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Here, we report the complete genome sequence of an Indian strain of chikungunya virus isolated from an infected patient from Hyderabad, Andhra Pradesh, India, during a massive outbreak in 2005-2006. The genome length spans 11,811 nucleotides and has a poly(A) tail of 29 residues at the 3' end.
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BACKGROUND: Triple negative breast cancer (TNBC) is the most aggressive subtype of breast cancer and exhibits high rate of chemoresistance, metastasis, and relapse. This can be attributed to the failure of conventional therapeutics to target a sub-population of slow cycling or quiescent cells called as cancer stem cells (CSCs). Therefore, elimination of CSCs is essential for effective TNBC treatment. PURPOSE: Research suggests that breast CSCs exhibit elevated glycolytic metabolism which directly contributes in maintenance of stemness, self-renewability and chemoresistance as well as in tumor progression. Therefore, this study aimed to target rewired metabolism which can serve as Achilles heel for CSCs population and have far reaching effect in TNBC treatment. METHODS: We used two preclinical models, zebrafish and nude mice to evaluate the fate of nanoparticles as well as the therapeutic efficacy of both piperlongumine (PL) and its nanomedicine (PL-NPs). RESULTS: In this context, we explored a phytochemical piperlongumine (PL) which has potent anti-cancer properties but poor pharmacokinetics impedes its clinical translation. So, we developed PLGA based nanomedicine for PL (PL-NPs), and demonstrated that it overcomes the pharmacokinetic limitations of PL, along with imparting advantages of selective tumor targeting through Enhanced Permeability and Retention (EPR) effect in zebrafish xenograft model. Further, we demonstrated that PL-NPs efficiently inhibit glycolysis in CSCs through inhibition of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) by modulating glutathione S-transferase pi 1 (GSTP1) and upregulation of fructose-1,6-bisphosphatase 1 (FBP1), a rate-limiting enzyme in gluconeogenesis. We also illustrated that inhibition of glycolysis results in overall tumor regression in two preclinical models. CONCLUSION: This study discusses novel mechanism of action by which PL acts on CSCSs. Taken together our study provides insight into development of PL based nanomedicine which could be exploited in clinics to achieve complete eradication of TNBC by targeting CSCs.
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Benzodioxoles , Neoplasias de la Mama Triple Negativas , Animales , Ratones , Humanos , Neoplasias de la Mama Triple Negativas/metabolismo , Pez Cebra/metabolismo , Nanomedicina , Ratones Desnudos , Línea Celular Tumoral , Recurrencia Local de Neoplasia/metabolismo , Células Madre Neoplásicas , Gliceraldehído-3-Fosfato Deshidrogenasas/metabolismo , Gliceraldehído-3-Fosfato Deshidrogenasas/farmacología , Gliceraldehído-3-Fosfato Deshidrogenasas/uso terapéutico , GlucólisisRESUMEN
BACKGROUND: The T cells, components of adaptive immunity participate in immune pathology of the autoimmune inflammatory disorder called rheumatoid arthritis (RA). The presence of TLRs on the surface of the CD8+ T cells and their ability to recognize bacterial moieties adds to the inflammatory burden in case of RA. It has been reported that the gut microbiome is necessary for the crucial shift in the balance between proinflammatory and anti-inflammatory cytokines. The altered gut microbiome and the presence of TLRs emphasizes on the microbiome driven inflammatory responses in case of RA. METHODS: Eighty-nine RA patients participated in this study. Clinical variations like disease duration, number of actively inflamed joints, number and type of bone deformities, CRP, RF, Anti-CCP, ESR, DAS 28 score were recorded for each patient. Co-culture of CD8+T cells and bacteria has been performed with proper culture condition. TLRs and inflammatory mediators' expression level were checked by both qPCR and flow cytometry analysis. RESULTS: We observed in the suppression of pro-inflammatory molecules like Granzyme B and IFNƳ and expression of TLR2 in CD8 + T cells upon treatment with Lactobacillus rhamnosus (L. rhamnosus). Moreover, L. rhamnosus activated CD8+T cells such that they could induce FOXP3 expression in CD4+T cells thereby skewing T cell population towards a regulatory phenotype. On the contrary, TLR4 engagement on CD8+T cell by Escherichia coli (E.coli) increased in inflammatory responses following ERK activation. CONCLUSIONS: Thus, we conclude that L. rhamnosus can effectively suppress CD8+T cell mediated inflammation by a simultaneous decrease of Th1 cells that may potentiate better treatment modalities for RA.
