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1.
Genes Dev ; 33(23-24): 1641-1656, 2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31727773

RESUMO

Angiopoietin-like protein 2 (ANGPTL2) is a secreted glycoprotein homologous to angiopoietins. Previous studies suggest that tumor cell-derived ANGPTL2 has tumor-promoting function. Here, we conducted mechanistic analysis comparing ANGPTL2 function in cancer progression in a murine syngeneic model of melanoma and a mouse model of translocation renal cell carcinoma (tRCC). ANGPTL2 deficiency in tumor cells slowed tRCC progression, supporting a tumor-promoting role. However, systemic ablation of ANGPTL2 accelerated tRCC progression, supporting a tumor-suppressing role. The syngeneic model also demonstrated a tumor-suppressing role of ANGPTL2 in host tumor microenvironmental cells. Furthermore, the syngeneic model showed that PDGFRα+ fibroblasts in the tumor microenvironment express abundant ANGPTL2 and contribute to tumor suppression. Moreover, host ANGPTL2 facilitates CD8+ T-cell cross-priming and enhances anti-tumor immune responses. Importantly, ANGPTL2 activates dendritic cells through PIR-B-NOTCH signaling and enhances tumor vaccine efficacy. Our study provides strong evidence that ANGPTL2 can function in either tumor promotion or suppression, depending on what cell type it is expressed in.


Assuntos
Proteínas Semelhantes a Angiopoietina/genética , Proteínas Semelhantes a Angiopoietina/metabolismo , Linfócitos T CD8-Positivos/imunologia , Carcinoma de Células Renais/fisiopatologia , Progressão da Doença , Melanoma/fisiopatologia , Transdução de Sinais , Proteína 2 Semelhante a Angiopoietina , Proteínas Semelhantes a Angiopoietina/deficiência , Proteínas Semelhantes a Angiopoietina/imunologia , Animais , Vacinas Anticâncer/imunologia , Carcinoma de Células Renais/imunologia , Células Dendríticas/imunologia , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica , Melanoma/imunologia , Camundongos , Transdução de Sinais/genética , Células Estromais/imunologia , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia
2.
Genes Dev ; 30(9): 1034-46, 2016 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-27151976

RESUMO

The tumor suppressor folliculin (FLCN) forms a repressor complex with AMP-activated protein kinase (AMPK). Given that AMPK is a master regulator of cellular energy homeostasis, we generated an adipose-specific Flcn (Adipoq-FLCN) knockout mouse model to investigate the role of FLCN in energy metabolism. We show that loss of FLCN results in a complete metabolic reprogramming of adipose tissues, resulting in enhanced oxidative metabolism. Adipoq-FLCN knockout mice exhibit increased energy expenditure and are protected from high-fat diet (HFD)-induced obesity. Importantly, FLCN ablation leads to chronic hyperactivation of AMPK, which in turns induces and activates two key transcriptional regulators of cellular metabolism, proliferator-activated receptor γ (PPARγ) coactivator-1α (PGC-1α) and estrogen-related receptor α (ERRα). Together, the AMPK/PGC-1α/ERRα molecular axis positively modulates the expression of metabolic genes to promote mitochondrial biogenesis and activity. In addition, mitochondrial uncoupling proteins as well as other markers of brown fat are up-regulated in both white and brown FLCN-null adipose tissues, underlying the increased resistance of Adipoq-FLCN knockout mice to cold exposure. These findings identify a key role of FLCN as a negative regulator of mitochondrial function and identify a novel molecular pathway involved in the browning of white adipocytes and the activity of brown fat.


Assuntos
Tecido Adiposo Bege/metabolismo , Metabolismo Energético/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Proteínas Proto-Oncogênicas/genética , Receptores de Estrogênio/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Quinases Ativadas por AMP/genética , Animais , Temperatura Baixa , Ativação Enzimática/genética , Regulação da Expressão Gênica/genética , Camundongos , Camundongos Knockout , Obesidade/enzimologia , Obesidade/genética , Oxirredução , Proteínas Proto-Oncogênicas/metabolismo , Receptores de Estrogênio/genética , Transdução de Sinais , Proteínas Supressoras de Tumor/metabolismo , Receptor ERRalfa Relacionado ao Estrogênio
3.
Cancer Sci ; 113(7): 2352-2367, 2022 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-35396773

RESUMO

Renal cell carcinoma with Xp11.2 translocation involving the TFE3 gene (TFE3-RCC) is a recently identified subset of RCC with unique morphology and clinical presentation. The chimeric PRCC-TFE3 protein produced by Xp11.2 translocation has been shown to transcriptionally activate its downstream target genes that play important roles in carcinogenesis and tumor development of TFE3-RCC. However, the underlying molecular mechanisms remain poorly understood. Here we show that in TFE3-RCC cells, PRCC-TFE3 controls heme oxygenase 1 (HMOX1) expression to confer chemoresistance. Inhibition of HMOX1 sensitized the PRCC-TFE3 expressing cells to genotoxic reagents. We screened for a novel chlorambucil-polyamide conjugate (Chb) to target PRCC-TFE3-dependent transcription, and identified Chb16 as a PRCC-TFE3-dependent transcriptional inhibitor of HMOX1 expression. Treatment of the patient-derived cancer cells with Chb16 exhibited senescence and growth arrest, and increased sensitivity of the TFE3-RCC cells to the genotoxic reagent etoposide. Thus, our data showed that the TFE3-RCC cells acquired chemoresistance through HMOX1 expression and that inhibition of HMOX1 by Chb16 may be an effective therapeutic strategy for TFE3-RCC.


Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/metabolismo , Clorambucila/farmacologia , Cromossomos Humanos X , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , Nylons , Translocação Genética
4.
J Immunol ; 203(11): 2899-2908, 2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31676673

RESUMO

Folliculin interacting protein 1 (Fnip1) is a cytoplasmic protein originally discovered through its interaction with the master metabolic sensor 5' AMP-activated protein kinase (AMPK) and Folliculin, a protein mutated in individuals with Birt-Hogg-Dubé Syndrome. In response to low energy, AMPK stimulates catabolic pathways such as autophagy to enhance energy production while inhibiting anabolic pathways regulated by the mechanistic target of rapamycin complex 1 (mTORC1). We previously found that constitutive disruption of Fnip1 in mice resulted in a lack of peripheral B cells because of a block in B cell development at the pre-B cell stage. Both AMPK and mTORC1 were activated in Fnip1-deficient B cell progenitors. In this study, we found inappropriate mTOR localization at the lysosome under nutrient-depleted conditions. Ex vivo lysine or arginine depletion resulted in increased apoptosis. Genetic inhibition of AMPK, inhibition of mTORC1, or restoration of cell viability with a Bcl-xL transgene failed to rescue B cell development in Fnip1-deficient mice. Fnip1-deficient B cell progenitors exhibited increased nuclear localization of transcription factor binding to IgHM enhancer 3 (TFE3) in developing B cells, which correlated with an increased expression of TFE3-target genes, increased lysosome numbers and function, and increased autophagic flux. These results indicate that Fnip1 modulates autophagy and energy response pathways in part through the regulation of AMPK, mTORC1, and TFE3 in B cell progenitors.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Linfócitos B/metabolismo , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Proteínas de Transporte/metabolismo , Homeostase , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Animais , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout
5.
Hum Mol Genet ; 27(15): 2712-2724, 2018 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-29767721

RESUMO

Birt-Hogg-Dubé (BHD) syndrome is a hereditary kidney cancer syndrome, which predisposes patients to develop kidney cancer, cutaneous fibrofolliculomas and pulmonary cysts. The responsible gene FLCN is a tumor suppressor for kidney cancer, which plays an important role in energy homeostasis through the regulation of mitochondrial oxidative metabolism. However, the process by which FLCN-deficiency leads to renal tumorigenesis is unclear. In order to clarify molecular pathogenesis of BHD-associated kidney cancer, we conducted whole-exome sequencing analysis using next-generation sequencing technology as well as metabolite analysis using liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry. Whole-exome sequencing analysis of BHD-associated kidney cancer revealed that copy number variations of BHD-associated kidney cancer are considerably different from those already reported in sporadic cases. In somatic variant analysis, very few variants were commonly observed in BHD-associated kidney cancer; however, variants in chromatin remodeling genes were frequently observed in BHD-associated kidney cancer (17/29 tumors, 59%). Metabolite analysis of BHD-associated kidney cancer revealed metabolic reprogramming toward upregulated redox regulation which may neutralize reactive oxygen species potentially produced from mitochondria with increased respiratory capacity under FLCN-deficiency. BHD-associated kidney cancer displays unique molecular characteristics that are completely different from sporadic kidney cancer, providing mechanistic insight into tumorigenesis under FLCN-deficiency as well as a foundation for development of novel therapeutics for kidney cancer.


Assuntos
Síndrome de Birt-Hogg-Dubé/patologia , Montagem e Desmontagem da Cromatina/genética , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor/genética , Síndrome de Birt-Hogg-Dubé/genética , Variações do Número de Cópias de DNA , Mutação em Linhagem Germinativa , Humanos , Neoplasias Renais/patologia , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Sequenciamento do Exoma
6.
Biochem Biophys Res Commun ; 522(4): 931-938, 2020 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-31806376

RESUMO

FLCN is a tumor suppressor gene which controls energy homeostasis through regulation of a variety of metabolic pathways including mitochondrial oxidative metabolism and autophagy. Birt-Hogg-Dubé (BHD) syndrome which is driven by germline alteration of the FLCN gene, predisposes patients to develop kidney cancer, cutaneous fibrofolliculomas, pulmonary cysts and less frequently, salivary gland tumors. Here, we report metabolic roles for FLCN in the salivary gland as well as their clinical relevance. Screening of salivary glands of BHD patients using ultrasonography demonstrated increased cyst formation in the salivary gland. Salivary gland tumors that developed in BHD patients exhibited an upregulated mTOR-S6R pathway as well as increased GPNMB expression, which are characteristics of FLCN-deficient cells. Salivary gland-targeted Flcn knockout mice developed cytoplasmic clear cell formation in ductal cells with increased mitochondrial biogenesis, upregulated mTOR-S6K pathway, upregulated TFE3-GPNMB axis and upregulated lipid metabolism. Proteomic and metabolite analysis using LC/MS and GC/MS revealed that Flcn inactivation in salivary gland triggers metabolic reprogramming towards the pentose phosphate pathway which consequently upregulates nucleotide synthesis and redox regulation, further supporting that Flcn controls metabolic homeostasis in salivary gland. These data uncover important roles for FLCN in salivary gland; metabolic reprogramming under FLCN deficiency might increase nucleotide production which may feed FLCN-deficient salivary gland cells to trigger tumor initiation and progression, providing mechanistic insight into salivary gland tumorigenesis as well as a foundation for development of novel therapeutics for salivary gland tumors.


