RESUMO
Insufficient telomerase activity, stemming from low telomerase reverse transcriptase (TERT) gene transcription, contributes to telomere dysfunction and aging pathologies. Besides its traditional function in telomere synthesis, TERT acts as a transcriptional co-regulator of genes pivotal in aging and age-associated diseases. Here, we report the identification of a TERT activator compound (TAC) that upregulates TERT transcription via the MEK/ERK/AP-1 cascade. In primary human cells and naturally aged mice, TAC-induced elevation of TERT levels promotes telomere synthesis, blunts tissue aging hallmarks with reduced cellular senescence and inflammatory cytokines, and silences p16INK4a expression via upregulation of DNMT3B-mediated promoter hypermethylation. In the brain, TAC alleviates neuroinflammation, increases neurotrophic factors, stimulates adult neurogenesis, and preserves cognitive function without evident toxicity, including cancer risk. Together, these findings underscore TERT's critical role in aging processes and provide preclinical proof of concept for physiological TERT activation as a strategy to mitigate multiple aging hallmarks and associated pathologies.
Assuntos
Envelhecimento , Metilação de DNA , Telomerase , Telomerase/metabolismo , Telomerase/genética , Humanos , Animais , Camundongos , DNA (Citosina-5-)-Metiltransferases/metabolismo , DNA (Citosina-5-)-Metiltransferases/genética , Senescência Celular , Regiões Promotoras Genéticas , DNA Metiltransferase 3B , Encéfalo/metabolismo , Telômero/metabolismo , Camundongos Endogâmicos C57BL , Masculino , Fator de Transcrição AP-1/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/genética , NeurogêneseRESUMO
Sex exerts a profound impact on cancer incidence, spectrum and outcomes, yet the molecular and genetic bases of such sex differences are ill-defined and presumptively ascribed to X-chromosome genes and sex hormones1. Such sex differences are particularly prominent in colorectal cancer (CRC) in which men experience higher metastases and mortality. A murine CRC model, engineered with an inducible transgene encoding oncogenic mutant KRASG12D and conditional null alleles of Apc and Trp53 tumour suppressors (designated iKAP)2, revealed higher metastases and worse outcomes specifically in males with oncogenic mutant KRAS (KRAS*) CRC. Integrated cross-species molecular and transcriptomic analyses identified Y-chromosome gene histone demethylase KDM5D as a transcriptionally upregulated gene driven by KRAS*-mediated activation of the STAT4 transcription factor. KDM5D-dependent chromatin mark and transcriptome changes showed repression of regulators of the epithelial cell tight junction and major histocompatibility complex class I complex components. Deletion of Kdm5d in iKAP cancer cells increased tight junction integrity, decreased cell invasiveness and enhanced cancer cell killing by CD8+ T cells. Conversely, iAP mice engineered with a Kdm5d transgene to provide constitutive Kdm5d expression specifically in iAP cancer cells showed an increased propensity for more invasive tumours in vivo. Thus, KRAS*-STAT4-mediated upregulation of Y chromosome KDM5D contributes substantially to the sex differences in KRAS* CRC by means of its disruption of cancer cell adhesion properties and tumour immunity, providing an actionable therapeutic strategy for metastasis risk reduction for men afflicted with KRAS* CRC.
Assuntos
Neoplasias Colorretais , Histona Desmetilases , Antígenos de Histocompatibilidade Menor , Caracteres Sexuais , Animais , Feminino , Humanos , Masculino , Camundongos , Linfócitos T CD8-Positivos/imunologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Modelos Animais de Doenças , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Camundongos Transgênicos , Antígenos de Histocompatibilidade Menor/genética , Antígenos de Histocompatibilidade Menor/metabolismo , Regulação para CimaRESUMO
During self-renewal, cell-type-defining features are drastically perturbed in mitosis and must be faithfully reestablished upon G1 entry, a process that remains largely elusive. Here, we characterized at a genome-wide scale the dynamic transcriptional and architectural resetting of mouse pluripotent stem cells (PSCs) upon mitotic exit. We captured distinct waves of transcriptional reactivation with rapid induction of stem cell genes and transient activation of lineage-specific genes. Topological reorganization at different hierarchical levels also occurred in an asynchronous manner and showed partial coordination with transcriptional resetting. Globally, rapid transcriptional and architectural resetting associated with mitotic retention of H3K27 acetylation, supporting a bookmarking function. Indeed, mitotic depletion of H3K27ac impaired the early reactivation of bookmarked, stem-cell-associated genes. However, 3D chromatin reorganization remained largely unaffected, suggesting that these processes are driven by distinct forces upon mitotic exit. This study uncovers principles and mediators of PSC molecular resetting during self-renewal.
