVs, not even at high concentrations far exceeding physiological levels. Instead, we found that only bacteria with active protein biosynthesis effectively cleave junctional proteins, which is followed by paracellular transmigration of C. jejuni through the epithelial cell layer. These findings shed new light on the pathogenic activities of HtrA and virulence strategies of C. jejuni.


Assuntos
Campylobacter jejuni , Humanos , Campylobacter jejuni/metabolismo , Serina Proteases/metabolismo , Serina Endopeptidases/metabolismo , Bactérias/metabolismo , Células Epiteliais/metabolismo , Junções Intercelulares/metabolismo
15.
Tissue Cell ; 87: 102329, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38367326

RESUMO

High-temperature requirement A1 (HtrA1), a multidomain serine protease acting on Extracellular matrix (ECM) rearrangement, is also secreted by osteoblasts and osteoclasts. Recent and conflicting literature highlights HtrA1's role as a controller of bone remodeling, proposing it as a possible target for pathologies with unbalanced bone resorption, like Osteoporosis (OP). To add knowledge on this molecule function in bone physiopathology, here we compared HtrA1 distribution in the ECM of healthy (H) and OP bone tissue, also examining its localization in the sites of new bone formation. HtrA1 was homogeneously expressed in the mature bone ECM of H tissue showing a 55.6 ± 16.4% of the stained area, with a significant (p=0.0001) decrease in OP percentage stained area (21.1 ± 13.1). Moreover, HtrA1 was present in the endosteum and cells involved in osteogenesis, mainly in those "entrapped" in woven bone, whereas osteocytes in mature lamellar bone were negative. Based on our previous observation in OP tissue of a significantly increased expression of Decorin and Osteocalcin, both involved in bone mineralization and remodeling and equally substrates for HtrA1, we speculate that HtrA1 by controlling the proper amount of Decorin and Osteocalcin favors normal bone maturation and mineralization. Besides, we suggest that late-osteoblasts and pre-osteocytes secrete HtrA1 in the adjacent matrix whilst proceeding with their maturation and that HtrA1 expression is further modified during the remodeling from woven to the lamellar bone. Overall, our data suggest HtrA1 as a positive regulator of bone matrix formation and maturation: its reduced expression in mature OP bone, affecting protein content and distribution, could hamper correct bone remodeling and mineralization.


Assuntos
Osteoporose , Serina Proteases , Humanos , Osteocalcina/metabolismo , Serina Proteases/metabolismo , Matriz Óssea/metabolismo , Decorina/metabolismo , Serina Peptidase 1 de Requerimento de Alta Temperatura A/genética , Serina Peptidase 1 de Requerimento de Alta Temperatura A/metabolismo , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo , Osso e Ossos/metabolismo , Matriz Extracelular/metabolismo , Osteoporose/genética
16.
J Immunol ; 212(7): 1172-1177, 2024 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-38372634

RESUMO

The activation of the CP/LP C3 proconvertase complex is a key event in complement activation and involves cleavage of C4 and C2 by the C1s protease (classical pathway) or the mannose-binding lectin-associated serine protease (MASP)-2 (lectin pathway). Efficient cleavage of C4 by C1s and MASP-2 involves exosites on the complement control protein and serine protease (SP) domains of the proteases. The complement control protein domain exosite is not involved in cleavage of C2 by the proteases, but the role of an anion-binding exosite (ABE) on the SP domains of the proteases has (to our knowledge) never been investigated. In this study, we have shown that the ABE on the SP of both C1s and MASP-2 is crucial for efficient cleavage of C2, with mutant forms of the proteases greatly impaired in their rate of cleavage of C2. We have additionally shown that the site of binding for the ABE of the proteases is very likely to be located on the von Willebrand factor domain of C2, with the precise area differing between the enzymes: whereas C1s requires two anionic clusters on the von Willebrand factor domain to enact efficient cleavage of C2, MASP-2 apparently only requires one. These data provide (to our knowledge) new information about the molecular determinants for efficient activation of C2 by C1s and MASP-2. The enhanced view of the molecular events underlying the early stages of complement activation provides further possible intervention points for control of this activation that is involved in a number of inflammatory diseases.


Assuntos
Ativação do Complemento , Lectina de Ligação a Manose , Serina Proteases Associadas a Proteína de Ligação a Manose , Complemento C1s , Complemento C4/metabolismo , Lectina de Ligação a Manose/metabolismo , Serina Proteases Associadas a Proteína de Ligação a Manose/metabolismo , Domínios Proteicos , Serina Endopeptidases/metabolismo , Serina Proteases/metabolismo , Fator de von Willebrand , Humanos , Células HEK293
17.
Clin Chim Acta ; 556: 117847, 2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38417778

RESUMO

Familial hypercholesterolemia (FH) is an inherited disorder characterized by increased low-density lipoprotein LDL) cholesterol and atherosclerotic cardiovascular disease. Although initial genetic analysis linked FH to LDL receptor mutations, subsequent work demonstrated that a gain-of-function mutation in the proprotein convertase subtilisin/kexin type 9 (PCSK9), which causes LDL-R degradation, was shown to be the cause of FH. In this review, we describe the history of research on FH, its clinical phenotyping and genotyping and advances in treatment with special focus on Japan.