Assuntos
Cistos/metabolismo , Cistos/patologia , Nucleotídeos/biossíntese , Proteínas Proto-Oncogênicas/metabolismo , Glândulas Salivares/metabolismo , Glândulas Salivares/patologia , Proteínas Supressoras de Tumor/metabolismo , Adulto , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Cistos/diagnóstico por imagem , Feminino , Ontologia Genética , Glicólise , Humanos , Masculino , Camundongos Knockout , Pessoa de Meia-Idade , Biogênese de Organelas , Via de Pentose Fosfato , Proteínas Proto-Oncogênicas/deficiência , Glândulas Salivares/diagnóstico por imagem , Serina-Treonina Quinases TOR/metabolismo , Proteínas Supressoras de Tumor/deficiência , Regulação para Cima
7.
Cancer Sci ; 110(6): 1897-1908, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31006167

RESUMO

Xp11.2 translocation renal cell carcinoma (Xp11 tRCC) is a rare sporadic pediatric kidney cancer caused by constitutively active TFE3 fusion proteins. Tumors in patients with Xp11 tRCC tend to recur and undergo frequent metastasis, in part due to lack of methods available to detect early-stage disease. Here we generated transgenic (Tg) mice overexpressing the human PRCC-TFE3 fusion gene in renal tubular epithelial cells, as an Xp11 tRCC mouse model. At 20 weeks of age, mice showed no histological abnormalities in kidney but by 40 weeks showed Xp11 tRCC development and related morphological and histological changes. MicroRNA (miR)-204-5p levels in urinary exosomes of 40-week-old Tg mice showing tRCC were significantly elevated compared with levels in control mice. MicroRNA-204-5p expression also significantly increased in primary renal cell carcinoma cell lines established both from Tg mouse tumors and from tumor tissue from 2 Xp11 tRCC patients. All of these lines secreted miR-204-5p-containing exosomes. Notably, we also observed increased miR-204-5p levels in urinary exosomes in 20-week-old renal PRCC-TFE3 Tg mice prior to tRCC development, and those levels were equivalent to those in 40-week-old Tg mice, suggesting that miR-204-5p increases follow expression of constitutively active TFE3 fusion proteins in renal tubular epithelial cells prior to overt tRCC development. Finally, we confirmed that miR-204-5p expression significantly increases in noncancerous human kidney cells after overexpression of a PRCC-TFE3 fusion gene. These findings suggest that miR-204-5p in urinary exosomes could be a useful biomarker for early diagnosis of patients with Xp11 tRCC.


Assuntos
Biomarcadores Tumorais/genética , Carcinoma de Células Renais/genética , Cromossomos Humanos X/genética , Neoplasias Renais/genética , MicroRNAs/genética , Translocação Genética , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Biomarcadores Tumorais/urina , Carcinoma de Células Renais/metabolismo , Carcinoma de Células Renais/urina , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Exossomos/genética , Humanos , Rim/anormalidades , Rim/metabolismo , Neoplasias Renais/metabolismo , Neoplasias Renais/urina , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , MicroRNAs/urina , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo
8.
Hum Mol Genet ; 26(2): 354-366, 2017 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-28007907

RESUMO

Germline H255Y and K508R missense mutations in the folliculin (FLCN) gene have been identified in patients with bilateral multifocal (BMF) kidney tumours and clinical manifestations of Birt-Hogg-Dubé (BHD) syndrome, or with BMF kidney tumours as the only manifestation; however, their impact on FLCN function remains to be determined. In order to determine if FLCN H255Y and K508R missense mutations promote aberrant kidney cell proliferation leading to pathogenicity, we generated mouse models expressing these mutants using BAC recombineering technology and investigated their ability to rescue the multi-cystic phenotype of Flcn-deficient mouse kidneys. Flcn H255Y mutant transgene expression in kidney-targeted Flcn knockout mice did not rescue the multi-cystic kidney phenotype. However, expression of the Flcn K508R mutant transgene partially, but not completely, abrogated the phenotype. Notably, expression of the Flcn K508R mutant transgene in heterozygous Flcn knockout mice resulted in development of multi-cystic kidneys and cardiac hypertrophy in some mice. These results demonstrate that both FLCN H255Y and K508R missense mutations promote aberrant kidney cell proliferation, but to different degrees. Based on the phenotypes of our preclinical models, the FLCN H255Y mutant protein has lost it tumour suppressive function leading to the clinical manifestations of BHD, whereas the FLCN K508R mutant protein may have a dominant negative effect on the function of wild-type FLCN in regulating kidney cell proliferation and, therefore, act as an oncoprotein. These findings may provide mechanistic insight into the role of FLCN in regulating kidney cell proliferation and facilitate the development of novel therapeutics for FLCN-deficient kidney cancer.