Assuntos
Cromatina/genética , Código das Histonas/genética , Histonas/genética , Mitose/genética , Células-Tronco Pluripotentes/fisiologia , Acetilação , Animais , Linhagem Celular , Drosophila/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Transcrição Gênica/genética , Ativação Transcricional/genéticaRESUMO
Colorectal cancer has served as a genetic and biological paradigm for the evolution of solid tumors, and these insights have illuminated early detection, risk stratification, prevention, and treatment principles. Employing the hallmarks of cancer framework, we provide a conceptual framework to understand how genetic alterations in colorectal cancer drive cancer cell biology properties and shape the heterotypic interactions across cells in the tumor microenvironment. This review details research advances pertaining to the genetics and biology of colorectal cancer, emerging concepts gleaned from immune and single-cell profiling, and critical advances and remaining knowledge gaps influencing the development of effective therapies for this cancer that remains a major public health burden.
Assuntos
Biomarcadores Tumorais/genética , Neoplasias Colorretais/genética , Biomarcadores Tumorais/imunologia , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/fisiopatologia , Neoplasias Colorretais/terapia , Humanos , Mutação/genética , Pesquisa/tendências , Microambiente Tumoral/imunologiaRESUMO
Prostate cancer is a leading cause of cancer-related mortality in men. The widespread use of androgen receptor (AR) inhibitors has generated an increased incidence of AR-negative prostate cancer, triggering the need for effective therapies for such patients. Here, analysis of public genome-wide CRISPR screens in human prostate cancer cell lines identified histone demethylase JMJD1C (KDM3C) as an AR-negative context-specific vulnerability. Secondary validation studies in multiple cell lines and organoids, including isogenic models, confirmed that small hairpin RNA (shRNA)-mediated depletion of JMJD1C potently inhibited growth specifically in AR-negative prostate cancer cells. To explore the cooperative interactions of AR and JMJD1C, we performed comparative transcriptomics of 1) isogenic AR-positive versus AR-negative prostate cancer cells, 2) AR-positive versus AR-negative prostate cancer tumors, and 3) isogenic JMJD1C-expressing versus JMJD1C-depleted AR-negative prostate cancer cells. Loss of AR or JMJD1C generates a modest tumor necrosis factor alpha (TNFα) signature, whereas combined loss of AR and JMJD1C strongly up-regulates the TNFα signature in human prostate cancer, suggesting TNFα signaling as a point of convergence for the combined actions of AR and JMJD1C. Correspondingly, AR-negative prostate cancer cells showed exquisite sensitivity to TNFα treatment and, conversely, TNFα pathway inhibition via inhibition of its downstream effector MAP4K4 partially reversed the growth defect of JMJD1C-depleted AR-negative prostate cancer cells. Given the deleterious systemic side effects of TNFα therapy in humans and the viability of JMJD1C-knockout mice, the identification of JMJD1C inhibition as a specific vulnerability in AR-negative prostate cancer may provide an alternative drug target for prostate cancer patients progressing on AR inhibitor therapy.
Assuntos
Histona Desmetilases com o Domínio Jumonji/genética , Oxirredutases N-Desmetilantes/genética , Neoplasias da Próstata/genética , Receptores Androgênicos/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Bases de Dados Genéticas , Histona Desmetilases/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Histona Desmetilases com o Domínio Jumonji/metabolismo , Masculino , Oxirredutases N-Desmetilantes/metabolismo , Regiões Promotoras Genéticas/efeitos dos fármacos , Próstata/patologia , Proteínas Serina-Treonina Quinases/genética , Receptores Androgênicos/genética , Transdução de Sinais/efeitos dos fármacos , Ativação Transcricional/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Toxoplasma gondii secretes a group of rhoptry-secreted kinases (ROPs), which play significant roles in promoting intracellular infection. T. gondii rhoptry organelle protein 17 (ROP17) is one of these important effector proteins. However, its role in modulating host cell response during infection remains poorly understood. Here, we reveal that ROP17 (genotype I) induces significant changes in the expression genes and transcription factors of host cells. HEK293T cells were transfected with PCMV-N-HA-ROP17 plasmid or empty control PCMV-N-HA plasmid. Transcriptomic analysis revealed 3138 differentially expressed genes (DEGs) in PCMV-N-HA-ROP17-transfected HEK293T cells, including 1456 upregulated, 1682 downregulated DEGs. Also, 715 of the DEGs were transcription factors (TFs), including 423 downregulated TFs and 292 upregulated TFs. Most differentially expressed TFs, whether belong to signal transduction, cancer-related pathways or immune-related pathways, were downregulated in ROP17-expressing cells. ROP17 also decreased alternative splicing events in host cells, presumably via alteration of the expression of genes involved in the alternative splicing pathway. Taken together, our findings suggest a novel strategy whereby T. gondii ROP17 manipulates various cellular processes, including immune response through reprogramming host gene expression to promote its own colonization and survival in the infected host cells.
