WIP media la función oncogénica de p53 mutante

Tesis Doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 21-07-2015Cancer genetics has been profoundly studied from the last 50 years, showing that p53 is mutated in many types of human tumors. On the contrary to what is generally believed, p53 mutations cause not only the loss of its tumor suppression function, but also they lead to an accumulation of the mutated protein and the generation of a new aberrant function, currently known as oncogenic gain-of-function. This gain-of-function seems to favor the acquisition of more aggressive tumoral phenotypes and is associated to a poor prognosis. Our work demonstrates a key role of the oncogenic gain-of-function of mutated p53 based on the maintenance of the cancer stem cells (CSCs), linking p53 gain-offunction to a higher tumor malignancy. In the present study, we have identified some of the essential components in the mechanism of action of several p53 mutants. We have observed that WIP, an actin-associated protein, is increased through the oncogenic gain-of-function of mutated p53 and this increment leads to the stabilization of YAP/TAZ and its subsequent nuclear translocation, favoring the maintenance and growth of the CSC/mesenchymal phenotype and the invasive capacity of the tumor. The signaling and recycling of membrane receptors, like EGFR or integrins, is one of the processes affected by mutated p53 in coordination with other member of p53 family, p63. From these exacerbated signaling pathways we identify that AKT activity is essential, specifically the AKT2 isoform is required for WIP stabilization, and the final YAP/TAZ stabilization and activity. The oncogenic mechanisms discovered here have a key role in the establishment and maintenance of CSCs and consequently in the promotion of tumorigenesis in vitro and in vivo in glioblastoma and breast cancer models. The knowledge of these mechanisms will facilitate the design of novel drugs against these therapeutic targets in the tumor types analyzed.En los últimos 50 años se ha estudiado profundamente la genética del cáncer y se ha observado que p53 aparece mutado en muchos tipos de tumores humanos. A pesar de lo que se cree de forma genérica, las mutaciones en p53 no solo causan una pérdida de la función supresora tumoral, si no también una acumulación de esta proteína generando una segunda función actualmente denominada ganancia de función oncogénica. Esta ganancia de función parece favorecer la adquisición de fenotipos tumorales más agresivos y con peor pronóstico. Nuestro trabajo demuestra que la ganancia de función oncogénica de p53 mutante es crucial para el mantenimiento de las cancer stem cells (CSC), relacionándola de esta forma con una mayor capacidad tumoral. En el presente estudio, hemos identificado algunos de los componentes esenciales en el mecanismo de acción de diferentes mutantes de p53. Hemos observado que WIP, una proteína asociada y reguladora del citoesqueleto de actina, se ve incrementada por la ganancia de función oncogénica de p53 mutante. Este aumento de WIP conduce a la estabilización de YAP/TAZ y su consecuente tránsito y actividad nuclear, favoreciendo el mantenimiento y el crecimiento del fenotipo de stem/mesenquimal y la capacidad invasora del tumor. Uno de los procesos que se ven afectados por p53 mutante es el incremento de la señalización y del reciclaje de receptores de la membrana, como EGFR o integrinas mediado por la interacción de p53 mutante con otro miembro de la familia de p53, la proteína p63. En esta señalización exacerbada de p53 mutante/p63 hemos identificado que la actividad de AKT, específicamente la isoforma AKT2, es esencial para la estabilización de WIP y la estabilización y la actividad de YAP/TAZ. Creemos que los mecanismos oncogénicos hallados tienen un papel primordial en el establecimiento y el mantenimiento de CSC y por lo tanto en el desarrollo de la tumorigénesis in vitro e in vivo en los modelos tumorales de glioblastoma y de carcinoma de mama. El conocimiento de estos mecanismos podrá facilitar el diseño de nuevos fármacos contra estas dianas terapéuticas en los tipos de tumores analizados.Este trabajo ha sido realizado en el laboratorio del Doctor Francisco Wandosell Jurado en el Centro de Biología Molecular Severo Ochoa (CSIC-UAM). Desde Enero del 2010 hasta Agosto del 2010 ha sido financiado con un contrato de la red CIBERNED (ISCIII). Desde Septiembre del 2010 hasta Agosto del 2014 con una beca/contrato FPI del CSIC asociada al proyecto GDYCII ref. SAF2012-39148-C03-01

