RESUMO
YAP plays a vital role in controlling growth and differentiation in various cell lineages. Although the expression of YAP in mice testicular and spermatogenic cells suggests its role in mammalian spermatogenesis, the role of YAP in the development of human male germ cells has not yet been determined. Using an in vitro model and a gene editing approach, we generated human spermatogonia stem cell-like cells (hSSLCs) from human embryonic stem cells (hESCs) and investigated the role of YAP in human spermatogenesis. The results showed that reducing YAP expression during the early stage of spermatogenic differentiation increased the number of PLZF+ hSSLCs and haploid spermatid-like cells. We also demonstrated that the up-regulation of YAP is essential for maintaining spermatogenic cell survival during the later stages of spermatogenic differentiation. The expression of YAP that deviates from this pattern results in a lower number of hSSLCs and an increased level of spermatogenic cell death. Taken together, our result demonstrates that the dynamic expression pattern of YAP is essential for human spermatogenesis. Modulating the level of YAP during human spermatogenesis could improve the production yield of male germ cells derived from hESCs, which could provide the optimization method for in vitro gametogenesis and gain insight into the application in the treatment of male infertility.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Diferenciação Celular , Células-Tronco Embrionárias Humanas , Espermatogênese , Fatores de Transcrição , Proteínas de Sinalização YAP , Masculino , Humanos , Células-Tronco Embrionárias Humanas/metabolismo , Células-Tronco Embrionárias Humanas/citologia , Proteínas de Sinalização YAP/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Espermatogônias/metabolismo , Espermatogônias/citologia , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Proteína com Dedos de Zinco da Leucemia Promielocítica/genéticaRESUMO
Lung adenocarcinoma is a malignant tumor with high morbidity and mortality. ZBTB16 plays a double role in various tumors; however, the potential mechanism of ZBTB16 in the pathophysiology of lung adenocarcinoma has yet to be elucidated. We herein observed a decreased expression of ZBTB16 mRNA and protein in lung adenocarcinoma and a significantly increased DNA methylation level of ZBTB16 in patients with lung adenocarcinoma. Analysis of public databases and patients' clinical data indicated a close association between ZBTB16 and patient survival. Ectopic expression of ZBTB16 in lung adenocarcinoma cells significantly inhibited cell proliferation, invasion, and migration. It also induced cell cycle arrest in the S phase. Meanwhile, mitotic catastrophe was induced, and DNA damage and apoptosis occurred. In line with these findings, the overexpression of ZBTB16 in xenograft mice resulted in the inhibition of tumor growth. Comprehensive analysis showed that WDHD1 was a potential target for ZBTB16. The overexpression of both isoforms of WDHD1 significantly reversed the ZBTB16-mediated inhibition of lung adenocarcinoma proliferation and cell cycle. These studies suggest that ZBTB16 impedes the progression of lung adenocarcinoma by interfering with WDHD1 transcription, making it a potential novel therapeutic target in the management of lung adenocarcinoma.
Assuntos
Adenocarcinoma de Pulmão , Pontos de Checagem do Ciclo Celular , Proliferação de Células , Replicação do DNA , Neoplasias Pulmonares , Animais , Feminino , Humanos , Masculino , Camundongos , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/patologia , Adenocarcinoma de Pulmão/metabolismo , Apoptose/genética , Pontos de Checagem do Ciclo Celular/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Metilação de DNA , Replicação do DNA/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/metabolismo , Camundongos Nus , Proteína com Dedos de Zinco da Leucemia Promielocítica/genética , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Transcrição Gênica/genéticaRESUMO
Glucocorticoids are important for proper organ maturation, and their levels are tightly regulated during development. Here, we use human cerebral organoids and mice to study the cell-type-specific effects of glucocorticoids on neurogenesis. We show that glucocorticoids increase a specific type of basal progenitors (co-expressing PAX6 and EOMES) that has been shown to contribute to cortical expansion in gyrified species. This effect is mediated via the transcription factor ZBTB16 and leads to increased production of neurons. A phenome-wide Mendelian randomization analysis of an enhancer variant that moderates glucocorticoid-induced ZBTB16 levels reveals causal relationships with higher educational attainment and altered brain structure. The relationship with postnatal cognition is also supported by data from a prospective pregnancy cohort study. This work provides a cellular and molecular pathway for the effects of glucocorticoids on human neurogenesis that relates to lasting postnatal phenotypes.