Assuntos
Imunidade Inata , Proteínas de Protozoários/metabolismo , Transdução de Sinais , Toxoplasma/imunologia , Toxoplasmose/parasitologia , Fatores de Virulência/metabolismo , Animais , Regulação para Baixo , Perfilação da Expressão Gênica , Células HEK293 , Humanos , Organelas/metabolismo , Proteínas de Protozoários/genética , Toxoplasma/fisiologia , Regulação para Cima , Fatores de Virulência/genéticaRESUMO
We constructed a new plasmid pIRESneo/ROP18/PLP1 that was injected intramuscularly into Kunming mice to evaluate its immune efficacy. The immunized mice exhibited significantly increased serum IgG2a levels, lymphocyte counts and Th1-type cytokine (IL-2, IL-12 and IFN-γ) levels. Moreover, the immunized mice exhibited longer survival times (44.7 ± 2.1 days for ROP18/PLP1 and 47.2 ± 2.9 days for ROP18/PLP1 + IL-18) and lower brain cyst burden (68.9% for ROP18/PLP1 and 72.4% for ROP18/PLP1 + IL-18) than control mice after T. gondii challenge. Our results demonstrate that the multiple-gene DNA vaccine including both ROP18 and PLP1 elicits greater protection against T. gondii challenge and stronger immunogenicity than single-gene vaccines.
Assuntos
Proteína Proteolipídica de Mielina/imunologia , Proteínas Serina-Treonina Quinases/imunologia , Vacinas Protozoárias , Toxoplasma/imunologia , Toxoplasmose Animal/prevenção & controle , Vacinas de DNA , Animais , Anticorpos Antiprotozoários/biossíntese , Encéfalo/parasitologia , Citocinas/análise , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Imunoglobulina G/análise , Imunoglobulina G/biossíntese , Injeções Intramusculares , Camundongos , Proteína Proteolipídica de Mielina/genética , Plasmídeos , Proteínas Serina-Treonina Quinases/genética , Proteínas de Protozoários , Vacinas Protozoárias/administração & dosagem , Vacinas Protozoárias/imunologia , Vacinas Protozoárias/normas , Organismos Livres de Patógenos Específicos , Baço/citologia , Baço/imunologia , Análise de Sobrevida , Toxoplasmose Animal/mortalidade , Vacinas de DNA/administração & dosagem , Vacinas de DNA/imunologia , Vacinas de DNA/normasRESUMO
BACKGROUND: Although the specific pathogenesis of preterm birth (PTB) has not been thoroughly clarified, it is known to be related to various factors, such as pregnancy complications, maternal socioeconomic factors, lifestyle habits, reproductive history, environmental and psychological factors, prenatal care, and nutritional status. PTB has serious implications for newborns and families and is associated with high mortality and complications. Therefore, the prediction of PTB risk can facilitate early intervention and reduce its resultant adverse consequences. AIM: To analyze the risk factors for PTB to establish a PTB risk prediction model and to assess postpartum anxiety and depression in mothers. METHODS: A retrospective analysis of 648 consecutive parturients who delivered at Shenzhen Bao'an District Songgang People's Hospital between January 2019 and January 2022 was performed. According to the diagnostic criteria for premature infants, the parturients were divided into a PTB group (n = 60) and a full-term (FT) group (n = 588). Puerperae were assessed by the Self-rating Anxiety Scale (SAS) and Self-rating Depression Scale (SDS), based on which the mothers with anxiety and depression symptoms were screened for further analysis. The factors affecting PTB were analyzed by univariate analysis, and the related risk factors were identified by logistic regression. RESULTS: According to univariate analysis, the PTB group was older than the FT group, with a smaller weight change and greater proportions of women who underwent artificial insemination and had gestational diabetes mellitus (P < 0.05). In addition, greater proportions of women with reproductive tract infections and greater white blood cell (WBC) counts (P < 0.05), shorter cervical lengths in the second trimester and lower neutrophil percentages (P < 0.001) were detected in the PTB group than in the FT group. The PTB group exhibited higher postpartum SAS and SDS scores than did the FT group (P < 0.0001), with a higher number of mothers experiencing anxiety and depression (P < 0.001). Multivariate logistic regression analysis revealed that a greater maternal