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Artritis Reumatoide , Lacticaseibacillus rhamnosus , Humanos , Linfocitos T CD8-positivos , Inflamación/metabolismo , Citocinas/metabolismo , Escherichia coli/metabolismoRESUMEN
Dendritic cells (DCs) undergo rapid metabolic reprogramming to generate signal-specific immune responses. The fine control of cellular metabolism underlying DC immune tolerance remains elusive. We have recently reported that NCoR1 ablation generates immune-tolerant DCs through enhanced IL-10, IL-27 and SOCS3 expression. In this study, we did comprehensive metabolic profiling of these tolerogenic DCs and identified that they meet their energy requirements through enhanced glycolysis and oxidative phosphorylation (OXPHOS), supported by fatty acid oxidation-driven oxygen consumption. In addition, the reduced pyruvate and glutamine oxidation with a broken TCA cycle maintains the tolerogenic state of the cells. Mechanistically, the AKT-mTOR-HIF-1α-axis mediated glycolysis and CPT1a-driven ß-oxidation were enhanced in these tolerogenic DCs. To confirm these observations, we used synthetic metabolic inhibitors and found that the combined inhibition of HIF-1α and CPT1a using KC7F2 and etomoxir, respectively, compromised the overall transcriptional signature of immunological tolerance including the regulatory cytokines IL-10 and IL-27. Functionally, treatment of tolerogenic DCs with dual KC7F2 and etomoxir treatment perturbed the polarization of co-cultured naïve CD4+ T helper (Th) cells towards Th1 than Tregs, ex vivo and in vivo. Physiologically, the Mycobacterium tuberculosis (Mtb) infection model depicted significantly reduced bacterial burden in BMcDC1 ex vivo and in CD103+ lung DCs in Mtb infected NCoR1DC-/-mice. The spleen of these infected animals also showed increased Th1-mediated responses in the inhibitor-treated group. These findings suggested strong involvement of NCoR1 in immune tolerance. Our validation in primary human monocyte-derived DCs (moDCs) showed diminished NCOR1 expression in dexamethasone-derived tolerogenic moDCs along with suppression of CD4+T cell proliferation and Th1 polarization. Furthermore, the combined KC7F2 and etomoxir treatment rescued the decreased T cell proliferative capacity and the Th1 phenotype. Overall, for the first time, we demonstrated here that NCoR1 mediated control of glycolysis and fatty acid oxidation fine-tunes immune tolerance versus inflammation balance in murine and human DCs.
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Interleucina-10 , Interleucina-27 , Humanos , Ratones , Animales , Interleucina-10/metabolismo , Interleucina-27/metabolismo , Células Dendríticas/metabolismo , Tolerancia Inmunológica , Glucólisis , Ácidos Grasos/metabolismo , Diferenciación Celular , Células Cultivadas , Co-Represor 1 de Receptor Nuclear/metabolismoRESUMEN
The antiviral state, an initial line of defense against viral infection, is established by a set of IFN-stimulated genes (ISGs) encoding antiviral effector proteins. The effector ISGs are transcriptionally regulated by type I IFNs mainly via activation of IFN-stimulated gene factor 3 (ISGF3). In this study, the regulatory elements of effector ISGs were characterized to determine the (epi)genetic features that enable their robust induction by type I IFNs in multiple cell types. We determined the location of regulatory elements, the DNA motifs, the occupancy of ISGF3 subunits (IRF9, STAT1, and STAT2) and other transcription factors, and the chromatin accessibility of 37 effector ISGs in murine dendritic cells. The IFN-stimulated response element (ISRE) and its tripartite version occurred most frequently in the regulatory elements of effector ISGs than in any other tested ISG subsets. Chromatin accessibility at their promoter regions was similar to most other ISGs but higher than at the promoters of inflammation-related cytokines, which were used as a reference gene set. Most effector ISGs (81.1%) had at least one ISGF3 binding region proximal to the transcription start site (TSS), and only a subset of effector ISGs (24.3%) was associated with three or more ISGF3 binding regions. The IRF9 signals were typically higher, and ISRE motifs were "stronger" (more similar to the canonical sequence) in TSS-proximal versus TSS-distal regulatory regions. Moreover, most TSS-proximal regulatory regions were accessible before stimulation in multiple cell types. Our results indicate that "strong" ISRE motifs and universally accessible promoter regions that permit robust, widespread induction are characteristic features of effector ISGs.