Assuntos
Síndrome de Birt-Hogg-Dubé/genética , Doenças Renais Císticas/genética , Neoplasias Renais/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor/genética , Animais , Síndrome de Birt-Hogg-Dubé/patologia , Cardiomegalia/genética , Cardiomegalia/patologia , Proliferação de Células/genética , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica , Mutação em Linhagem Germinativa , Humanos , Rim/patologia , Doenças Renais Císticas/patologia , Neoplasias Renais/patologia , Camundongos , Camundongos Knockout , Mutação de Sentido Incorreto
9.
Histopathology ; 75(2): 254-265, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30908700

RESUMO

AIMS: Xp11 rearrangement in renal cell carcinoma (RCC) typically involves gene fusion to the gene encoding transcription factor E3 (TFE3), a member of the microphthalmia-associated transcription factor family on chromosome Xp11.2. Dual-colour break-apart fluorescence in-situ hybridisation (FISH) is recommended to confirm histological diagnoses. Recently, RNA-binding motif protein 10 (RBM10), encoded by a gene on chromosome Xp11.3, was identified as a chimeric partner of TFE3; thus, RBM10-TFE3 fusion results from paracentric inversion. RBM10-TFE3 RCC may yield a false-negative result in FISH analysis of TFE3 expression. The aim of the present study was to investigate the clinicopathological features of RBM10-TFE3 RCC. METHODS AND RESULTS: Ten patients with RBM10-TFE3 RCC aged 31-71 years were investigated. Histological analysis, immunostaining, dual-colour break-apart FISH for TFE3, reverse transcription polymerase chain reaction and sequencing analysis were performed. No patient had a history of exposure to chemotherapy. Two of these patients died of RCC, and three were alive but developed metastases. Microscopically, the tumours were composed of a mixed architecture of tubulocystic and papillary patterns with scattered psammoma bodies. The tumours showed strong nuclear immunoreactivity for TFE3. FISH showed consistent closely spaced split signals in the RCCs of four patients, and polysomic signals with occasional closely spaced split signals in the RCCs of six patients. Of the latter six patients, five had renal failure, and four developed tumours in kidneys subjected to haemodialysis. CONCLUSIONS: The present study suggests that the carcinogenesis of RBM10-TFE3 RCC in some, but not all, patients may be associated with chronic kidney disease. The aggressive nature of RBM10-TFE3 RCC should be considered, as five patients experienced metastases.


Assuntos
Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Carcinoma de Células Renais/genética , Neoplasias Renais/genética , Proteínas de Ligação a RNA/genética , Adulto , Idoso , Carcinoma de Células Renais/complicações , Carcinoma de Células Renais/patologia , Inversão Cromossômica , Cromossomos Humanos X , Feminino , Humanos , Hibridização in Situ Fluorescente , Neoplasias Renais/complicações , Neoplasias Renais/patologia , Masculino , Pessoa de Meia-Idade , Fusão Oncogênica , Insuficiência Renal Crônica/complicações , Insuficiência Renal Crônica/genética , Translocação Genética
10.
Proc Natl Acad Sci U S A ; 112(13): E1624-31, 2015 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-25775561

RESUMO

Folliculin (FLCN)-interacting proteins 1 and 2 (FNIP1, FNIP2) are homologous binding partners of FLCN, a tumor suppressor for kidney cancer. Recent studies have revealed potential functions for Flcn in kidney; however, kidney-specific functions for Fnip1 and Fnip2 are unknown. Here we demonstrate that Fnip1 and Fnip2 play critical roles in kidney tumor suppression in cooperation with Flcn. We observed no detectable phenotype in Fnip2 knockout mice, whereas Fnip1 deficiency produced phenotypes similar to those seen in Flcn-deficient mice in multiple organs, but not in kidneys. We found that absolute Fnip2 mRNA copy number was low relative to Fnip1 in organs that showed phenotypes under Fnip1 deficiency but was comparable to Fnip1 mRNA copy number in mouse kidney. Strikingly, kidney-targeted Fnip1/Fnip2 double inactivation produced enlarged polycystic kidneys, as was previously reported in Flcn-deficient kidneys. Kidney-specific Flcn inactivation did not further augment kidney size or cystic histology of Fnip1/Fnip2 double-deficient kidneys, suggesting pathways dysregulated in Flcn-deficient kidneys and Fnip1/Fnip2 double-deficient kidneys are convergent. Heterozygous Fnip1/homozygous Fnip2 double-knockout mice developed kidney cancer at 24 mo of age, analogous to the heterozygous Flcn knockout mouse model, further supporting the concept that Fnip1 and Fnip2 are essential for the tumor-suppressive function of Flcn and that kidney tumorigenesis in human Birt-Hogg-Dubé syndrome may be triggered by loss of interactions among Flcn, Fnip1, and Fnip2. Our findings uncover important roles for Fnip1 and Fnip2 in kidney tumor suppression and may provide molecular targets for the development of novel therapeutics for kidney cancer.


Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Proteínas de Transporte/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias Renais/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Alelos , Animais , Proteínas Reguladoras de Apoptose/genética , Síndrome de Birt-Hogg-Dubé/genética , Proteínas de Transporte/genética , Modelos Animais de Doenças , Feminino , Rim/patologia , Neoplasias Renais/genética , Neoplasias Renais/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , Doenças Renais Policísticas/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor/genética
11.
Lab Invest ; 97(3): 343-351, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-27991910

RESUMO

Hereditary renal cell carcinomas (RCCs) are life-threatening disorders not only for the patients but also for their relatives. Birt-Hogg-Dubé syndrome (BHD) is an autosomal dominant disorder caused by germline mutations in the folliculin gene (FLCN). The protein product, FLCN, functions as a tumor suppressor, and the affected patients have high risks of developing multiple RCCs. The carcinogenic mechanisms stemming from FLCN dysfunction have been investigated using rodent models and human RCC tissues. However, very limited information has been available about in vitro signaling of human renal cells with genetically mutant FLCN. Herein, we established a new cell line, BHD-F59RSVT, from a BHD patient's chromophobe RCC by transfecting SV40 large T antigen. We investigated FLCN mutations, chromosome profiles, and cytopathologic characteristics of the cell line. BHD-F59RSVT reflected the patient's FLCN germline mutation, a 3-nt deletion in exon 13 (c.1528_1530delGAG). Neither somatic mutation nor loss of heterozygosity of FLCN was detectable. Chromosome 17p11.2 of the FLCN proximal region demonstrated a trimodal pattern. Genome-wide chromosomal analysis revealed a loss of chromosome 16 and mosaic segmental gains in chromosome 7. BHD-F59RSVT cells were positive when immunostained for cytokeratin 7, supporting their origin from distal convoluted tubules. Western blotting analysis demonstrated severely suppressed FLCN expression at the protein level. The collective findings indicate that the established cell line will be suitable for functional analysis of the typical phenotype of BHD-associated RCC with suppressed FLCN expression.


Assuntos
Síndrome de Birt-Hogg-Dubé/genética , Carcinoma de Células Renais/genética , Mutação em Linhagem Germinativa , Neoplasias Renais/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor/genética , Síndrome de Birt-Hogg-Dubé/complicações , Carcinoma de Células Renais/complicações , Carcinoma de Células Renais/patologia , Linhagem Celular Transformada , Linhagem Celular Tumoral , Variações do Número de Cópias de DNA , Análise Mutacional de DNA/métodos , Saúde da Família , Feminino , Humanos , Neoplasias Renais/complicações , Neoplasias Renais/patologia , Perda de Heterozigosidade , Masculino , Pessoa de Meia-Idade , Linhagem , Proteínas Proto-Oncogênicas/metabolismo , Cariotipagem Espectral/métodos , Células Tumorais Cultivadas , Proteínas Supressoras de Tumor/metabolismo
12.
Stem Cells ; 34(4): 1068-82, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-27095138

RESUMO

Folliculin (FLCN) is an autosomal dominant tumor suppressor gene that modulates diverse signaling pathways required for growth, proliferation, metabolism, survival, motility, and adhesion. FLCN is an essential protein required for murine embryonic development, embryonic stem cell (ESC) commitment, and Drosophila germline stem cell maintenance, suggesting that Flcn may be required for adult stem cell homeostasis. Conditional inactivation of Flcn in adult hematopoietic stem/progenitor cells (HSPCs) drives hematopoietic stem cells (HSC) into proliferative exhaustion resulting in the rapid depletion of HSPC, loss of all hematopoietic cell lineages, acute bone marrow (BM) failure, and mortality after 40 days. HSC that lack Flcn fail to reconstitute the hematopoietic compartment in recipient mice, demonstrating a cell-autonomous requirement for Flcn in HSC maintenance. BM cells showed increased phosphorylation of Akt and mTorc1, and extramedullary hematopoiesis was significantly reduced by treating mice with rapamycin in vivo, suggesting that the mTorc1 pathway was activated by loss of Flcn expression in hematopoietic cells in vivo. Tfe3 was activated and preferentially localized to the nucleus of Flcn knockout (KO) HSPCs. Tfe3 overexpression in HSPCs impaired long-term hematopoietic reconstitution in vivo, recapitulating the Flcn KO phenotype, and supporting the notion that abnormal activation of Tfe3 contributes to the Flcn KO phenotype. Flcn KO mice develop an acute histiocytic hyperplasia in multiple organs, suggesting a novel function for Flcn in macrophage development. Thus, Flcn is intrinsically required to maintain adult HSC quiescence and homeostasis, and Flcn loss leads to BM failure and mortality in mice.