weight change, the presence of gestational diabetes mellitus, a shorter cervical length in the second trimester, a greater WBC count, and the presence of maternal anxiety and depression were risk factors for PTB (P < 0.01). Moreover, the risk score of the FT group was lower than that of the PTB group, and the area under the curve of the risk score for predicting PTB was greater than 0.9. CONCLUSION: This study highlights the complex interplay between postpartum anxiety and PTB, where maternal anxiety may be a potential risk factor for PTB, with PTB potentially increasing the incidence of postpartum anxiety in mothers. In addition, a greater maternal weight change, the presence of gestational diabetes mellitus, a shorter cervical length, a greater WBC count, and postpartum anxiety and depression were identified as risk factors for PTB.
RESUMO
Telomere dynamics are linked to aging hallmarks, and age-associated telomere loss fuels the development of epithelial cancers. In Apc-mutant mice, the onset of DNA damage associated with telomere dysfunction has been shown to accelerate adenoma initiation via unknown mechanisms. Here, we observed that Apc-mutant mice engineered to experience telomere dysfunction show accelerated adenoma formation resulting from augmented cell competition and clonal expansion. Mechanistically, telomere dysfunction induces the repression of EZH2, resulting in the derepression of Wnt antagonists, which causes the differentiation of adjacent stem cells and a relative growth advantage to Apc-deficient telomere dysfunctional cells. Correspondingly, in this mouse model, GSK3ß inhibition countered the actions of Wnt antagonists on intestinal stem cells, resulting in impaired adenoma formation of telomere dysfunctional Apc-mutant cells. Thus, telomere dysfunction contributes to cancer initiation through altered stem cell dynamics, identifying an interception strategy for human APC-mutant cancers with shortened telomeres.
Assuntos
Proteína da Polipose Adenomatosa do Colo , Células-Tronco , Telômero , Animais , Camundongos , Telômero/metabolismo , Proteína da Polipose Adenomatosa do Colo/genética , Proteína da Polipose Adenomatosa do Colo/metabolismo , Células-Tronco/metabolismo , Células-Tronco/patologia , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Adenoma/patologia , Adenoma/genética , Adenoma/metabolismo , Intestinos/patologia , Diferenciação Celular , Humanos , Glicogênio Sintase Quinase 3 beta/metabolismo , Glicogênio Sintase Quinase 3 beta/genética , Dano ao DNA , Camundongos Endogâmicos C57BL , Via de Sinalização WntRESUMO
Inactivation of adenomatous polyposis coli (APC) is common across many cancer types and serves as a critical initiating event in most sporadic colorectal cancers. APC deficiency activates WNT signaling, which remains an elusive target for cancer therapy, prompting us to apply the synthetic essentiality framework to identify druggable vulnerabilities for APC-deficient cancers. Tryptophan 2,3-dioxygenase 2 (TDO2) was identified as a synthetic essential effector of APC-deficient colorectal cancer. Mechanistically, APC deficiency results in the TCF4/ß-catenin-mediated upregulation of TDO2 gene transcription. TDO2 in turn activates the Kyn-AhR pathway, which increases glycolysis to drive anabolic cancer cell growth and CXCL5 secretion to recruit macrophages into the tumor microenvironment. Therapeutically, APC-deficient colorectal cancer models were susceptible to TDO2 depletion or pharmacologic inhibition, which impaired cancer cell proliferation and enhanced antitumor immune profiles. Thus, APC deficiency activates a TCF4-TDO2-AhR-CXCL5 circuit that affects multiple cancer hallmarks via autonomous and nonautonomous mechanisms and illuminates a genotype-specific vulnerability in colorectal cancer. SIGNIFICANCE: This study identifies critical effectors in the maintenance of APC-deficient colorectal cancer and demonstrates the relationship between APC/WNT pathway and kynurenine pathway signaling. It further determines the tumor-associated macrophage biology in APC-deficient colorectal cancer, informing genotype-specific therapeutic targets and the use of TDO2 inhibitors. This article is highlighted in the In This Issue feature, p. 1599.