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Factores de Restricción Antivirales , Cromatina , Animales , Ratones , Cromatina/genética , Motivos de Nucleótidos , Regiones Promotoras Genéticas/genética , Elementos de Respuesta/genética , Interferones/metabolismoRESUMEN
CMTM6, a type 3 transmembrane protein, is known to stabilize the expression of programmed cell death ligand 1 (PD-L1) and hence facilitates the immune evasion of tumor cells. Recently, we demonstrated that CMTM6 is a major driver of cisplatin resistance in oral squamous cell carcinomas (OSCC). However, the detailed mechanism of how CMTM6 rewires cisplatin resistance in OSCC is yet to be explored. RNA sequencing analysis of cisplatin-resistant OSCC lines stably expressing Nt shRNA and CMTM6 shRNA revealed that CMTM6 might be a potential regulator of the ribosome biogenesis network. Knocking down CMTM6 significantly inhibited transcription of 47S precursor rRNA and hindered the nucleolar structure, indicating reduced ribosome biogenesis. When CMTM6 was ectopically over-expressed in CMTM6KD cells, almost all ribosomal machinery components were rescued. Mechanistically, CMTM6 induced the expression of C-Myc, which promotes RNA polymerase I mediated rDNA transcription. In addition to this, CMTM6 was also found to regulate the AKT-mTORC1-dependent ribosome biogenesis and protein synthesis in cisplatin-resistant lines. The nude mice and zebrafish xenograft experiments indicate that blocking ribosome synthesis either by genetic inhibitor (CMTM6KD) or pharmacological inhibitor (CX-5461) significantly restores cisplatin-mediated cell death in chemoresistant OSCC. Overall, our study suggests that CMTM6 is a major regulator of the ribosome biogenesis network and targeting the ribosome biogenesis network is a viable target to overcome chemoresistance in OSCC. The novel combination of CX-5461 and cisplatin deserves further clinical investigation in advanced OSCC.
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Carcinoma de Células Escamosas , Neoplasias de Cabeza y Cuello , Neoplasias de la Boca , Animales , Antígeno B7-H1 , Carcinoma de Células Escamosas/genética , Muerte Celular , Línea Celular Tumoral , Cisplatino/farmacología , ADN Ribosómico , Humanos , Ligandos , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Ratones Desnudos , Neoplasias de la Boca/tratamiento farmacológico , Neoplasias de la Boca/patología , Proteínas Proto-Oncogénicas c-akt , ARN Polimerasa I , ARN Interferente Pequeño , Ribosomas , Carcinoma de Células Escamosas de Cabeza y Cuello , Pez Cebra/genéticaRESUMEN
The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a major global health concern. This virus infects the upper respiratory tract and causes pneumonia-like symptoms. So far, few studies have shown alterations in nasopharyngeal (NP) microbial diversity, enrichment of opportunistic pathogens and their role in co-infections during respiratory infections. Therefore, we hypothesized that microbial diversity changes, with increase in the population of opportunistic pathogens, during SARS-CoV2 infection in the nasopharynx, which may be involved in co-infection in COVID-19 patients. The 16S rRNA variable regions, V1-V9, of NP samples of control and COVID-19 (symptomatic and asymptomatic) patients were sequenced using the Oxford Nanopore™ technology. Comprehensive bioinformatics analysis for determining alpha/beta diversities, non-metric multidimensional scaling, correlation studies, canonical correspondence analysis, linear discriminate analysis, and dysbiosis index were used to analyze the control and COVID-19-specific NP microbiomes. We observed significant dysbiosis in the COVID-19 NP microbiome with an increase in the abundance of opportunistic pathogens at genus and species levels in asymptomatic/symptomatic patients. The significant abundance of Mycobacteria spp. and Mycoplasma spp. in symptomatic patients suggests their association and role in co-infections in COVID-19 patients. Furthermore, we found strong correlation of enrichment of Mycobacteria and Mycoplasma with the occurrences of chest pain and fever in symptomatic COVID-19 patients. This is the first study from India to show the abundance of Mycobacteria and Mycoplasma opportunistic pathogens in non-hospitalized COVID-19 patients and their relationship with symptoms, indicating the possibility of co-infections.