Assuntos
Hiperlipoproteinemia Tipo II , Pró-Proteína Convertase 9 , Humanos , Pró-Proteína Convertase 9/genética , Serina Endopeptidases/metabolismo , Pró-Proteína Convertases/genética , Pró-Proteína Convertases/metabolismo , Pró-Proteína Convertases/uso terapêutico , Japão , Receptores de LDL/genética , Receptores de LDL/metabolismo , Hiperlipoproteinemia Tipo II/diagnóstico , Hiperlipoproteinemia Tipo II/genética , Mutação
18.
Infect Genet Evol ; 118: 105564, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38307396

RESUMO

This pilot study aimed to investigate genetic factors that may have contributed to the milder clinical outcomes of COVID-19 in Brazilian indigenous populations. 263 Indigenous from the Araweté, Kararaô, Parakanã, Xikrin do Bacajá, Kayapó and Munduruku peoples were analyzed, 55.2% women, ages ranging from 10 to 95 years (average 49.5 ± 20.7). Variants in genes involved in the entry of SARS-CoV-2 into the host cell (ACE1 rs1799752 I/D, ACE2 rs2285666 C/T, ACE2 rs73635825 A/G and TMPRSS2 rs123297605 C/T), were genotyped in indigenous peoples from the Brazilian Amazon, treated during the SARS-CoV-2 pandemic between 2020 and 2021. The distribution of genotypes did not show any association with the presence or absence of IgG antibodies. Additionally, the influence of genetic variations on the severity of the disease was not examined extensively because a significant number of indigenous individuals experienced the disease with either mild symptoms or no symptoms. It is worth noting that the frequencies of risk alleles were found to be lower in Indigenous populations compared to both continental populations and Brazilians. Indigenous Brazilian Amazon people exhibited an ethnic-specific genetic profile that may be associated with a milder disease, which could explain the unexpected response they demonstrated to COVID-19, being less impacted than Brazilians.


Assuntos
COVID-19 , Peptidil Dipeptidase A , Serina Endopeptidases , Feminino , Humanos , Masculino , Enzima de Conversão de Angiotensina 2/genética , Brasil/epidemiologia , COVID-19/epidemiologia , COVID-19/genética , Peptidil Dipeptidase A/genética , Projetos Piloto , SARS-CoV-2/fisiologia , Serina Endopeptidases/genética , Índios Sul-Americanos
19.
BMC Infect Dis ; 24(1): 112, 2024 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-38254046

RESUMO

BACKGROUND: The World Health Organization (WHO) declared Coronavirus Disease 2019 (COVID-19) a global pandemic on March 11, 2020. Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection has killed millions of people and had a terrible effect on society. The transmembrane protease serine 2 (TMPRSS2) enzyme is essential in the initial phases of the interplay between the SARSCoV-2 and the host cells by assisting viral entrance. METHODS: This observational case-control study involved 150 participants, 100 adult patients with COVID-19, 50 of whom appeared healthy and had no history of or symptoms of COVID-19 infection when the study was conducted. Between January and April 2022, patients were taken as inpatients in isolation units or through recruitment from the COVID-19 clinic at Kasr Al-Ainy Cairo University Hospitals. According to the National Institutes of Health guidelines (2021), they were categorised into three categories: mild, moderate, and severe. TMPRSS2 p.(Val197Met) variant genotyping was evaluated using TaqMan Real-Time PCR. RESULTS: The study showed a substantial difference between the mild and severe COVID-19 patient groups regarding their TMPRSS2 (p.Val197Met) genotypes (P value = 0.046). The C allele was significantly more prevalent in the mild, moderate and severe COVID-19 patient categories (77.8%, 89.7% and 91.7%, respectively) and the control group (80%). Meanwhile, the T allele was more prevalent in the mild (22.2%) and control (20%) groups. There was a statistically significant difference in allelic distribution between the mild and severe groups (P value = 0.034). CONCLUSION: The study showed a connection between the TMPRSS2 gene variant p.(Val197Met) and the degree of illness. We concluded that the T(mutant) allele was protective against severe COVID-19 because it was linked to lesser disease severity.


Assuntos
COVID-19 , Serina Endopeptidases , Adulto , Humanos , Alelos , Estudos de Casos e Controles , COVID-19/genética , Genótipo , SARS-CoV-2/patogenicidade , Serina Endopeptidases/genética , Estados Unidos
20.
Nat Commun ; 15(1): 889, 2024 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-38291024

RESUMO

Omicron emerged following COVID-19 vaccination campaigns, displaced previous SARS-CoV-2 variants of concern worldwide, and gave rise to lineages that continue to spread. Here, we show that Omicron exhibits increased infectivity in primary adult upper airway tissue relative to Delta. Using recombinant forms of SARS-CoV-2 and nasal epithelial cells cultured at the liquid-air interface, we show that mutations unique to Omicron Spike enable enhanced entry into nasal tissue. Unlike earlier variants of SARS-CoV-2, our findings suggest that Omicron enters nasal cells independently of serine transmembrane proteases and instead relies upon metalloproteinases to catalyze membrane fusion. Furthermore, we demonstrate that this entry pathway unlocked by Omicron Spike enables evasion from constitutive and interferon-induced antiviral factors that restrict SARS-CoV-2 entry following attachment. Therefore, the increased transmissibility exhibited by Omicron in humans may be attributed not only to its evasion of vaccine-elicited adaptive immunity, but also to its superior invasion of nasal epithelia and resistance to the cell-intrinsic barriers present therein.


Assuntos
COVID-19 , Interferons , Adulto , Humanos , SARS-CoV-2/genética , Vacinas contra COVID-19 , Mucosa Nasal , Serina Endopeptidases/genética , Serina Proteases , Glicoproteína da Espícula de Coronavírus/genética
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