Assuntos
Polipose Adenomatosa do Colo , Neoplasias Colorretais , Dioxigenases , Polipose Adenomatosa do Colo/genética , Polipose Adenomatosa do Colo/metabolismo , Polipose Adenomatosa do Colo/patologia , Neoplasias Colorretais/metabolismo , Dioxigenases/metabolismo , Humanos , Triptofano , Triptofano Oxigenase/metabolismo , Microambiente Tumoral , Via de Sinalização Wnt/genética , beta Catenina/genética , beta Catenina/metabolismoRESUMO
BACKGROUND: Tumor-associated macrophages (TAMs) are key regulators of the complex interplay between cancer and the immune microenvironment. Tumor cell-derived spondin 2 (SPON2) is an extracellular matrix glycoprotein that has complicated roles in recruitment of macrophages and neutrophils during inflammation. Overexpression of SPON2 has been shown to promote tumor cell migration in colorectal cancer (CRC). However, the mechanism by which SPON2 regulates the accumulation of TAMs in the tumor microenvironment (TME) of CRC is unknown. METHODS: Immunohistochemistry was used to examine SPON2 expression in clinical CRC tissues. In vitro migration assays, transendothelial migration assays (iTEM), and cell adhesion assays were used to investigate the effects of SPON2 on monocyte/macrophage migration. Subcutaneous tumor formation and orthotopic implantation assays were performed in C57 BL/6 mice to confirm the effects of SPON2 on TAM infiltration in tumors. RESULTS: SPON2 expression is positively correlated with M2-TAM infiltration in clinical CRC tumors and poor prognosis of CRC patients. In addition, SPON2 promotes cytoskeletal remodeling and transendothelial migration of monocytes by activating integrin ß1/PYK2 axis. SPON2 may indirectly induce M2-polarization through upregulating cytokines including IL10, CCL2 and CSF1 expression in tumor cells. Blocking M2 polarization and Macrophage depletion inhibited the SPON2-induced tumors growth and invasion. Furthermore, blocking the SPON2/integrin ß1/PYK2 axis impairs the transendothelial migration of monocytes and cancer-promoting functions of TAMs in vivo. CONCLUSIONS: Our findings demonstrate that SPON2-driven M2-TAM infiltration plays an important role during CRC tumor growth and metastasis. SPON2 may be a valuable biomarker guiding the use of macrophage-targeting strategies and a potential therapeutic target in advanced CRC.
Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/patologia , Proteínas da Matriz Extracelular/metabolismo , Quinase 2 de Adesão Focal/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/secundário , Proteínas de Neoplasias/metabolismo , Macrófagos Associados a Tumor/imunologia , Animais , Apoptose , Biomarcadores Tumorais/genética , Proliferação de Células , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/metabolismo , Proteínas da Matriz Extracelular/genética , Feminino , Quinase 2 de Adesão Focal/genética , Humanos , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Proteínas de Neoplasias/genética , Prognóstico , Taxa de Sobrevida , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Amyloid-induced neurodegeneration plays a central role in Alzheimer's disease (AD) pathogenesis. Here, we show that telomerase reverse transcriptase (TERT) haploinsufficiency decreases BDNF and increases amyloid-ß (Aß) precursor in murine brain. Moreover, prior to disease onset, the TERT locus sustains accumulation of repressive epigenetic marks in murine and human AD neurons, implicating TERT repression in amyloid-induced neurodegeneration. To test the impact of sustained TERT expression on AD pathobiology, AD mouse models were engineered to maintain physiological levels of TERT in adult neurons, resulting in reduced Aß accumulation, improved spine morphology, and preserved cognitive function. Mechanistically, integrated profiling revealed that TERT interacts with ß-catenin and RNA polymerase II at gene promoters and upregulates gene networks governing synaptic signaling and learning processes. These TERT-directed transcriptional activities do not require its catalytic activity nor telomerase RNA. These findings provide genetic proof-of-concept for somatic TERT gene activation therapy in attenuating AD progression including cognitive decline.