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COVID-19 , Coinfección , Mycobacterium , Mycoplasma , Coinfección/epidemiología , Disbiosis , Humanos , Nasofaringe , ARN Ribosómico 16S/genética , ARN Viral , SARS-CoV-2RESUMEN
Rheumatoid arthritis (RA) is an autoimmune disorder of unknown etiology with aberrant immunological responses leading to inflammation, swelling and pain of the joints. CD8+ T cells have been known to be one of the major immune modulators in the progression of RA and the presence of toll-like receptors (TLRs) on these cells further accentuate their role in RA. Herein, we report an increased expression of TLR7 in the endosomes of CD8+ T cells of RA patients correlating with disease severity. The stimulation of TLR7 with Imiquimod (IMQ) in these CD8+ T cells drives the signalling cascade via NFkB and pERK activation and hence an increase in the mRNA transcripts of signature cytokines and cytolytic enzymes. However, a parallel synthesis of Tristetraprolin (TTP), an mRNA destabilizing protein prevents the translation of the mRNA transcripts, leading to a rapid degeneration of the target mRNA. We thus report that a direct TLR7 ligation by its agonist increases cytokine transcript signature but not an equivalent protein surge.
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Artritis Reumatoide , Receptor Toll-Like 7 , Linfocitos T CD8-positivos/metabolismo , Citocinas/metabolismo , Humanos , Mediadores de Inflamación , ARN Mensajero , Receptor Toll-Like 7/genética , Receptor Toll-Like 7/metabolismo , Receptores Toll-LikeRESUMEN
During the course of the COVID-19 pandemic, large-scale genome sequencing of SARS-CoV-2 has been useful in tracking its spread and in identifying variants of concern (VOC). Viral and host factors could contribute to variability within a host that can be captured in next-generation sequencing reads as intra-host single nucleotide variations (iSNVs). Analysing 1347 samples collected till June 2020, we recorded 16 410 iSNV sites throughout the SARS-CoV-2 genome. We found â¼42% of the iSNV sites to be reported as SNVs by 30 September 2020 in consensus sequences submitted to GISAID, which increased to â¼80% by 30th June 2021. Following this, analysis of another set of 1774 samples sequenced in India between November 2020 and May 2021 revealed that majority of the Delta (B.1.617.2) and Kappa (B.1.617.1) lineage-defining variations appeared as iSNVs before getting fixed in the population. Besides, mutations in RdRp as well as RNA-editing by APOBEC and ADAR deaminases seem to contribute to the differential prevalence of iSNVs in hosts. We also observe hyper-variability at functionally critical residues in Spike protein that could alter the antigenicity and may contribute to immune escape. Thus, tracking and functional annotation of iSNVs in ongoing genome surveillance programs could be important for early identification of potential variants of concern and actionable interventions.
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Evolución Molecular , Variación Genética/genética , Genoma Viral/genética , Interacciones Huésped-Patógeno/genética , SARS-CoV-2/genética , Desaminasas APOBEC-1/genética , Adenosina Desaminasa/genética , Animales , COVID-19/epidemiología , COVID-19/prevención & control , COVID-19/virología , Chlorocebus aethiops , ARN Polimerasa Dependiente de ARN de Coronavirus/genética , Bases de Datos Genéticas , Evasión Inmune/genética , India/epidemiología , Filogenia , Proteínas de Unión al ARN/genética , SARS-CoV-2/clasificación , SARS-CoV-2/crecimiento & desarrollo , Glicoproteína de la Espiga del Coronavirus/genética , Células VeroRESUMEN
The response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is largely impacted by the level of virus exposure and status of the host immunity. The nature of protection shown by direct asymptomatic contacts of coronavirus disease 2019 (COVID-19)-positive patients is quite intriguing. In this study, we have characterized the antibody titer, SARS-CoV-2 surrogate virus neutralization, cytokine levels, single-cell T-cell receptor (TCR), and B-cell receptor (BCR) profiling in asymptomatic direct contacts, infected cases, and controls. We observed significant increase in antibodies with neutralizing amplitude in asymptomatic contacts along with cytokines such as Eotaxin, granulocyte-colony stimulating factor (G-CSF), interleukin 7 (IL-7), migration inhibitory factor (MIF), and macrophage inflammatory protein-1α (MIP-1α). Upon single-cell RNA (scRNA) sequencing, we explored the dynamics of the adaptive immune response in few representative asymptomatic close contacts and COVID-19-infected patients. We reported direct asymptomatic contacts to have decreased CD4+ naive T cells with concomitant increase in CD4+ memory and CD8+ Temra cells along with expanded clonotypes compared to infected patients. Noticeable proportions of class switched memory B cells were also observed in them. Overall, these findings gave an insight into the nature of protection in asymptomatic contacts.