WIP Drives Tumor Progression through YAP/TAZ-Dependent Autonomous Cell Growth

In cancer, the deregulation of growth signaling pathways drives changes in the cell¿s architecture and its environment that allow autonomous growth of tumors. These cells then acquire a tumor-initiating ¿stemness¿ phenotype responsible for disease advancement to more aggressive stages. Here, we show that high levels of the actin cytoskeleton-associated protein WIP (WASP-interacting protein) correlates with tumor growth, both of which are linked to the tumor-initiating cell phenotype. We find that WIP controls tumor growth by boosting signals that stabilize the YAP/TAZ complex via a mechanism mediated by the endocytic/endosomal system. When WIP levels are high, the ß-catenin Adenomatous polyposis coli (APC)-axin-GSK3 destruction complex is sequestered to the multi-vesicular body compartment, where its capacity to degrade YAP/TAZ is inhibited. YAP/TAZ stability is dependent on Rac, p21-activated kinase (PAK) and mammalian diaphanous-related formin (mDia), and is Hippo independent. This close biochemical relationship indicates an oncogenic role for WIP in the physiology of cancer pathology by increasing YAP/TAZ stability.MINECO (SAF2013-45937-R to I.M.A.) and MINECO/FEDER (SAF2015-70368-R to I.M.A. and F.W.), the European Union (EU-FP7-2009-CT222887 to F.W.), the Instituto de Salud Carlos III Centro de Investigación Biomédica en Red (CIBERNED to F.W.) and by a grant from ISCIII-RETIC (RD12/0036/0009 to R.G.E.)Peer Reviewe

Síndrome De McKittrick-Wheelock Mediante Cirugía Transanal Mínimamente invasiva, dispositivo SILS.

Presentamos el caso de un paciente varón de 80 años de edad sin alergias conocidas y con antecedentes de riesgo cardiovascular, diabetes mellitus tipo II, insuficiencia renal (IR) crónica estadio III, nefropatía diabética, colon irritable de dos años de evolución e ingreso previo por insufiencia renal aguda de causa prerrenal, hiperpotasemia y acidosis metabólica que precisó diálisis

Transcription factor NFE2L2/NRF2 is a regulator of macroautophagy genes

Autophagy is a highly coordinated process that is controlled at several levels including transcriptional regulation. Here, we identify the transcription factor NFE2L2/NRF2 (nuclear factor, erythroid 2 like 2) as a regulator of autophagy gene expression and its relevance in a mouse model of Alzheimer disease (AD) that reproduces impaired APP (amyloid β precursor protein) and human (Hs)MAPT/TAU processing, clearance and aggregation. We screened the chromatin immunoprecipitation database ENCODE for 2 proteins, MAFK and BACH1, that bind the NFE2L2-regulated enhancer antioxidant response element (ARE). Using a script generated from the JASPAR's consensus ARE sequence, we identified 27 putative AREs in 16 autophagy-related genes. Twelve of these sequences were validated as NFE2L2 regulated AREs in 9 autophagy genes by additional ChIP assays and quantitative RT-PCR on human and mouse cells after NFE2L2 activation with sulforaphane. Mouse embryo fibroblasts of nfe2l2-knockout mice exhibited reduced expression of autophagy genes, which was rescued by an NFE2L2 expressing lentivirus, and impaired autophagy flux when exposed to hydrogen peroxide. NFE2L2-deficient mice co-expressing HsAPP(V717I) and HsMAPT(P301L), exhibited more intracellular aggregates of these proteins and reduced neuronal levels of SQSTM1/p62, CALCOCO2/NDP52, ULK1, ATG5 and GABARAPL1. Also, colocalization of HsAPP(V717I) and HsMAPT(P301L) with the NFE2L2-regulated autophagy marker SQSTM1/p62 was reduced in the absence of NFE2L2. In AD patients, neurons expressing high levels of APP or MAPT also expressed SQSTM1/p62 and nuclear NFE2L2, suggesting their attempt to degrade intraneuronal aggregates through autophagy. This study shows that NFE2L2 modulates autophagy gene expression and suggests a new strategy to combat proteinopathies

Transcription factor NRF2 uses the Hippo pathway effector TAZ to induce tumorigenesis in glioblastomas