Assuntos
Doença de Alzheimer , Telomerase , Camundongos , Humanos , Animais , Doença de Alzheimer/genética , Telomerase/genética , Peptídeos beta-Amiloides/metabolismo , Cognição , Neurônios/metabolismoRESUMO
Loss of imprinting (LOI) results in severe developmental defects, but the mechanisms preventing LOI remain incompletely understood. Here, we dissect the functional components of the imprinting control region of the essential Dlk1-Dio3 locus (called IG-DMR) in pluripotent stem cells. We demonstrate that the IG-DMR consists of two antagonistic elements: a paternally methylated CpG island that prevents recruitment of TET dioxygenases and a maternally unmethylated non-canonical enhancer that ensures expression of the Gtl2 lncRNA by counteracting de novo DNA methyltransferases. Genetic or epigenetic editing of these elements leads to distinct LOI phenotypes with characteristic alternations of allele-specific gene expression, DNA methylation, and 3D chromatin topology. Although repression of the Gtl2 promoter results in dysregulated imprinting, the stability of LOI phenotypes depends on the IG-DMR, suggesting a functional hierarchy. These findings establish the IG-DMR as a bipartite control element that maintains imprinting by allele-specific restriction of the DNA (de)methylation machinery.
Assuntos
Alelos , Proteínas de Ligação ao Cálcio/genética , Metilação de DNA/genética , Peptídeos e Proteínas de Sinalização Intercelular/genética , Animais , Cromossomos/genética , Impressão Genômica/genética , Iodeto Peroxidase/genética , Camundongos , Regiões Promotoras Genéticas/genética , RNA Longo não Codificante/genéticaRESUMO
Toxoplasma gondii secretes a number of virulence-related effector proteins, such as the rhoptry protein 18 (ROP18). To further broaden our understanding of the molecular functions of ROP18, we examined the transcriptional response of human embryonic kidney cells (HEK293T) to ROP18 of type I T. gondii RH strain. Using RNA-sequencing, we compared the transcriptome of ROP18-expressing HEK293T cells to control HEK293T cells. Our analysis revealed that ROP18 altered the expression of 750 genes (467 upregulated genes and 283 downregulated genes) in HEK293T cells. Gene ontology (GO) and pathway enrichment analyses showed that differentially expressed genes (DEGs) were significantly enriched in extracellular matrix- and immune-related GO terms and pathways. KEGG pathway enrichment analysis revealed that DEGs were involved in several disease-related pathways, such as nervous system diseases and eye disease. ROP18 significantly increased the alternative splicing pattern "retained intron" and altered the expression of 144 transcription factors (TFs). These results provide new insight into how ROP18 may influence biological processes in the host cells via altering the expression of genes, TFs, and pathways. More in vitro and in vivo studies are required to substantiate these findings.
Assuntos
Toxoplasma , Células HEK293 , Humanos , Proteínas de Protozoários , Toxoplasma/genética , Virulência , Fatores de VirulênciaRESUMO
BACKGROUND: Infection with the apicomplexan protozoan parasite T. gondii can cause severe and potentially fatal cerebral and ocular disease, especially in immunocompromised individuals. The anticoccidial ionophore drug monensin has been shown to have anti-Toxoplasma gondii properties. However, the comprehensive molecular mechanisms that underlie the effect of monensin on T. gondii are still largely unknown. We hypothesized that analysis of T. gondii transcriptional changes induced by monensin treatment can reveal new aspects of the mechanism of action of monensin against T. gondii. METHODS: Porcine kidney (PK)-15 cells were infected with tachyzoites of T. gondii RH strain. Three hours post-infection, PK-15 cells were treated with 0.1 µM monensin, while control cells were treated with medium only. PK-15 cells containing intracellular tachyzoites were harvested at 6 and 24 h post-treatment, and the transcriptomic profiles of T. gondii-infected PK-15 cells were examined using high-throughput RNA sequencing (RNA-seq). Quantitative real-time PCR was used to verify the expression of 15 differentially expressed genes (DEGs) identified by RNA-seq analysis. RESULTS: A total of 4868 downregulated genes and three upregulated genes were identified in monensin-treated T. gondii, indicating that most of T. gondii genes were suppressed by monensin. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of T. gondii DEGs showed that T. gondii metabolic and cellular pathways were significantly downregulated. Spliceosome, ribosome, and protein processing in endoplasmic reticulum were the top three most significantly enriched pathways out of the 30 highly enriched pathways detected in T. gondii. This result suggests that monensin, via down-regulation of protein biosynthesis in T. gondii, can limit the parasite growth and proliferation. CONCLUSIONS: Our findings provide a comprehensive insight into T. gondii genes and pathways with altered expression following monensin treatment. These data can be further explored to achieve better understanding of the specific mechanism of action of monensin against T. gondii.