Assuntos
Córtex Cerebral , Glucocorticoides , Neurogênese , Proteína com Dedos de Zinco da Leucemia Promielocítica , Neurogênese/efeitos dos fármacos , Neurogênese/fisiologia , Humanos , Animais , Camundongos , Glucocorticoides/farmacologia , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/metabolismo , Córtex Cerebral/citologia , Feminino , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Gravidez , Neurônios/metabolismo , Neurônios/efeitos dos fármacos , Organoides/efeitos dos fármacos , Organoides/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Células-Tronco Neurais/efeitos dos fármacos , Células-Tronco Neurais/metabolismo , MasculinoRESUMO
The transcription factor promyelocytic leukemia zinc finger (PLZF, also known as ZBTB16) is critical for the self-renewal of spermatogonial stem cells (SSCs). However, the function of PLZF in SSCs is not clear. Here, we found that PLZF acted as an epigenetic regulator of stem cell maintenance and self-renewal of germ cells. The PLZF protein interacts with the ten-eleven translocation 1 (TET1) protein and subsequently acts as a modulator to regulate the expression of self-renewal-related genes. Furthermore, Transcription Factor 7-like 2 (TCF7L2) is promoted by the coordination of PLZF and Tri-methylation of lysine 4 on histone H3 (H3K4me3). In addition, testicular single-cell sequencing indicated that TCF7L2 is commonly expressed in the PLZF cluster. We demonstrated that PLZF directly targets TCF7L2 and alters the landscape of histone methylation in the SSCs nucleus. Meanwhile, the RD domain and Zn finger domain of PLZF synergize with H3K4me3 and directly upregulate TCF7L2 expression at the transcriptional level. Additionally, we identified a new association between PLZF and the histone methyltransferase EZH2 at the genomic level. Our study identified a new association between PLZF and H3K4me3, established the novel PLZF&TET1-H3K4me3-TCF7L2 axis at the genomic level which regulates undifferentiated spermatogonia, and provided a platform for studying germ cell development in male domestic animals.
Assuntos
Fatores de Transcrição Kruppel-Like , Espermatogônias , Masculino , Animais , Espermatogônias/metabolismo , Proteína com Dedos de Zinco da Leucemia Promielocítica/genética , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Fatores de Transcrição Kruppel-Like/genética , Testículo/metabolismo , Fatores de Transcrição/metabolismoRESUMO
PURPOSE: Androgen-refractory prostate cancer (ARPC) is one of the aggressive human cancers with metastatic capacity and resistance to androgen deprivation therapy (ADT). The present study investigated the genes responsible for ARPC progression and ADT resistance, and their regulatory mechanisms. METHODS: Transcriptome analysis, co-immunoprecipitation, confocal microscopy, and FACS analysis were performed to determine differentially-expressed genes, integrin α3ß4 heterodimer, and cancer stem cell (CSC) population. miRNA array, 3'-UTR reporter assay, ChIP assay, qPCR, and immunoblotting were used to determine differentially-expressed microRNAs, their binding to integrin transcripts, and gene expressions. A xenograft tumor model was used to assess tumor growth and metastasis. RESULTS: Metastatic ARPC cell lines (PC-3 and DU145) exhibiting significant downregulation of ZBTB16 and AR showed significantly upregulated ITGA3 and ITGB4. Silencing either one of the integrin α3ß4 heterodimer significantly suppressed ARPC survival and CSC population. miRNA array and 3'-UTR reporter assay revealed that miR-200c-3p, the most strongly downregulated miRNA in ARPCs, directly bound to 3'-UTR of ITGA3 and ITGB4 to inhibit the gene expression. Concurrently, miR-200c-3p also increased PLZF expression, which, in turn, inhibited integrin α3ß4 expression. Combination treatment with miR-200c-3p mimic and AR inhibitor enzalutamide showed synergistic inhibitory effects on ARPC cell survival in vitro and tumour growth and metastasis of ARPC xenografts in vivo, and the combination effect was greater than the mimic alone. CONCLUSION: This study demonstrated that miR-200c-3p treatment of ARPC is a promising therapeutic approach to restore the sensitivity to anti-androgen therapy and inhibit tumor growth and metastasis.