Transcription factor NRF2 orchestrates a cellular defense against oxidative stress and, so far, has been involved in tumor progression by providing a metabolic adaptation to tumorigenic demands and resistance to chemotherapeutics. In this study, we discover that NRF2 also propels tumorigenesis in gliomas and glioblastomas by inducing the expression of the transcriptional co-activator TAZ, a protein of the Hippo signaling pathway that promotes tumor growth. The expression of the genes encoding NRF2 (NFE2L2) and TAZ (WWTR1) showed a positive correlation in 721 gliomas from The Cancer Genome Atlas database. Moreover, NRF2 and TAZ protein levels also correlated in immunohistochemical tissue arrays of glioblastomas. Genetic knock-down of NRF2 decreased, while NRF2 overexpression or chemical activation with sulforaphane, increased TAZ transcript and protein levels. Mechanistically, we identified several NRF2-regulated functional enhancers in the regulatory region of WWTR1. The relevance of the new NRF2/TAZ axis in tumorigenesis was demonstrated in subcutaneous and intracranial grafts. Thus, intracranial inoculation of NRF2-depleted glioma stem cells did not develop tumors as determined by magnetic resonance imaging. Forced TAZ overexpression partly rescued both stem cell growth in neurospheres and tumorigenicity. Hence, NRF2 not only enables tumor cells to be competent to proliferate but it also propels tumorigenesis by activating the TAZ-mediated Hippo transcriptional program.This study was funded by the Spanish Ministry of Economy and Competitiveness (MINECO) under the grant SAF2016-76520-R. ME is recipient of a postdoctoral contract Juan de la Cierva; DL and NRA of a FPU contract of MINECO; MP and RFG of a FPI contracts of Autonomous University of Madrid. RG has been funded by the AECC Scientific Foundation

Experimental Meningococcal Sepsis in Congenic Transgenic Mice Expressing Human Transferrin

Severe meningococcal sepsis is still of high morbidity and mortality. Its management may be improved by an experimental model allowing better understanding of its pathophysiology. We developed an animal model of meningococcal sepsis in transgenic BALB/c mice expressing human transferrin. We studied experimental meningococcal sepsis in congenic transgenic BALB/c mice expressing human transferrin by transcriptional profiling using microarray analysis of blood and brain samples. Genes encoding acute phase proteins, chemokines and cytokines constituted the largest strongly regulated groups. Dynamic bioluminescence imaging further showed high blood bacterial loads that were further enhanced after a primary viral infection by influenza A virus. Moreover, IL-1 receptor–associated kinase–3 (IRAK-3) was induced in infected mice. IRAK-3 is a negative regulator of Toll-dependant signaling and its induction may impair innate immunity and hence result in an immunocompromised state allowing bacterial survival and systemic spread during sepsis. This new approach should enable detailed analysis of the pathophysiology of meningococcal sepsis and its relationships with flu infection

Translocated LPS Might Cause Endotoxin Tolerance in Circulating Monocytes of Cystic Fibrosis Patients

Cystic Fibrosis (CF) is an inherited pleiotropic disease that results from abnormalities in the gene codes of a chloride channel. The lungs of CF patients are chronically infected by several pathogens but bacteraemia have rarely been reported in this pathology. Besides that, circulating monocytes in CF patients exhibit a patent Endotoxin Tolerance (ET) state since they show a significant reduction of the inflammatory response to bacterial stimulus. Despite a previous description of this phenomenon, the direct cause of ET in CF patients remains unknown. In this study we have researched the possible role of microbial/endotoxin translocation from a localized infection to the bloodstream as a potential cause of ET induction in CF patients. Plasma analysis of fourteen CF patients revealed high levels of LPS compared to healthy volunteers and patients who suffer from Chronic Obstructive Pulmonary Disease. Experiments in vitro showed that endotoxin concentrations found in plasma of CF patients were enough to induce an ET phenotype in monocytes from healthy controls. In agreement with clinical data, we failed to detect bacterial DNA in CF plasma. Our results suggest that soluble endotoxin present in bloodstream of CF patients causes endotoxin tolerance in their circulating monocytes

Wiskott-Aldrich syndrome protein-mediated actin dynamics control type-I interferon production in plasmacytoid dendritic cells

Mutations in Wiskott-Aldrich syndrome (WAS) protein (WASp), a regulator of actin dynamics in hematopoietic cells, cause WAS, an X-linked primary immunodeficiency characterized by recurrent infections and a marked predisposition to develop autoimmune disorders. The mechanisms that link actin alterations to the autoimmune phenotype are still poorly understood. We show that chronic activation of plasmacytoid dendritic cells (pDCs) and elevated type-I interferon (IFN) levels play a role in WAS autoimmunity. WAS patients display increased expression of type-I IFN genes and their inducible targets, alteration in pD