Assuntos
Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Diferenciação Celular/genética , Estrona/genética , Células-Tronco Hematopoéticas/patologia , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Células da Medula Óssea/patologia , Linhagem da Célula/genética , Proliferação de Células/genética , Desenvolvimento Embrionário/genética , Células-Tronco Hematopoéticas/metabolismo , Homeostase/genética , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Camundongos Knockout
13.
Pol J Pathol ; 68(4): 284-290, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29517197

RESUMO

The entity of hereditary leiomyomatosis renal cell carcinoma (HLRCC)-associated RCC has been proposed and integrated into the recent International Society of Urologic Pathology (ISUP) of renal tumors. This tumor is characterized by presence of cutaneous and/or uterine leiomyomas and RCC and autosomal dominant hereditary form. Grossly, HLRCC arising in the kidney show the solid tumor with frequent partial cystic area. Microscopically, the tumor typically shows papillary RCC, type 2, with eosinophilic large nucleoli reminiscent of cytomegaloviral inclusion and perinuclear clearing/haloes. Immunohistochemically, tumor cells show the overexpression for 2SC and reduced expression of FH. Germline mutation of fumarate hydratase (FH) gene, the HLRCC responsible gene mapped to chromosome 1q43, has been identified in patients with HLRCC. As the renal cancer in patients with HLRCC generally behave aggressively even in a small size, complete surgical resection and retroperitoneal lymph node resection should be performed promptly when the tumor is discovered. The surveillance of renal tumor in FH gene germline mutation-positive patients should be started from the early age using ultrasound sonography or magnetic resonance imaging.


Assuntos
Carcinoma de Células Renais/patologia , Neoplasias Renais/patologia , Leiomiomatose/patologia , Síndromes Neoplásicas Hereditárias/patologia , Neoplasias Cutâneas/patologia , Neoplasias Uterinas/patologia , Biomarcadores Tumorais/análise , Biomarcadores Tumorais/genética , Biópsia , Carcinoma de Células Renais/química , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/cirurgia , Análise Mutacional de DNA , Diagnóstico Diferencial , Feminino , Predisposição Genética para Doença , Humanos , Imuno-Histoquímica , Neoplasias Renais/química , Neoplasias Renais/genética , Neoplasias Renais/cirurgia , Leiomiomatose/química , Leiomiomatose/genética , Leiomiomatose/cirurgia , Masculino , Mutação , Síndromes Neoplásicas Hereditárias/genética , Síndromes Neoplásicas Hereditárias/cirurgia , Fenótipo , Valor Preditivo dos Testes , Neoplasias Cutâneas/química , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/cirurgia , Resultado do Tratamento , Neoplasias Uterinas/química , Neoplasias Uterinas/genética , Neoplasias Uterinas/cirurgia
14.
Hum Mol Genet ; 23(21): 5706-19, 2014 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-24908670

RESUMO

Cardiac hypertrophy, an adaptive process that responds to increased wall stress, is characterized by the enlargement of cardiomyocytes and structural remodeling. It is stimulated by various growth signals, of which the mTORC1 pathway is a well-recognized source. Here, we show that loss of Flcn, a novel AMPK-mTOR interacting molecule, causes severe cardiac hypertrophy with deregulated energy homeostasis leading to dilated cardiomyopathy in mice. We found that mTORC1 activity was upregulated in Flcn-deficient hearts, and that rapamycin treatment significantly reduced heart mass and ameliorated cardiac dysfunction. Phospho-AMP-activated protein kinase (AMPK)-alpha (T172) was reduced in Flcn-deficient hearts and nonresponsive to various stimulations including metformin and AICAR (5-amino-1-ß-D-ribofuranosyl-imidazole-4-carboxamide). ATP levels were elevated and mitochondrial function was increased in Flcn-deficient hearts, suggesting that excess energy resulting from up-regulated mitochondrial metabolism under Flcn deficiency might attenuate AMPK activation. Expression of Ppargc1a, a central molecule for mitochondrial metabolism, was increased in Flcn-deficient hearts and indeed, inactivation of Ppargc1a in Flcn-deficient hearts significantly reduced heart mass and prolonged survival. Ppargc1a inactivation restored phospho-AMPK-alpha levels and suppressed mTORC1 activity in Flcn-deficient hearts, suggesting that up-regulated Ppargc1a confers increased mitochondrial metabolism and excess energy, leading to inactivation of AMPK and activation of mTORC1. Rapamycin treatment did not affect the heart size of Flcn/Ppargc1a doubly inactivated hearts, further supporting the idea that Ppargc1a is the critical element leading to deregulation of the AMPK-mTOR-axis and resulting in cardiac hypertrophy under Flcn deficiency. These data support an important role for Flcn in cardiac homeostasis in the murine model.


Assuntos
Cardiomegalia/genética , Cardiomegalia/metabolismo , Estrona/genética , Inativação Gênica , Complexos Multiproteicos/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Trifosfato de Adenosina/biossíntese , Animais , Cardiomegalia/complicações , Cardiomegalia/tratamento farmacológico , Cardiomegalia/patologia , Linhagem Celular , Modelos Animais de Doenças , Ativação Enzimática , Insuficiência Cardíaca/etiologia , Insuficiência Cardíaca/patologia , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Camundongos Transgênicos , Renovação Mitocondrial , Tamanho do Órgão/efeitos dos fármacos , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo , Fosforilação , Transdução de Sinais , Sirolimo/farmacologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Função Ventricular/efeitos dos fármacos
15.
Clin Genet ; 90(5): 403-412, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27220747