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Inmunidad Adaptativa/inmunología , COVID-19/inmunología , Genómica/métodos , SARS-CoV-2/inmunología , Análisis de la Célula Individual/métodos , Inmunidad Adaptativa/genética , Adulto , Anticuerpos Antivirales/inmunología , COVID-19/genética , COVID-19/virología , Citocinas/inmunología , Citocinas/metabolismo , Femenino , Perfilación de la Expresión Génica/métodos , Humanos , Masculino , Células B de Memoria/inmunología , Células B de Memoria/metabolismo , Células B de Memoria/virología , Persona de Mediana Edad , SARS-CoV-2/fisiología , Análisis de Secuencia de ARN/métodos , Linfocitos T/inmunología , Linfocitos T/metabolismo , Linfocitos T/virología , Adulto JovenRESUMEN
Purpose: Aurora kinase B (AURKB) plays a pivotal role in the regulation of mitosis and is gaining prominence as a therapeutic target in cancers; however, the role of AURKB in retinoblastoma (RB) has not been studied. The purpose of this study was to determine if AURKB plays a role in RB, how its expression is regulated, and whether it could be specifically targeted. Methods: The protein expression of AURKB was determined using immunohistochemistry in human RB patient specimens and immunoblotting in cell lines. Pharmacological inhibition and shRNA-mediated knockdown were used to understand the role of AURKB in cell viability, apoptosis, and cell cycle distribution. Cell viability in response to AURKB inhibition was also assessed in enucleated RB specimens. Immunoblotting was employed to determine the protein levels of phospho-histone H3, p53, p21, and MYCN. Chromatin immunoprecipitation-qPCR was performed to verify the binding of MYCN on the promoter region of AURKB. Results: The expression of AURKB was found to be markedly elevated in human RB tissues, and the overexpression significantly correlated with optic nerve and anterior chamber invasion. Targeting AURKB with small-molecule inhibitors and shRNAs resulted in reduced cell survival and increased apoptosis and cell cycle arrest at the G2/M phase. More importantly, primary RB specimens showed decreased cell viability in response to pharmacological AURKB inhibition. Additional studies have demonstrated that the MYCN oncogene regulates the expression of AURKB in RB. Conclusions: AURKB is overexpressed in RB, and targeting it could serve as a novel therapeutic strategy to restrict tumor cell growth.
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Aurora Quinasa B/genética , Regulación Enzimológica de la Expresión Génica/fisiología , Terapia Molecular Dirigida , Inhibidores de Proteínas Quinasas/farmacología , Neoplasias de la Retina/enzimología , Retinoblastoma/enzimología , Apoptosis/efectos de los fármacos , Compuestos Aza/farmacología , Benzamidas/farmacología , Ciclo Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Humanos , Immunoblotting , Inmunohistoquímica , Indoles/farmacología , Organofosfatos/farmacología , Quinazolinas/farmacología , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Neoplasias de la Retina/patología , Retinoblastoma/patología , Células Tumorales CultivadasRESUMEN
BACKGROUND: Rheumatoid arthritis (RA) is an autoimmune disorder with genetic and environmental causes often linked with the disease etiology. A disrupted metabolism has often been a characteristic of RA and an altered metabolic state of immune cells has been associated with their phenotypic and functional changes. The energy in the form of ATP produced by the metabolically active cells may thus initiate a cascade of immune responses there by influencing the disease pathogenesis or progression. AIM OF THE STUDY: Through this study we have focused on determining the role of ATP in etiology of RA and aberrant cellular functions. METHODS: Blood samples of 80 healthy controls (HC) and 95 RA patients were screened for extracellular ATP concentration, transcriptome analyses, an inflammatory mediator and the results were statistically analysed. RESULTS: In this study, ATP is shown to be excessive in the plasma of RA patients (453.5 ± 16.09% in RA vs. 233.9 ± 10.07% in HC, p <0.0001) and significantly increases with the disease severity. The abundant extracellular ATP could activate circulating cytotoxic CD8+T cells in RA patients to produce Granzyme B. CONCLUSION: Plasma ATP is thus identified to have a significant potential in progression and prognosis of RA and may thus be studied further to design better therapeutic approaches for the disease.
