The Transposon Bari-Jheh in Drosophila melanogaster: adaptive phenotypes, molecular mechanisms, and genetic inheritance = El transposón Bari-Jheh en Drosophila melanogaster: fenotipos adaptativos, mecanismos moleculares y herencia genética

Abstract

Elucidating the fitness effects of natural genetic variants is one of the current major challenges in evolutionary biology. Understanding the interplay between genotype, phenotype and environment is necessary to make accurate predictions of important biological outcomes such as stress resistance or yield in economically important plants and animals, and disease in humans. Based on population frequency patterns and footprints of selection at the DNA level, the transposable element Bari-Jheh, inserted in the intergenic region of Juvenile Hormone Epoxy Hydrolase (Jheh) genes, was previously identified as putatively adaptive. However, the adaptive effect of this mutation remained elusive. In this thesis, we integrate information on transcription factor binding sites, available ChIP-Seq data, ChIP-PCR data generated by ourselves, gene expression analyses and phenotypic assays to identify the functional and the mechanistic underpinnings of Bari-Jheh. We show that Bari-Jheh adds extra antioxidant response elements upstream of Jheh1 and Jheh2 genes. Furthermore, we show that Bari-Jheh adds H3K27me3 histone marks and that this marks change under oxidative stress conditions. Thus, we observe that Bari-Jheh affects the expression of adjacent genes in non-stress and stress conditions. We find that Bari-Jheh is associated with down regulation of Jheh2 and Jheh3 under non stress conditions, as previously reported, and we find that Bari-Jheh is associated with up regulation of Jheh1 and/or Jheh2 and down regulation of up-stream gene Jheh3 under oxidative stress conditions. Besides these two molecular mechanisms described for Bari-Jheh under oxidative stress, we found that this transposable element is associated with resistance to oxidative stress induced by two different compounds relevant for natural D. melanogaster populations. The replicability of fitness effects on different genetic backgrounds also suggests that epistatic interactions do not seem to dominate the genetic architecture of oxidative stress resistance. We show that the resistance to oxidative stress is associated with the up-regulation of Jheh1 or Jheh2 indicating that the upregulation of only one of these genes is enough to confer resistance to oxidative stress. Moreover, since oxidative stress is produced in the gut by the organisms against bacterial infection, we further show that Bari-Jheh is associated with Pseudomonas entomophila oral infection. Thus, Bari-Jheh influences gene expression and enables stress response through two different

mechanisms, by adding cis-regulatory sequences and by adding histone marks, leading to changes in two ecologically relevant phenotypes. We also analyzed the dominance effect of Bari-Jheh. Although adaptive mutations are often considered to be dominant, it has been recently shown that a substantial proportion of adaptive mutations should display heterozygote advantage. The comparison of the survival curves of heterozygous and the two corresponding homozygous flies indicated that the dominance effect of Bari-Jheh depends on the genetic background. Our results add to the limited number of studies in which the dominance effect of adaptive mutations has been empirically estimated and highlights the complexity of their inheritance. Finally, we also find that TEs other than Bari-Jheh might be playing a role in the D. melanogaster response to oxidative stress. Overall our results contribute to the understanding of resistance to oxidative stress in natural populations and highlight the role of transposable elements in environmental adaptation.

Elucidar los efectos sobre la eficacia biológica de las variantes genéticas naturales es uno de los grandes retos actuales de la biología evolutiva. En esta tesis caracterizamos el elemento móvil Bari-Jheh, que está insertado en la región intergénica de los genes que codifican para las epoxi hidrolasas de la Hormona Juvenil (Jheh). Bari-Jheh se identificó como putativamente adaptativo si bien no se había encontrado el efecto adaptativo de esta mutación. Integrando información sobre los sitios de unión de factores de transcripción, datos disponibles de ChIP-Seq, análisis de expresión génica y ensayos fenotípicos identificamos el efecto adaptativo y los mecanismos moleculares por los que actúa Bari-Jheh. Mostramos que Bari-Jheh añade elementos adicionales de respuesta al estrés oxidativo aguas arriba de los genes Jheh1 y Jheh2. Además mostramos que Bari-Jheh añade marcas de histonas y que estas marcas cambian en condiciones de estrés oxidativo. En concordancia con estos datos Bari-Jheh está asociado con un incremento de la expresión de Jheh1 y/o Jheh2 y una disminución en la expresión de Jheh3. Además Bari-Jheh está asociado con resistencia a estrés oxidativo. Como el estrés oxidativo puede ser inducido de manera endógena en el intestino de Drosophila por la infección de bacterias Gram negativas, mostramos también que Bari-Jheh está asociado con la tolerancia a infección por Pseudomonas enotmophila. Además, mostramos que aparte de Bari-Jheh, otros elementos móviles podrían estar jugando un papel en la respuesta de D. melanogaster al estrés oxidativo. Finalmente, estudiamos el efecto de dominancia de Bari-Jheh. Si bien las mutaciones adaptativas son frecuentemente consideradas dominantes, recientemente se ha mostrado que una proporción importante de mutaciones adaptativas deberían mostrar ventajas en heterocigosis. La comparación de las curvas de supervivencia de moscas heterocigotas con los correspondientes homocigotos indica que la dominancia de Bari-Jheh depende del fondo genético. Nuestros resultados se añaden al limitado número de estudios en los que se ha estimado empíricamente la dominancia de una mutación adaptativa y destacan la complejidad de su herencia. En general, nuestros resultados contribuyen a la comprensión de la resistencia al estrés oxidativo en poblaciones naturales y ponen de relieve el papel de los elementos móviles en la adaptación ambiental.