Assuntos
MicroRNAs , Neoplasias da Próstata , Masculino , Humanos , Regulação para Baixo/genética , Neoplasias da Próstata/patologia , Androgênios , Proteína com Dedos de Zinco da Leucemia Promielocítica/genética , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Antagonistas de Androgênios , Linhagem Celular Tumoral , MicroRNAs/genética , MicroRNAs/metabolismo , Regulação Neoplásica da Expressão Gênica , Proliferação de Células/genéticaRESUMO
In the process of orthodontic tooth movement (OTM), periodontal ligament fibroblasts (PDLFs) must undergo osteogenic differentiation. OTM increased the expression of Zinc finger and BTB domain-containing 16 (ZBTB16), which is implicated in osteogenic differentiation. Our goal was to investigate the mechanism of PDLF osteogenic differentiation mediated by ZBTB16. The OTM rat model was established, and PDLFs were isolated and exposed to mechanical force. Hematoxylin-eosin staining, Alizarin Red staining, immunofluorescence, and immunohistochemistry were carried out. The alkaline phosphatase (ALP) activity was measured. Dual-luciferase reporter gene assay and chromatin immunoprecipitation assay were conducted. In OTM models, ZBTB16 was significantly expressed. Additionally, there was an uneven distribution of PDLFs in the OTM group, as well as an increase in fibroblasts and inflammatory infiltration. ZBTB16 interference hindered PDLF osteogenic differentiation and decreased Wnt and ß-catenin levels. Meanwhile, ZBTB16 activated the Wnt/ß-catenin pathway. ZBTB16 also enhanced the expression of the osteogenic molecules osterix, osteocalcin (OCN), osteopontin (OPN), and bone sialo protein (BSP) at mRNA and protein levels. The interactions between Wnt1 and ZBTB16, as well as GCN5 and ZBTB16, were also verified. The adeno-associated virus-shZBTB16 injection also proved to inhibit osteogenic differentiation and reduce tooth movement distance in in vivo tests. ZBTB16 was up-regulated in OTM. Through acetylation modification of ZBTB16, GCN5 regulated the Wnt/ß-catenin signaling pathway and further mediated PDLF osteogenic differentiation.
Assuntos
Osteogênese , beta Catenina , Ratos , Animais , Osteogênese/genética , beta Catenina/metabolismo , Acetilação , Técnicas de Movimentação Dentária , Ligamento Periodontal , Via de Sinalização Wnt/genética , Diferenciação Celular , Células Cultivadas , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Histona Acetiltransferases/metabolismoRESUMO
The cryopreservation of spermatogonia stem cells (SSCs) has been widely used as an alternative treatment for infertility. However, cryopreservation itself induces cryoinjury due to oxidative and osmotic stress, leading to reduction in the survival rate and functionality of SSCs. Glial-derived neurotrophic factor family receptor alpha 1 (GFRα1) and promyelocytic leukemia zinc finger (PLZF) are expressed during the self-renewal and differentiation of SSCs, making them key tools for identifying the functionality of SSCs. To the best of our knowledge, the involvement of GFRα1 and PLZF in determining the functionality of SSCs after cryopreservation with therapeutic intervention is limited. Therefore, the purpose of this review is to determine the role of GFRα1 and PLZF as biomarkers for evaluating the functionality of SSCs in cryopreservation with therapeutic intervention. Therapeutic intervention, such as the use of antioxidants, and enhancement in cryopreservation protocols, such as cell encapsulation, cryoprotectant agents (CPA), and equilibrium of time and temperature increase the expression of GFRα1 and PLZF, resulting in maintaining the functionality of SSCs. In conclusion, GFRα1 and PLZF have the potential as biomarkers in cryopreservation with therapeutic intervention of SSCs to ensure the functionality of the stem cells.
Assuntos
Criopreservação , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial , Proteína com Dedos de Zinco da Leucemia Promielocítica , Espermatogônias , Células-Tronco , Humanos , Masculino , Biomarcadores/metabolismo , Fator Neurotrófico Derivado de Linhagem de Célula Glial/genética , Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/genética , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Proteína com Dedos de Zinco da Leucemia Promielocítica/genética , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Espermatogônias/metabolismo , Células-Tronco/metabolismo , Testículo/metabolismo , Dedos de ZincoRESUMO
Group 3 innate lymphoid cells (ILC3s) play important roles in maintaining intestinal homeostasis by protecting the host from pathogen infections and tissue inflammation. The transcription factor PLZF (promyelocytic leukemia zinc finger), encoded by zinc finger BTB domain containing 16 (Zbtb16), is highly and transiently expressed in ILC precursors (ILCPs). However, the role of PLZF in regulating ILC3 development and function remains unknown. Here, we show that PLZF was specifically expressed in mature intestinal ILC3s compared with other ILC subsets. PLZF was dispensable for ILC3 development. However, PLZF deficiency in ILC3s resulted in increased innate interleukin-22 (IL-22) secretion and protection against gut infection and inflammation. Mechanistically, PLZF negatively regulated IL-22 expression by ILC3s in a cell-intrinsic manner by binding to the IL-22 promoter region for transcriptional repression. Together, our data suggest that PLZF restricts intestinal ILC3 function to regulate gut immune homeostasis.