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Artritis Reumatoide , Adenosina Trifosfato , Artritis Reumatoide/sangre , Linfocitos T CD8-positivos , Humanos , Plasma , Índice de Severidad de la EnfermedadRESUMEN
Dendritic cell (DC) activation and cytokine production is tightly regulated. In this study, we found that Zbtb10 expression is activation dependent and it is essential for the immunogenic function of cDC1. Zbtb10 knockdown (KD) significantly reduced the expression of co-stimulatory genes CD80 and CD86 along with cytokines including IL-12, IL-6, and IL-10, in activated cDC1 Mutu-DC line. Consequently, the clonal expansion of CD44+ effector T cells in co-cultured CD4+ T cells was drastically reduced owing to significantly reduced IL-2. At the same time, these CD44+ effector T cells were unable to differentiate toward Tbet+ IFNγ+ Th1 subtype. Instead, an increased frequency of Th2 cells expressing GATA3+ and IL-13+ was observed. Interestingly, in Zbtb10 KD condition the co-cultured T cells depicted increased expression of PD1 and LAG3, the T-cell anergic markers. Moreover, the global transcriptome analysis identified that Zbtb10 is pertinent for DC activation and its depletion in cDC1 completely shuts down their immune responses. Mechanistic analysis revealed that Zbtb10 KD enhanced the expression of NKRF (NF-κB repressing factor) leading to drastic suppression of NF-κB related genes. Zbtb10 KD abrogated p65 and RelB nuclear translocation, thereby controlling the activation and maturation of cDC1 and the ensuing adaptive T cell responses.
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Citocinas/biosíntesis , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Factores de Transcripción/metabolismo , Animales , Biomarcadores , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Línea Celular , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Activación de Linfocitos/inmunología , Ratones , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismoRESUMEN
Rewiring tumor cells to undergo drug-induced apoptosis is a promising way to overcome chemoresistance. Therefore, identifying causative factors for chemoresistance is of high importance. Unbiased global proteome profiling of sensitive, early, and late cisplatin-resistant oral squamous cell carcinoma (OSCC) lines identified CMTM6 as a top-ranked upregulated protein. Analyses of OSCC patient tumor samples demonstrated significantly higher CMTM6 expression in chemotherapy (CT) nonresponders as compared with CT responders. In addition, a significant association between higher CMTM6 expression and poorer relapse-free survival in esophageal squamous cell carcinoma, head and neck squamous cell carcinoma, and lung squamous cell carcinoma was observed from Kaplan-Meier plot analysis. Stable knockdown (KD) of CMTM6 restored cisplatin-mediated cell death in chemoresistant OSCC lines. Upon CMTM6 overexpression in CMTM6-KD lines, the cisplatin-resistant phenotype was rescued. The patient-derived cell xenograft model of chemoresistant OSCC displaying CMTM6 depletion restored the cisplatin-induced cell death and tumor burden substantially. The transcriptome analysis of CMTM6-KD and control chemoresistant cells depicted enrichment of the Wnt signaling pathway. We demonstrated that CMTM6 interaction with membrane-bound Enolase-1 stabilized its expression, leading to activation of Wnt signaling mediated by AKT-glycogen synthase kinase-3ß. CMTM6 has been identified as a stabilizer of programmed cell death ligand 1. Therefore, as CMTM6 facilitates tumor cells for immune evasion and mediates cisplatin resistance, it could be a promising therapeutic target for treating therapy-resistant OSCC.
