Sex-dependent and sex-independent regulatory systems of size variation in natural populations

Size of organs/organisms is a polygenic trait. Many of the growth-regulatory genes constitute conserved growth signaling pathways. However, how these multiple genes are orchestrated at the systems level to attain the natural variation in size including sexual size dimorphism is mostly unknown. Here we take a multi-layered systems omics approach to study size variation in the Drosophila wing. We show that expression levels of many critical growth regulators such as Wnt and TGF beta pathway components significantly differ between sexes but not between lines exhibiting size differences within each sex, suggesting a primary role of these regulators in sexual size dimorphism. Only a few growth genes including a receptor of steroid hormone ecdysone exhibit association with between-line size differences. In contrast, we find that between-line size variation is largely regulated by genes with a diverse range of cellular functions, most of which have never been implicated in growth. In addition, we show that expression quantitative trait loci (eQTLs) linked to these novel growth regulators accurately predict population-wide, between-line wing size variation. In summary, our study unveils differential gene regulatory systems that control wing size variation between and within sexes.

Elucidating regulatory mechanisms shaping the transcriptome and proteome of the gut immune response in Drosophila melanogaster

Michael Vincent Frochaux

Work on Drosophila melanogaster paved the way to our current understanding of modern genetics. Since then, this model organism has contributed greatly to various fields such as neurobiology, development, and immunology. The discovery and analysis of the various pathways of the Drosophila immune response led the way to the in-depth characterization of the gut immune response. Furthermore, a genome-wide association study on a population of Drosophila identified resistance to infection by the entomopathogenic bacteria Pseudomonas entomophila (P.e.) as a complex phenotype, with highly resistant and susceptible lines, and highlighted genes and pathways mediating inter-individual differences. However, the molecular mechanisms mediating the resistance to enteric infection remain largely uncharacterized. This study uses the same Drosophila population to analyze what are the molecular mechanisms mediating the resistance to enteric infection. We analyzed the genetic determinants of gene expression variation among the most resistant and susceptible lines in response to infection using expression quantitative trait loci (eQTL) analysis. We also characterized the mode of action of these eQTLs into cis- and trans-acting. Furthermore, we identified the gene nutcracker (ntc) as the only differentially-expressed gene between resistance classes. We then demonstrate that loss of function ntc mutants are significantly more susceptible to infection and then identify a single nucleotide polymorphism (SNP) located in a transcription factor binding site (TFBS) which affects the binding affinity of the transcription factor Broad leading to allele-specific expression variation of the gene ntc. If the transcriptomic response to infection has been thoroughly characterized, it is not the case for the proteomic response. We thus sought to characterize for the first time the proteomic response of the Drosophila to enteric infection. We performed mass spectrometry and RNA sequencing on dissected guts from flies infected by two Gram-negative bacteria, Erwinia carotovora carotovora 15 (Ecc15) or Pseudomonas entomophila, 4h and 16h after infection. We found that a large portion of the measurable proteome (12%) varies after infection and that protein changes are strongly time- and infection-dependent. We showed the relatively poor correlation between gene expression and protein abundance in the Drosophila enteric immune response. Finally, we performed a screen of several proteins that were identified in our proteomics work but had previously not been found when assessing the gene expression response to infection using loss-of-function mutants and identifying 7 genes that modulate overall susceptibility to P.e. infection. In summary, this work analyze the genetic determinants of gene expression variation in the DGRP population upon infection and characterize them according to their mode of action. Furthermore, we describe how a non-coding variant lowers resistance to infection by modulating ntc gene expression through altered Broad repressor binding. Finally, it provides the first comprehensive characterization of the Drosophila gut proteome upon infection by Gram-negative bacteria which can be the basis of future proteomic analysis of the enteric immune response

EPFL2019

Genomic Variation and Its Impact on Gene Expression in Drosophila melanogaster

Monica Albarca Aguilera, Bart Deplancke, Jeffrey David Jensen, Andreas Massouras, Sebastian Martin Waszak

Understanding the relationship between genetic and phenotypic variation is one of the great outstanding challenges in biology. To meet this challenge, comprehensive genomic variation maps of human as well as of model organism populations are required. Here, we present a nucleotide resolution catalog of single-nucleotide, multi-nucleotide, and structural variants in 39 Drosophila melanogaster Genetic Reference Panel inbred lines. Using an integrative, local assembly-based approach for variant discovery, we identify more than 3.6 million distinct variants, among which were more than 800,000 unique insertions, deletions (indels), and complex variants (1 to 6,000 bp). While the SNP density is higher near other variants, we find that variants themselves are not mutagenic, nor are regions with high variant density particularly mutation-prone. Rather, our data suggest that the elevated SNP density around variants is mainly due to population-level processes. We also provide insights into the regulatory architecture of gene expression variation in adult flies by mapping cis-expression quantitative trait loci (cis-eQTLs) for more than 2,000 genes. Indels comprise around 10% of all cis-eQTLs and show larger effects than SNP cis-eQTLs. In addition, we identified two-fold more gene associations in males as compared to females and found that most cis-eQTLs are sex-specific, revealing a partial decoupling of the genomic architecture between the sexes as well as the importance of genetic factors in mediating sex-biased gene expression. Finally, we performed RNA-seq-based allelic expression imbalance analyses in the offspring of crosses between sequenced lines, which revealed that the majority of strong cis-eQTLs can be validated in heterozygous individuals.

Public Library of Science2012

Elucidating regulatory mechanisms shaping the transcriptome and proteome of the gut immune response in Drosophila melanogaster

Michael Vincent Frochaux

EPFL2019