Assuntos
Imunidade Inata , Linfócitos , Proteína com Dedos de Zinco da Leucemia Promielocítica , Humanos , Expressão Gênica , Inflamação/metabolismo , Fatores de Transcrição/metabolismo , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismoRESUMO
Clarifying the pathogenesis of asthma and/or identifying the specific pathway underlying oral asthma tolerance (OT) would be of great significance. In our previous study, promyelocytic leukemia zinc finger (PLZF), which reportedly regulates memory phenotypes, was found to promote ovalbumin (OVA)-induced OT. Therefore, this study aimed to explore the regulatory effects of PLZF on memory phenotypes in asthma and OT mouse models. We found that Zbtb16 (encoding PLZF) and PLZF+ cells were highly increased in OT lungs compared with asthmatic lungs. PLZF was co-expressed with GATA3, and IL-4+PLZF+ cells were significantly lower in OT lungs than in asthmatic lungs. Notably, memory cells were decreased in OT mice, and these mice had PLZF+ cells that expressed lower levels of CD44 than those of asthmatic mice. When Zbtb16 was overexpressed in splenic lymphocytes, the number of CD44+ cells decreased. There were increased memory cells in splenic lymphocytes after treatment with the supernatant of OVA-treated airway epithelial cells; however, this was reversed by Zbtb16 overexpression. Early respiratory syncytial virus infection increased memory cells and reduced PLZF+ cells in the OT mice. Collectively, these results indicate that PLZF may reduce the proportion of memory cells, thereby, promoting the establishment of OT.
Assuntos
Asma , Proteína com Dedos de Zinco da Leucemia Promielocítica , Animais , Camundongos , Asma/imunologia , Asma/metabolismo , Receptores de Hialuronatos , Pulmão , Linfócitos , Ovalbumina , Fenótipo , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Infecções por Vírus Respiratório Sincicial/imunologia , Infecções por Vírus Respiratório Sincicial/metabolismoRESUMO
Long non-coding RNA ELFN1 antisense RNA 1 (ELFN1-AS1) has been reported as a cancer driver in many human malignancies. This study was conducted to investigate the function of ELFN1-AS1 in gastric cancer (GC) and its mechanism of action. Bioinformatics analysis revealed increased expression of ELFN1-AS1 in GC, and abundant expression of ELFN1-AS1 was observed in the acquired GC cell lines. Knockdown of ELFN1-AS1 in GC cells weakened cell proliferation, invasion, migration, and resistance to apoptosis. ELFN1-AS1 was mainly localized in the nuclei of GC cells. ELFN1-AS1 recruited DNA methyltransferases to the promoter region of ZBTB16 and induced transcriptional repression of ZBTB16 through methylation modification. Furthermore, downregulation of ZBTB16 activated the phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) signaling pathway and restored the proliferation and invasiveness of GC cells. In vivo, downregulation of ELFN1-AS1 reduced the growth rate of xenograft tumors in mice. In summary, this study demonstrates that ELFN1-AS1 recruits DNA methyltransferases to the promoter region of ZBTB16 to induce its transcriptional repression, which further augments the development of GC by activating the PI3K/AKT signaling pathway.