Assuntos
Interações Hospedeiro-Parasita/genética , Monensin/farmacologia , Toxoplasma/efeitos dos fármacos , Toxoplasma/genética , Transcriptoma , Animais , Linhagem Celular , Regulação para Baixo , Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Rim/citologia , Suínos , Toxoplasmose Animal , Regulação para CimaRESUMO
Toxoplasma gondii is a leading cause of foodborne illness and consumption of undercooked pig meat is a major risk factor for acquiring toxoplasmosis, which causes a substantial burden on society. Here, we used isobaric tags for relative and absolute quantification (iTRAQ) labelling coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify cellular proteins and pathways altered during T. gondii infection in pigs. We also used parallel reaction monitoring-based LC-MS/MS to verify the levels of protein expression of infected spleens and mesenteric lymph nodes (MLNs). At 6 days post-infection (dpi), 156, 391, 170, 292, and 200 differentially expressed proteins (DEPs) were detected in the brain, liver, lung, MLNs and spleen, respectively. At 18 dpi, 339, 351, 483, 388, and 303 DEPs were detected in the brain, liver, lung, MLNs and spleen, respectively. Although proteins involved in immune responses were upregulated in all infected tissues, protein expression signature in infected livers was dominated by downregulation of the metabolic processes. By weighted gene co-expression network analysis, we could further show that all proteins were clustered into 25 co-expression modules and that the pink module significantly correlated with the infection status. We also identified 163 potential anti-T. gondii proteins (PATPs) and provided evidence that two PATPs (HSP70.2 and PDIA3) can reduce T. gondii burden in porcine macrophages in vitro. This comprehensive proteomics analysis reveals new facets in the pathogenesis of T. gondii infection and identifies key proteins that may contribute to the pig's defense against this infection.
RESUMO
Oncogenic KRAS (KRAS*) is a key tumor maintenance gene in pancreatic ductal adenocarcinoma (PDAC), motivating pharmacologic targeting of KRAS* and its effectors. Here, we explored mechanisms involving the tumor microenvironment (TME) as a potential basis for resistance to targeting KRAS*. Using the inducible Kras G12D;Trp53 -/- PDAC mouse model, gain-of-function screens of epigenetic regulators identified HDAC5 as the top hit enabling KRAS* independent tumor growth. HDAC5-driven escaper tumors showed a prominent neutrophil-to-macrophage switch relative to KRAS*-driven tumors. Mechanistically, HDAC5 represses Socs3, a negative regulator of chemokine CCL2, resulting in increased CCL2, which recruits CCR2+ macrophages. Correspondingly, enforced Ccl2 promotes macrophage recruitment into the TME and enables tumor recurrence following KRAS* extinction. These tumor-associated macrophages in turn provide cancer cells with trophic support including TGFß to enable KRAS* bypass in a SMAD4-dependent manner. Our work uncovers a KRAS* resistance mechanism involving immune cell remodeling of the PDAC TME. SIGNIFICANCE: Although KRAS* is required for PDAC tumor maintenance, tumors can recur following KRAS* extinction. The capacity of PDAC cancer cells to alter the TME myeloid cell composition to support KRAS*-independent tumor growth illuminates novel therapeutic targets that may enhance the effectiveness of therapies targeting KRAS* and its pathway components.See related commentary by Carr and Fernandez-Zapico, p. 910.This article is highlighted in the In This Issue feature, p. 890.