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Cisplatino/farmacología , Proteínas de Unión al ADN/metabolismo , Resistencia a Antineoplásicos/efectos de los fármacos , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Proteínas de la Mielina/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Vía de Señalización Wnt/efectos de los fármacos , Animales , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Biomarcadores de Tumor/metabolismo , Carcinoma de Células Escamosas/patología , Muerte Celular , Línea Celular Tumoral , Neoplasias Esofágicas/genética , Carcinoma de Células Escamosas de Esófago/tratamiento farmacológico , Carcinoma de Células Escamosas de Esófago/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Glucógeno Sintasa Quinasa 3 beta/genética , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Neoplasias de Cabeza y Cuello/genética , Humanos , Proteínas con Dominio MARVEL , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Neoplasias de la Boca/tratamiento farmacológico , Neoplasias de la Boca/patología , Proteínas de la Mielina/genética , Fosfopiruvato Hidratasa/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/tratamiento farmacológico , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Proteínas Supresoras de Tumor/metabolismo , Regulación hacia Arriba/efectos de los fármacosRESUMEN
OBJECTIVE: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has affected the whole world, including Odisha, a state in eastern India. Many people have migrated to the state from different countries as well as other states during this SARS-CoV-2 pandemic. The aim of this study was to analyse the receptor-binding domain (RBD) sequence of the spike protein from isolates collected from throat swab samples of COVID-19-positive patients and further to assess the RBD affinity for angiotensin-converting enzyme 2 (ACE2) of different species, including humans. METHODS: Whole-genome sequencing for 35 clinical SARS-CoV-2 isolates from COVID-19-positive patients was performed by ARTIC amplicon-based sequencing. Sequence analysis and phylogenetic analysis were performed for the spike region and the RBD region of all isolates. The interaction between the RBD and ACE2 of five different species was also analysed. RESULTS: The spike region of 32 isolates showed one or multiple alterations in nucleotide bases in comparison with the Wuhan reference strain. One of the identified mutations, at position 1204 (Ref A, RMRC 22 C), in the RBD coding region of the spike protein showed stronger binding affinity for human ACE2. Furthermore, RBDs of all the Indian isolates showed binding affinity for ACE2 of different species. CONCLUSION: As mutant RBD showed stronger interaction with human ACE2, it could potentially result in higher infectivity. The binding affinity of the RBDs for ACE2 of all five species studied suggests that the virus can infect a wide variety of animals, which could also act as natural reservoir for SARS-CoV-2.
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Enzima Convertidora de Angiotensina 2/análisis , Enzima Convertidora de Angiotensina 2/metabolismo , SARS-CoV-2/genética , Análisis de Secuencia , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/metabolismo , Secuenciación Completa del Genoma , Animales , Sitios de Unión , Humanos , India/epidemiología , Mutación , Filogenia , Unión Proteica , Dominios ProteicosRESUMEN
Plasmacytoid dendritic cells (DCs) are reported to induce robust type-I interferon (IFN) response, whereas cDC1 DCs develop moderate type-I IFN response upon TLR9 stimulation. It is very interesting to understand how this signaling under TLR9 is tightly regulated for the induction of type-I IFNs. Here, we report co-repressor protein NCoR1 as the major factor fine-tuning the signaling pathways regulating IFN-ß expression under TLR9 in cDC1 DCs. We found that NCoR1 knockdown induced a robust IFN-ß-mediated antiviral response upon TLR9 activation in cDC1 DCs. At the molecular level, we showed that NCoR1 directly repressed MyD88-IRF7 signaling axis in cDC1 cells. Therefore, NCoR1 depletion enhanced pIRF7 levels, IFN-ß secretion, and downstream pSTAT1-pSTAT2 signaling, leading to sustained induction of IFN stimulatory genes. Integrative genomic analysis depicted strong enrichment of an antiviral gene-module in CpG-activated NCoR1 knockdown DCs upon TLR9 activation. Moreover, we confirmed our findings in primary DCs derived from splenocytes of WT and NCoR1 DC-/- animals, which showed protection from Sendai and Vesicular Stomatitis viruses upon CpG activation. Ultimately, we identified that NCoR1-HDAC3 complex is involved in repressing the type-I IFN response in cDC1 DCs.