Assuntos
MicroRNAs , RNA Longo não Codificante , Neoplasias Gástricas , Animais , Linhagem Celular Tumoral , Proliferação de Células/genética , DNA , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Humanos , Metiltransferases/genética , Camundongos , MicroRNAs/genética , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteína com Dedos de Zinco da Leucemia Promielocítica/genética , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Longo não Codificante/metabolismo , Neoplasias Gástricas/patologiaRESUMO
T helper 2 (Th2) cytokine production by invariant natural killer T (iNKT) cells is involved in the development of asthma, but the regulation of Th2 cytokines in iNKT cells remains unknown. Although it is known that progranulin (PGRN) induces the production of Th2 cytokines in iNKT cells in vivo, the underlying mechanism is not clear. This study aims to investigate the role of PGRN in iNKT cells. The effects of PGRN on the differentiation of iNKT cells was detected by flow cytometry. Then stimulation of iNKT cells and airway resistance were carried out to evaluate the function of PGRN on iNKT cells. Furthermore, the mechanisms of PGRN in regulating iNKT cells was investigated by RT-PCR, WB, confocal and luciferase reporter assays. The absolute number of iNKT cells decreased in PGRN KO mice despite an increase in the percentage of iNKT cells. Furthermore, analyzing the subsets of iNKT cells, we found that NKT2 cells and their IL-4 production were reduced. Mechanistically, the decrease in NKT2 cells in the PGRN KO mice was caused by increased expression of enhancer of zeste homolog 2 (EZH2), that in turn caused increased degradation and altered nuclear localization of PLZF. Interestingly, PGRN signaling decreased expression of EZH2 and treatment of the PGRN KO mice with the EZH2 specific inhibitor GSK343 rescued the defect in NKT2 differentiation, IL-4 generation, and PLZF expression. Altogether, We have revealed a new pathway (PGRN-EZH2-PLZF), which regulates the Th2 responses of iNKT cells and provides a potentially new target for asthma treatment.
Assuntos
Asma , Proteína Potenciadora do Homólogo 2 de Zeste , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Animais , Diferenciação Celular , Citocinas , Interleucina-4 , Camundongos , Camundongos Endogâmicos C57BL , ProgranulinasRESUMO
Objective: To explore the effects of Nd(2)O(3) exposure to rare earth particles on the secretion of sex hormones, cytochrome P450 family member 11A1 (CYP11A1) , spermatogenesis markers promyelocytic leukemia zinc finger protein (PLZF) and retinoic acid stimulating gene 8 (STRA8) protein in C57 BL/6J male mice. Methods: In March 2021, Forty-eight male C57 BL/6J mice aged 6-8 weeks divided into control group and Nd(2)O(3) exposure low, medium and high dose groups (exposing doses of 62.5, 125.0, 250.0 mg/ml Nd(2)O(3)) , 12 per group. The mice in the Nd(2)O(3) groups were perfused with different doses of Nd(2)O(3) suspension by a one-time non-exposing tracheal instillation method, and the control group was perfused with an equal volume of normal saline, with a volume of 0.1 ml, to establish a mouse reproductive function injury model. After 28 days of exposure, the mice's body weight, testes and epididymis were weighed, and the organ coefficients were calculated; the two epididymis were taken to make a sperm suspension to determine the sperm count, survival rate, and deformity rate; inductively coupled plasma mass spectrometry (ICP-MS) method was used to detect the content of Nd in mouse testis tissue; HE staining was used to detect testicular tissue pathological changes and quantitative analysis; enzyme-linked immunosorbent assay (ELISA) method was used to detect serum luteinizing hormone (LH) and follicle stimulating hormone (FSH) and testosterone (T) content; western blot was used to detect the protein levels of CYP11A1, PLZF and STRA8 in testicular tissues. Results: Compared with the control group, with the increase of the exposure dose, the Nd content in the testis of the mice showed an increasing trend, the sperm survival rate and LH showed a decreasing trend, and the sperm deformity rate showed an increasing trend (P<0.05) ; Pathological showed that the number of sperm in the seminiferous tubules of the testicular tissue in the Nd(2)O(3) medium and high dose groups was significantly reduced, and the germinal epithelial disintegration, intraepithelial vacuolization, and exfoliation of spermatogenic cells and supporting cells occurred; The height of germinal epithelium was significantly reduced, and the percentage of damaged seminiferous tubules showed an increasing trend (P<0.05) ; FSH and T levels in serum in the middle and high dose groups of Nd(2)O(3), and CYP11A1, PLZF and STRA8 proteins in testicular tissues showed a downward trend with increasing dose (P<0.05) . Conclusion: The rare earth particulate Nd(2)O(3) may interfere with the expression of CYP11A1, PLZF and STRA8 protein, thereby causing the disorder of sex hormone secretion in the body, the maintenance of spermatogonia and the obstruction of the process of meiosis, causing reproductive function damage.