Assuntos
Oncogenes/fisiologia , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Humanos , Neoplasias Pancreáticas/patologia , Microambiente Tumoral , Neoplasias PancreáticasRESUMO
Genetic inactivation of PTEN is common in prostate cancer and correlates with poorer prognosis. We previously identified CHD1 as an essential gene in PTEN-deficient cancer cells. Here, we sought definitive in vivo genetic evidence for, and mechanistic understanding of, the essential role of CHD1 in PTEN-deficient prostate cancer. In Pten and Pten/Smad4 genetically engineered mouse models, prostate-specific deletion of Chd1 resulted in markedly delayed tumor progression and prolonged survival. Chd1 deletion was associated with profound tumor microenvironment (TME) remodeling characterized by reduced myeloid-derived suppressor cells (MDSC) and increased CD8+ T cells. Further analysis identified IL6 as a key transcriptional target of CHD1, which plays a major role in recruitment of immunosuppressive MDSCs. Given the prominent role of MDSCs in suppressing responsiveness to immune checkpoint inhibitors (ICI), our genetic and tumor biological findings support combined testing of anti-IL6 and ICI therapies, specifically in PTEN-deficient prostate cancer. SIGNIFICANCE: We demonstrate a critical role of CHD1 in MDSC recruitment and discover CHD1/IL6 as a major regulator of the immunosuppressive TME of PTEN-deficient prostate cancer. Pharmacologic inhibition of IL6 in combination with immune checkpoint blockade elicits robust antitumor responses in prostate cancer.This article is highlighted in the In This Issue feature, p. 1241.
Assuntos
Proteínas de Ligação a DNA/metabolismo , PTEN Fosfo-Hidrolase/genética , Neoplasias da Próstata/genética , Evasão Tumoral/genética , Microambiente Tumoral/imunologia , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica/imunologia , Humanos , Masculino , Camundongos Transgênicos , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/patologia , Proteína Smad4/genética , Microambiente Tumoral/genéticaRESUMO
Germline telomere maintenance defects are associated with an increased incidence of inflammatory diseases in humans, yet whether and how telomere dysfunction causes inflammation are not known. Here, we show that telomere dysfunction drives pATM/c-ABL-mediated activation of the YAP1 transcription factor, up-regulating the major pro-inflammatory factor, pro-IL-18. The colonic microbiome stimulates cytosolic receptors activating caspase-1 which cleaves pro-IL-18 into mature IL-18, leading to recruitment of interferon (IFN)-γ-secreting T cells and intestinal inflammation. Correspondingly, patients with germline telomere maintenance defects exhibit DNA damage (γH2AX) signaling together with elevated YAP1 and IL-18 expression. In mice with telomere dysfunction, telomerase reactivation in the intestinal epithelium or pharmacological inhibition of ATM, YAP1, or caspase-1 as well as antibiotic treatment, dramatically reduces IL-18 and intestinal inflammation. Thus, telomere dysfunction-induced activation of the ATM-YAP1-pro-IL-18 pathway in epithelium is a key instigator of tissue inflammation.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ciclo Celular/metabolismo , Inflamação/patologia , Telômero/patologia , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Antibacterianos/uso terapêutico , Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Caspase 1/metabolismo , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/genética , Criança , Colo/metabolismo , Colo/microbiologia , Colo/patologia , Gastroenteropatias/patologia , Microbioma Gastrointestinal/efeitos dos fármacos , Microbioma Gastrointestinal/fisiologia , Humanos , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Inflamação/microbiologia , Interleucina-18/genética , Interleucina-18/metabolismo , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Camundongos , Camundongos Mutantes , Fosforilação , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , Transdução de Sinais , Telomerase/genética , Telomerase/metabolismo , Proteínas de Sinalização YAPRESUMO
We characterized the porcine tissue transcriptional landscapes that follow Toxoplasma gondii infection. RNAs were isolated from liver, spleen, cerebral cortex, lung, and mesenteric lymph nodes (MLNs) of T. gondii-infected and uninfected (control) pigs at days 6 and 18 postinfection, and were analyzed using next-generation sequencing (RNA-seq). T. gondii altered the expression of 178, 476, 199, 201, and 362 transcripts at 6 dpi and 217, 223, 347, 119, and 161 at 18 dpi in the infected brain, liver, lung, MLNs and spleen, respectively. The differentially expressed transcripts (DETs) were grouped into five expression patterns and 10 sub-clusters. Gene Ontology enrichment and pathway analysis revealed that immune-related genes dominated the overall transcriptomic signature and that metabolic processes, such as steroid biosynthesis, and metabolism of lipid and carboxylic acid, were downregulated in infected tissues. Co-expression network analysis identified transcriptional modules associated with host immune response to infection. These findings not only show how T. gondii infection alters porcine transcriptome in a tissue-specific manner, but also offer a gateway for testing new hypotheses regarding human response to T. gondii infection.