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Células Dendríticas/metabolismo , Factor 7 Regulador del Interferón/metabolismo , Interferón Tipo I/metabolismo , Factor 88 de Diferenciación Mieloide/metabolismo , Co-Represor 1 de Receptor Nuclear/metabolismo , Receptor Toll-Like 9/metabolismo , Animales , Células Cultivadas , Femenino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Transducción de Señal/fisiologíaRESUMEN
The autoimmune inflammatory disease, Rheumatoid arthritis (RA), has known imbalances in energy metabolism and superoxide levels thus may have an etiology associated with mitochondrial dysfunction. We thus evaluated the presence of a differential mitochondrial proteome as well as other characteristics including mitochondrial mass, membrane potential (Ψm), total cellular ATP and superoxide levels. Eighteen mitochondrial proteins were down-regulated while four were up-regulated in RA patients in comparison to the healthy controls (HC). A significant decrease in mitochondrial Ψm, superoxides and cellular ATP levels was observed in RA with constant mitochondrial mass suggesting mitochondrial dysfunction responsible for functional disparity in RA.
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Artritis Reumatoide/metabolismo , Mitocondrias/metabolismo , Proteínas Mitocondriales/metabolismo , Proteómica/métodos , Adenosina Trifosfato/metabolismo , Estudios de Casos y Controles , Cromatografía Liquida , Humanos , Potencial de la Membrana Mitocondrial , Mapas de Interacción de Proteínas , Superóxidos/metabolismo , Espectrometría de Masas en TándemRESUMEN
Circular RNAs (circRNAs) are a large family of noncoding RNAs that have emerged as novel regulators of gene expression. However, little is known about the function of circRNAs in pancreatic ß-cells. Here, transcriptomic analysis of mice pancreatic islet RNA-sequencing data identified 77 differentially expressed circRNAs between mice fed with a normal diet and a high-fat diet. Surprisingly, multiple circRNAs were derived from the intron 2 of the preproinsulin 2 (Ins2) gene and are termed as circular intronic (ci)-Ins2. The expression of ci-Ins2 transcripts in mouse pancreatic islets, and ßTC6 cells were confirmed by reverse transcription PCR, DNA sequencing, and RNase R treatment experiments. The level of ci-Ins2 was altered in ßTC6 cells upon exposure to elevated levels of palmitate and glucose. Computational analysis predicted the interaction of several RNA-binding proteins with ci-Ins2 and their flanking region, suggesting their role in the ci-Ins2 function or biogenesis. Additionally, bioinformatics analysis predicted the association of several microRNAs with ci-Ins2. Gene ontology and pathway analysis of genes targeted by miRNAs associated with ci-Ins2 suggested the regulation of several key biological processes. Together, our findings indicate that differential expression of circRNAs, especially ci-Ins2 transcripts, may regulate ß-cell function and may play a critical role in the development of diabetes.
Asunto(s)
Insulinas/genética , ARN Circular , Empalme Alternativo , Secuencia de Bases , Biología Computacional/métodos , Exones , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Células Secretoras de Insulina/metabolismo , Intrones , Interferencia de ARN , Empalme del ARN , Factores de Empalme de ARN/metabolismo , TranscriptomaRESUMEN
Krüppel-associated box zinc finger proteins (KZFPs) constitute the largest family of mammalian transcription factors, but most remain completely uncharacterized. While initially proposed to primarily repress transposable elements, recent reports have revealed that KFZPs contribute to a wide variety of other biological processes. Using murine and human in vitro and in vivo models, we demonstrate here that one poorly studied KZFP, ZFP30, promotes adipogenesis by directly targeting and activating a retrotransposon-derived Pparg2 enhancer. Through mechanistic studies, we further show that ZFP30 recruits the co-regulator KRAB-associated protein 1 (KAP1), which, surprisingly, acts as a ZFP30 co-activator in this adipogenic context. Our findings provide an understanding of both adipogenic and KZFP-KAP1 complex-mediated gene regulation, showing that the KZFP-KAP1 axis can also function in a non-repressive manner.