Assuntos
Enzima de Clivagem da Cadeia Lateral do Colesterol , Neodímio , Sêmen , Animais , Masculino , Camundongos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Enzima de Clivagem da Cadeia Lateral do Colesterol/metabolismo , Hormônio Foliculoestimulante/metabolismo , Hormônio Luteinizante/metabolismo , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Sêmen/metabolismo , Contagem de Espermatozoides , Espermatogênese , Testículo , Testosterona/metabolismo , Neodímio/toxicidade , Óxidos/toxicidadeRESUMO
CONTEXT: Central serous chorioretinopathy (CSC) is a severe ocular disease characterized by fluid accumulation under the retina and abnormalities in the underlying vascular layer, the choroid. CSC has a striking prevalence in males of 80% to 90% of total patients. Corticosteroids are the most pronounced extrinsic risk factor for CSC. Choroidal endothelial cells (CECs) are important for the vascular integrity of the choroid, but the effects of corticosteroid effects in these cells are unknown. OBJECTIVE: We aimed to reveal the potential steroidal contribution to CSC. METHOD: We characterized the expression of the glucocorticoid, mineralocorticoid, and androgen receptor in the human choroid using immunohistochemistry. Using RNA-sequencing, we describe the cortisol response in human CECs derived from 5 male and 5 female postmortem donors. RESULTS: The glucocorticoid receptor was highly expressed in the human choroid, whereas no to minimal expression of the mineralocorticoid and androgen receptors was observed. The extensive transcriptional response to cortisol in human primary cultured CECs showed interindividual differences but very few sex differences. Several highly regulated genes such as ZBTB16 (log2 fold change males 7.9; females 6.2) provide strong links to choroidal vascular regulation. CONCLUSIONS: The glucocorticoid receptor predominantly mediates the response to cortisol in human CECs. Interindividual differences are an important determinant regarding the cortisol response in human cultured CECs, whereas intrinsic sex differences appear less pronounced. The marked response of particular target genes in endothelial cells to cortisol, such as ZBTB16, warrants further investigation into their potential role in the pathophysiology of CSC and other vascular conditions.
Assuntos
Coriorretinopatia Serosa Central/patologia , Corioide/patologia , Hidrocortisona/metabolismo , Receptores de Glucocorticoides/metabolismo , Idoso , Idoso de 80 Anos ou mais , Biópsia , Corioide/citologia , Células Endoteliais/metabolismo , Feminino , Células Endoteliais da Veia Umbilical Humana , Humanos , Masculino , Pessoa de Meia-Idade , Cultura Primária de Células , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , RNA-Seq , Fatores SexuaisRESUMO
Pomalidomide and lenalidomide are immunomodulatory agents that were derived from thalidomide. Cereblon (CRBN) is a common direct target of thalidomide and related compounds and works as a Cullin Ring 4 E3 ubiquitin ligase (CRL4) with DDB1, CUL4, and ROC1. The substrate specificity of CRL4CRBN is modulated by thalidomide-related compounds. While lenalidomide is approved for the treatment of several diseases including multiple myeloma, 5q- syndrome, mantle cell lymphoma, and follicular lymphoma, pomalidomide is approved only for the treatment of lenalidomide-resistant multiple myeloma. Here we show that PLZF/ZBTB16 and its fusion proteins are pomalidomide-dependent neosubstrates of CRL4CRBN. PLZF joins to RARα or potentially other partner genes, and the translocation causes leukemias, such as acute promyelocytic leukemia and T-cell acute lymphoblastic leukemia. We demonstrate that pomalidomide treatment induces PLZF-RARα degradation, resulting in antiproliferation of leukemic cells expressing PLZF-RARα. This study highlights a potential therapeutic role of pomalidomide as a degrader of leukemogenic fusion proteins.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteína com Dedos de Zinco da Leucemia Promielocítica/genética , Talidomida/análogos & derivados , Ubiquitina-Proteína Ligases/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Sequência de Aminoácidos , Células HEK293 , Humanos , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Alinhamento de Sequência , Talidomida/metabolismo , Ubiquitina-Proteína Ligases/metabolismoRESUMO
OBJECTIVE: The purpose of this study is to identify novel biomarkers for the prognosis of Ewing's sarcoma based on bioinformatics analysis. METHODS: The GSE63157 and GSE17679 datasets contain patient and healthy control microarray data that were downloaded from the Gene Expression Omnibus (GEO) database and analyzed through R language software to obtain differentially expressed genes (DEGs). Firstly, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional enrichment, protein-protein interaction (PPI) networks, and Cytoscape Molecular Complex Detection (MCODE) plug-in were then used to compute the highest scores of the module. After survival analysis, the hub genes were lastly obtained from the two module genes. RESULTS: A total of 1181 DEGs were identified from the two GSEs. Through MCODE and survival analysis, we obtain 53 DEGs from the module and 29 overall survival- (OS-) related genes. ZBTB16 was the only downregulated gene after Venn diagrams. Survival analysis indicates that there was a significant correlation between the high expression of ZBTB16 and the OS of Ewing's sarcoma (ES), and the low expression group had an unfavorable OS when compared to the high expression group. CONCLUSIONS: High expression of ZBTB16 may serve as a predictor biomarker of poor prognosis in ES patients.