RESUMO

Birt-Hogg-Dubé syndrome (BHD) is a rare genetic disorder characterized by fibrofolliculomas, pulmonary cysts and renal cell carcinomas (RCCs). The affected individuals inherit germline mutations in the folliculin gene (FLCN). We investigated the mutation spectrum and clinicopathologic findings of 312 patients from 120 different families (119 Japanese and 1 Taiwanese). A total of 31 different FLCN sequence variants were identified. The majority were c.1285dupC (n = 34), c.1533_1536delGATG (n = 25), and c.1347_1353dupCCACCCT (n = 19). Almost all patients presented with pulmonary cysts. The incidence of RCCs in FLCN mutation carriers over the age of 40 was 34.8% (40/115). Fifty-five RCC lesions were surgically resected; most were either chromophobe RCC (n = 24; 43.6%) or hybrid oncocytic/chromophobe tumors (19; 34.5%). Seventy-six of 156 FLCN mutation carriers (120 probands and 36 sibs, 48.7%) had skin papules; however, cutaneous manifestations were so subtle that only one patient voluntarily consulted dermatologists. Japanese Asian BHD families have three FLCN mutational hotspots. Recurrent episodes of pneumothoraces are the major symptoms suggestive of a BHD diagnosis in our cohort. Characteristic features of lung and kidney lesions may be more informative than fibrofolliculomas as diagnostic criteria for BHD in the Japanese Asian population.


Assuntos
Síndrome de Birt-Hogg-Dubé/genética , Carcinoma de Células Renais/genética , Epidemiologia Molecular , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Síndrome de Birt-Hogg-Dubé/epidemiologia , Síndrome de Birt-Hogg-Dubé/fisiopatologia , Carcinoma de Células Renais/epidemiologia , Carcinoma de Células Renais/fisiopatologia , Cistos/fisiopatologia , Feminino , Mutação em Linhagem Germinativa , Humanos , Japão/epidemiologia , Pulmão/fisiopatologia , Masculino , Pessoa de Meia-Idade , Mutação , Pele/fisiopatologia
16.
Int J Urol ; 23(3): 204-10, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26608100

RESUMO

Birt-Hogg-Dubé syndrome is an autosomal dominantly inherited disease that predisposes patients to develop fibrofolliculoma, lung cysts and bilateral multifocal renal tumors, histologically hybrid oncocytic/chromophobe tumors, chromophobe renal cell carcinoma, oncocytoma, papillary renal cell carcinoma and clear cell renal cell carcinoma. The predominant forms of Birt-Hogg-Dubé syndrome-associated renal tumors, hybrid oncocytic/chromophobe tumors and chromophobe renal cell carcinoma are typically less aggressive, and a therapeutic principle for these tumors is a surgical removal with nephron-sparing. The timing of surgery is the most critical element for postoperative renal function, which is one of the important prognostic factors for Birt-Hogg-Dubé syndrome patients. The folliculin gene (FLCN) that is responsible for Birt-Hogg-Dubé syndrome was isolated as a novel tumor suppressor for kidney cancer. Recent studies using murine models for FLCN, a protein encoded by the FLCN gene, and its two binding partners, folliculin-interacting protein 1 (FNIP1) and folliculin-interacting protein 2 (FNIP2), have uncovered important roles for FLCN, FNIP1 and FNIP2 in cell metabolism, which include AMP-activated protein kinase-mediated energy sensing, Ppargc1a-driven mitochondrial oxidative phosphorylation and mTORC1-dependent cell proliferation. Birt-Hogg-Dubé syndrome is a hereditary hamartoma syndrome, which is triggered by metabolic alterations under a functional loss of FLCN/FNIP1/FNIP2 complex, a critical regulator of kidney cell proliferation rate; a mechanistic insight into the FLCN/FNIP1/FNIP2 pathway could provide us a basis for developing new therapeutics for kidney cancer.


Assuntos
Síndrome de Birt-Hogg-Dubé , Neoplasias Renais , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor/genética , Animais , Síndrome de Birt-Hogg-Dubé/genética , Síndrome de Birt-Hogg-Dubé/patologia , Síndrome de Birt-Hogg-Dubé/terapia , Proteínas de Transporte/metabolismo , Técnicas de Inativação de Genes , Humanos , Neoplasias Renais/genética , Neoplasias Renais/patologia , Neoplasias Renais/terapia , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Complexos Multiproteicos/metabolismo , Mutação , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologia , Neoplasias Experimentais/terapia , Nefrectomia , Proteínas Proto-Oncogênicas/metabolismo , Ratos , Sirolimo/administração & dosagem , Sirolimo/uso terapêutico , Serina-Treonina Quinases TOR/metabolismo , Proteínas Supressoras de Tumor/metabolismo
17.
Blood ; 120(6): 1254-61, 2012 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-22709692

