MicroRNA-mediated regulation of p53 in Drosophila: a new role in adaptation to nutrient deprivation = Regulación de p53 por microARNs en Drosophila: una nueva función en la adaptación a la escasez nutricional

Abstract

The integration of nutrient status to metabolic homeostasis at the cellular and organismal level is a complex process performed in multicellular organisms, and the ability of an organism to respond to nutritional stress is critical for its survival. In the last few years, the tumour suppressor protein p53 has emerged as an important regulator of several metabolic pathways such as glycolysis, oxidative phosphorylation and autophagy, that triggers a cellular adaptive response to fluctuations in nutrient availability, a function that may contribute not only to tumour suppression activities of this molecule but also to its non-cancer-associated functions.

A better understanding of the molecular mechanisms underlying nutritional stress-mediated p53 activation and the potential role of p53 in organismal homeostasis is important to improve our current knowledge about p53 biology and its role in disease. In this regard, Drosophila is a very attractive model system whereby the physiology and the molecular mechanism that control metabolic homeostasis display significant parallels with humans. The fat body (FB), a functional analogue of vertebrate liver and adipose tissue, functions as a key sensor that couples nutrient status and energy expenditure. In conditions of nutrient deprivation, the FB supplies energy to the rest of the body by mobilizing stored glycogen and triacylglycerides (TAGs) to circulation in the form of trehalose and diacylglycerides (DAGs), respectively. The Drosophila homologue of p53 (Dp53) shares significant amino acid identity with mammalian p53, including specific residues frequently associated with human cancer. Like it mammalian counterpart, Dp53 promotes apoptotic cell death and cell cycle arrest in response to several stresses.

The results presented in this thesis reveal a fundamental role of Dp53 in the organismal adaptation to nutrient deprivation. The depletion of Dp53 activity levels specifically in FB cells accelerates the consumption of the main energy stores, reduces the levels of sugars in the animal, and compromises organismal survival upon fasting. We present evidence that Dp53 has an impact on energy balance independently on the regulation of the endocrine signaling network (adipokinetic hormone and insulin) involved in metabolic homeostasis and the metabolic enzymes involved in the mobilization of energy resources upon starvation. Interestingly, we unveil a cell autonomous role of Dp53 in modulating the metabolic changes of FB cells to nutrient deprivation. We also identified the molecular mechanism by which Dp53 is activated by nutrient conditions in FB cells. We show that microRNAs (miRNAs), an abundant class of endogenous non-coding RNAs measuring 22-23 nucleotides in lenght, regulate dp53 expression by targeting its 3’ UTR, and we identify miR-305 as a major regulatory element. Interestingly, two elements involved in the biogenesis of miRNAs, Drosha and Dicer, and one catalytic component of the RISC complex, Argonaute-1, are downregulated during starvation, thus alleviating miR-305-dependent targeting of the dp53-3’UTR and contributing to organismal resistance to starvation. These results open up new avenues towards the molecular understanding of p53 activation under metabolic stress and reveal the participation of p53 in nutrient sensing and metabolic adaptation at the organismal level.

During this thesis we also analyzed the classical tumour supressor functions of Dp53 and showed that upon ionizing radiation (IR)-induced DNA damage, Dp53 only regulates the early apoptotic response and induces DNA repair. Moreover, we present evidence that the miRNA machinery targets Dp53 in epithelial cells, thus preventing induction of programmed cell death. However, Dp53 activation upon DNA damage does not rely on the alleviation of miR305-mediated repression of the dp53-3’UTR.

As p53 is found in primitive organisms, and cancer is unlikely to have significantly influenced evolution, suppressing tumour formation was almost certainly not the original function of this gene. In the last few years, a link between p53 and metabolic homeostasis has been uncovered, and together with our results, we will like to speculate that regulation of cellular metabolism could be one of the primary functions of p53.

Los organismos han desarrollado a lo largo de la evolución una amplia gama de estrategias para mantener un estado homeostático interno a pesar de ciertas fluctuaciones ambientales tales como la disponibilidad de nutrientes. En los últimos años, p53 ha surgido como un importante regulador de varias rutas metabólicas, capaz de desencadenar una respuesta celular adaptativa a fluctuaciones en la disponibilidad de nutrientes, una función que puede contribuir no sólo a la capacidad supresora de tumores de p53, sino también a otras funciones más fisiológicas no asociadas con el cáncer. El homólogo de Drosophila de p53 (Dp53) comparte una identidad significativa en secuencia de aminoácidos con p53 de mamíferos, capaz de promover funciones similares en respuesta a varios tipos de estrés.

Los resultados presentados en esta tesis revelan un papel fundamental de Dp53 en la adaptación del organismo a la privación de nutrientes. Dp53 se requiere específicamente en el ‘cuerpo graso’ de Drosophila, un análogo funcional del hígado y el tejido adiposo de los vertebrados. La reducción de los niveles de actividad de Dp53 específicamente en este tejido acelera el consumo de las principales reservas de energía, reduce los niveles de azúcares en el animal, y compromete la supervivencia del organismo en ayuno. El impacto de Dp53 en el balance de energía es independiente a la regulación de las hormonas involucradas en el mantenimiento de la homeostasis metabólica y de las enzimas metabólicas implicadas en la movilización de los recursos energéticos. Desvelamos un papel autónomo-celular de Dp53 en la modulación de los cambios metabólicos que padecen las células del ‘cuerpo graso’ ante la privación de nutrientes. También identificamos el mecanismo molecular por el cual Dp53 se activa en condiciones de escasez nutricional en las células del ‘cuerpo graso’. Demostramos que los microRNAs (miRNAs) son capaces de regular la expresión dp53 e identificamos miR-305 como principal regulador. En una situación de privación de nutrientes, los niveles de expresión de varios elementos implicados en la biogénesis de los miRNAs se ven altamente reducidos, lo cual conlleva a la liberación de la represión mediada por miR-305, activando así Dp53 y contribuyendo a la resistencia del organismo frente a una situación de carencia nutricional.