Assuntos
Biomarcadores Tumorais/metabolismo , Biologia Computacional , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Sarcoma de Ewing/metabolismo , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Ontologia Genética , Redes Reguladoras de Genes/efeitos dos fármacos , Humanos , Estimativa de Kaplan-Meier , Prognóstico , Mapas de Interação de Proteínas/genética , Sarcoma de Ewing/genética , Sarcoma de Ewing/patologiaRESUMO
Spermatogonial stem cells (SSCs) are essential for male fertility. Here, we report that mouse SSC generation is driven by a transcription factor (TF) cascade controlled by the homeobox protein, RHOX10, which acts by driving the differentiation of SSC precursors called pro-spermatogonia (ProSG). We identify genes regulated by RHOX10 in ProSG in vivo and define direct RHOX10-target genes using several approaches, including a rapid temporal induction assay: iSLAMseq. Together, these approaches identify temporal waves of RHOX10 direct targets, as well as RHOX10 secondary-target genes. Many of the RHOX10-regulated genes encode proteins with known roles in SSCs. Using an in vitro ProSG differentiation assay, we find that RHOX10 promotes mouse ProSG differentiation through a conserved transcriptional cascade involving the key germ-cell TFs DMRT1 and ZBTB16. Our study gives important insights into germ cell development and provides a blueprint for how to define TF cascades.
Assuntos
Proteínas de Homeodomínio/metabolismo , Transdução de Sinais , Espermatogônias/citologia , Células-Tronco/citologia , Células-Tronco/metabolismo , Fatores de Transcrição/metabolismo , Animais , Sequência de Bases , Diferenciação Celular/genética , Regulação da Expressão Gênica no Desenvolvimento , Genoma , Células Germinativas/metabolismo , Células HEK293 , Humanos , Masculino , Camundongos Endogâmicos C57BL , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Transcrição Gênica , Ativação Transcricional/genéticaRESUMO
RORγt+ lymphocytes, including interleukin 17 (IL-17)-producing gamma delta T (γδT17) cells, T helper 17 (Th17) cells, and group 3 innate lymphoid cells (ILC3s), are important immune regulators. Compared to Th17 cells and ILC3s, γδT17 cell metabolism and its role in tissue homeostasis remains poorly understood. Here, we report that the tissue milieu shapes splenic and intestinal γδT17 cell gene signatures. Conditional deletion of mitochondrial transcription factor A (Tfam) in RORγt+ lymphocytes significantly affects systemic γδT17 cell maintenance and reduces ILC3s without affecting Th17 cells in the gut. In vivo deletion of Tfam in RORγt+ lymphocytes, especially in γδT17 cells, results in small intestine tissue remodeling and increases small intestine length by enhancing the type 2 immune responses in mice. Moreover, these mice show dysregulation of the small intestine transcriptome and metabolism with less body weight but enhanced anti-helminth immunity. IL-22, a cytokine produced by RORγt+ lymphocytes inhibits IL-13-induced tuft cell differentiation in vitro, and suppresses the tuft cell-type 2 immune circuit and small intestine lengthening in vivo, highlighting its key role in gut tissue remodeling.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Proteínas de Grupo de Alta Mobilidade/metabolismo , Intestino Delgado/imunologia , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Animais , Diferenciação Celular , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Feminino , Perfilação da Expressão Gênica , Proteínas de Grupo de Alta Mobilidade/deficiência , Proteínas de Grupo de Alta Mobilidade/genética , Homeostase/imunologia , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Intestino Delgado/citologia , Intestino Delgado/metabolismo , Masculino , Metaboloma , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Organoides , Proteína com Dedos de Zinco da Leucemia Promielocítica/genética , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Subpopulações de Linfócitos T/citologia , Células Th17/citologia , Células Th17/imunologia , Células Th17/metabolismoRESUMO
BACKGROUND: The importance of spermatogonial stem cells (SSCs) in spermatogenesis is crucial and intrinsic factors and extrinsic signals mediate fate decisions of SSCs. Among endogenous regulators, microRNAs (miRNAs) play critical role in spermatogenesis. However, the mechanisms which individual miRNAs regulate self- renewal and differentiation of SSCs are unknown. The aim of this study was to investigate effects of miRNA-30a-5p inhibitor on fate determinations of SSCs. METHODS: SSCs were isolated from testes of neonate mice (3-6 days old) and their purities were performed by flow cytometry with ID4 and Thy1 markers. Cultured cells were transfected with miRNA- 30a-5p inhibitor. Evaluation of the proliferation (GFRA1, PLZF and ID4) and differentiation (C-Kit & STRA8) markers of SSCs were accomplished by immunocytochemistry and western blot 48 h after transfection. RESULTS: Based on the results of flow cytometry with ID4 and Thy1 markers, percentage of purity of SSCs was about 84.3 and 97.4 % respectively. It was found that expression of differentiation markers after transfection was significantly higher in miRNA-30a- 5p inhibitor group compared to other groups. The results of proliferation markers evaluation also showed decrease of GFRA1, PLZF and ID4 protein in SSCs transfected with miRNA-30a-5p inhibitor compared to the other groups. CONCLUSIONS: It can be concluded that inhibition of miRNA-30a-5p by overexpression of differentiation markers promotes differentiation of Spermatogonial Stem Cells.