RESUMO

Birt-Hogg-Dubé (BHD) syndrome is an autosomal dominant disorder characterized by cutaneous fibrofolliculomas, pulmonary cysts, and kidney malignancies. Affected individuals carry germ line mutations in folliculin (FLCN), a tumor suppressor gene that becomes biallelically inactivated in kidney tumors by second-hit mutations. Similar to other factors implicated in kidney cancer, FLCN has been shown to modulate activation of mammalian target of rapamycin (mTOR). However, its precise in vivo function is largely unknown because germ line deletion of Flcn results in early embryonic lethality in animal models. Here, we describe mice deficient in the newly characterized folliculin-interacting protein 1 (Fnip1). In contrast to Flcn, Fnip1(-/-) mice develop normally, are not susceptible to kidney neoplasia, but display a striking pro-B cell block that is entirely independent of mTOR activity. We show that this developmental arrest results from rapid caspase-induced pre-B cell death, and that a Bcl2 transgene reconstitutes mature B-cell populations, respectively. We also demonstrate that conditional deletion of Flcn recapitulates the pro-B cell arrest of Fnip1(-/-) mice. Our studies thus demonstrate that the FLCN-FNIP complex deregulated in BHD syndrome is absolutely required for B-cell differentiation, and that it functions through both mTOR-dependent and independent pathways.


Assuntos
Linfócitos B/fisiologia , Síndrome de Birt-Hogg-Dubé/genética , Proteínas de Transporte/genética , Diferenciação Celular/genética , Deleção de Genes , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor/genética , Animais , Linfócitos B/imunologia , Linfócitos B/metabolismo , Proteínas de Transporte/metabolismo , Proteínas de Transporte/fisiologia , Diferenciação Celular/imunologia , Células Cultivadas , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas/fisiologia , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Especificidade da Espécie , Proteínas Supressoras de Tumor/metabolismo , Proteínas Supressoras de Tumor/fisiologia
19.
Genes (Basel) ; 14(1)2023 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-36672892

RESUMO

The microphthalmia-associated transcription factor/transcription factor E (MiT/TFE) family of transcription factors are evolutionarily conserved, basic helix-loop-helix leucine zipper (bHLH-Zip) transcription factors, consisting of MITF, TFEB, TFE3, and TFEC. MiT/TFE proteins, with the exception of TFEC, are involved in the development of renal cell carcinoma (RCC). Most of the MiT/TFE transcription factor alterations seen in sporadic RCC cases of MiT family translocation renal cell carcinoma (tRCC) are chimeric proteins generated by chromosomal rearrangements. These chimeric MiT/TFE proteins retain the bHLH-Zip structures and act as oncogenic transcription factors. The germline variant of MITF p.E318K has been reported as a risk factor for RCC. E 318 is present at the SUMOylation consensus site of MITF. The p.E318K variant abrogates SUMOylation on K 316, which results in alteration of MITF transcriptional activity. Only a few cases of MITF p.E318K RCC have been reported, and their clinical features have not yet been fully described. It would be important for clinicians to recognize MITF p.E318K RCC and consider MITF germline testing for undiagnosed familial RCC cases. This review outlines the involvement of the MiT/TFE transcription factors in RCC, both in sporadic and hereditary cases. Further elucidation of the molecular function of the MiT/TFE family is necessary for better diagnosis and treatment of these rare diseases.


Assuntos
Carcinoma de Células Renais , Fator de Transcrição Associado à Microftalmia , Humanos , Fator de Transcrição Associado à Microftalmia/genética , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética
20.
Pathol Res Pract ; 242: 154313, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36669395

RESUMO

It is possible that PRCCs may still contain a variety of unknown histologic subtypes. Some PRCCs express high expression of TFE3 protein without TFE3 gene rearrangement, but no reports have investigated the significance of this. Here we attempted to examine clinicopathological and molecular significance of the TFE3-immunopositive PRCC. We reviewed the histology and immunohistochemistry in 58 PRCCs. TFE3 immunoexpression was recognized in 7 cases. Because TFE3 immunostaining shows false-positive, to ensure the integrity of TFE3 immunostaining, the immunostaining was performed under strict control of internal controls and western blotting was performed on 2 positive cases and 5 negative cases, and differences in protein expression between two groups were confirmed. Significant immunohistochemical expressions of autophagy/lysosome proteins were observed in TFE3-positive group. No TFE3 gene arrangement was detected in all positive cases by fluorescence in situ hybridization. Whole-exome sequencing was performed on 6 TFE3-positive and 2 TFE3-negative cases. Gain of chromosome 7 was found in five of 6 TFE3-positive cases (83%). TFE3-positive group was correlated significantly with higher pTstage, cNstage, WHO/ISUP nuclear grade, and decreased OS. TFE3-immunopositive PRCC group had a poorer prognosis than TFE3-negative PRCC group and showed correlation with expressions of autophagy/lysosome proteins, suggesting that enhancement of autophagy/lysosome function drives an environment of energy metabolism that is favorable for cancer. It is necessary to recognize that there is TFE3-immunopositive group without TFE3 gene rearrangement within PRCC. Because of its aggressive biological behaviour, TFE3 can act as a biomarker in PRCC; moreover, autophagy-inhibiting drugs may have therapeutic effects on TFE3-immunopositive PRCC.


Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Humanos , Carcinoma de Células Renais/patologia , Neoplasias Renais/patologia , Hibridização in Situ Fluorescente , Fatores de Transcrição/genética , Translocação Genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética
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