Assuntos
Células-Tronco Germinativas Adultas/fisiologia , MicroRNAs/fisiologia , Espermatogênese/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Células-Tronco Germinativas Adultas/metabolismo , Animais , Animais Recém-Nascidos , Western Blotting , Autorrenovação Celular , Citometria de Fluxo , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Imuno-Histoquímica , Proteínas Inibidoras de Diferenciação/metabolismo , Masculino , Camundongos , MicroRNAs/antagonistas & inibidores , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Proteínas Proto-Oncogênicas c-kit/metabolismo , Antígenos Thy-1/metabolismoRESUMO
Conventional T cells are selected by peptide-MHC expressed by cortical epithelial cells in the thymus, and not by cortical thymocytes themselves that do not express MHC I or MHC II. Instead, cortical thymocytes express non-peptide presenting MHC molecules like CD1d and MR1, and promote the selection of PLZF+ iNKT and MAIT cells, respectively. Here, we report an inducible class-I transactivator mouse that enables the expression of peptide presenting MHC I molecules in different cell types. We show that MHC I expression in DP thymocytes leads to expansion of peptide specific PLZF+ innate-like (PIL) T cells. Akin to iNKT cells, PIL T cells differentiate into three functional effector subsets in the thymus, and are dependent on SAP signaling. We demonstrate that PIL and NKT cells compete for a narrow niche, suggesting that the absence of peptide-MHC on DP thymocytes facilitates selection of non-peptide specific lymphocytes.
Assuntos
Antígenos de Histocompatibilidade Classe I/metabolismo , Imunidade Inata , Timócitos/imunologia , Timo/imunologia , Animais , Diferenciação Celular/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Camundongos , Camundongos Transgênicos , Células T Invariantes Associadas à Mucosa/imunologia , Células T Matadoras Naturais/imunologia , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Timócitos/metabolismo , Timo/citologiaRESUMO
Optineurin (OPTN) is a multifunctional protein that mediates a network of cellular processes regulating membrane trafficking, inflammatory responses and autophagy. The OPTN-rich interactome includes Group I metabotropic glutamate receptors (mGluR1 and 5), members of the Gαq/11 protein receptor family. Recent evidence has shown that mGluR5, in addition to its canonical Gαq/11 protein-coupled signaling, regulates autophagic machinery via mTOR/ULK1 and GSK3ß/ZBTB16 pathways in both Alzheimer's and Huntington's disease mouse models. Despite its potential involvement, the role of OPTN in mediating mGluR5 downstream signaling cascades remains largely unknown. Here, we employed a CRISPR/Cas9 OPTN-deficient STHdhQ7/Q7 striatal cell line and global OPTN knockout mice to investigate whether Optn gene deletion alters both mGluR5 canonical and noncanonical signaling. We find that OPTN is required for mGluR5-activated Ca2+ flux and ERK1/2 signaling following receptor activation in STHdhQ7/Q7 cells and acute hippocampal slices. Deletion of OPTN impairs both GSK3ß/ZBTB16 and mTOR/ULK1 autophagic signaling in STHdhQ7/Q7 cells. Furthermore, mGluR5-dependent regulation of GSK3ß/ZBTB16 and mTOR/ULK1 autophagic signaling is impaired in hippocampal slices of OPTN knockout mice. Overall, we show that the crosstalk between OPTN and mGluR5 can have major implication on receptor signaling and therefore potentially contribute to the pathophysiology of neurodegenerative diseases.