Gene expression has to withstand stochastic, environmental and genomic perturbations. For example, in the latter case, 0.5–1% of the human genome is typically variable between any two unrelated individuals. Such diversity might create problematic variability in the activity of gene regulatory networks, and ultimately, in cell behaviors. Using multigenerational selection experiments, we find that for the Drosophila proneural network, the effect of genomic diversity is dampened by miR-9a-mediated regulation of senseless expression. Reducing miR-9a regulation of the Senseless transcription factor frees the genomic landscape to exert greater phenotypic influence. Whole genome sequencing identified genomic loci that potentially exert such effects. A larger set of sequence variants, including variants within proneural network genes, exhibit these characteristics when miR-9a concentration is reduced. These findings reveal that microRNA-target interactions may be a key mechanism by which the impact of genomic diversity on cell behavior is dampened.

Metabolic conditions affect the developmental tempo of most animal species. Consequently, developmental gene regulatory networks (GRNs) must faithfully adjust their dynamics to a variable time scale. We find evidence that layered weak repression of genes provides the necessary coupling between GRN output and cellular metabolism. Using a mathematical model that replicates such a scenario, we find that lowering metabolism corrects developmental errors that otherwise occur when different layers of repression are lost. Through mutant analysis, we show that gene expression dynamics are unaffected by loss of repressors, but only when cellular metabolism is reduced. We further show that when metabolism is lowered, formation of a variety of sensory organs in Drosophila is normal despite loss of individual repressors of transcription, mRNA stability, and protein stability. We demonstrate the universality of this phenomenon by experimentally eliminating the entire microRNA family of repressors, and find that all microRNAs are rendered unnecessary when metabolism is reduced. Thus, layered weak repression provides robustness through error frequency suppression, and may provide an evolutionary route to a shorter reproductive cycle. ! 2. CC-BY-ND 4.0 International license It is made available under a (which was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.

Small ubiquitin-like modifier (SUMO) modification modulates the expression of defense genes in Drosophila, activated by the Toll/nuclear factor-κB and immune-deficient/nuclear factor-κB signaling networks. We have, however, limited understanding of the SUMO-modulated regulation of the immune response and lack information on SUMO targets in the immune system. In this study, we measured the changes to the SUMO proteome in S2 cells in response to a lipopolysaccharide challenge and identified 1619 unique proteins in SUMO-enriched lysates. A confident set of 710 proteins represents the immune-induced SUMO proteome and analysis suggests that specific protein domains, cellular pathways, and protein complexes respond to immune stress. A small subset of the confident set was validated by in-bacto SUMOylation and shown to be bona-fide SUMO targets. These include components of immune signaling pathways such as Caspar, Jra, Kay, cdc42, p38b, 14-3-3ε, as well as cellular proteins with diverse functions, many being components of protein complexes, such as prosß4, Rps10b, SmD3, Tango7, and Aats-arg. Caspar, a human FAF1 ortholog that negatively regulates immune-deficient signaling, is SUMOylated at K551 and responds to treatment with lipopolysaccharide in cultured cells. Our study is one of the first to describe SUMO proteome for the Drosophila immune response. Our data and analysis provide a global framework for the understanding of SUMO modification in the host response to pathogens.

Over the last decade, microRNAs have emerged as critical regulators in the expression and function of animal genomes. This review article discusses the relationship between microRNA-mediated regulation and the biology of the fruit fly Drosophila melanogaster. We focus on the roles that microRNAs play in tissue growth, germ cell development, hormone action, and the development and activity of the central nervous system. We also discuss the ways in which microRNAs affect robustness. Many gene regulatory networks are robust; they are relatively insensitive to the precise values of reaction constants and concentrations of molecules acting within the networks. MicroRNAs involved in robustness appear to be nonessential under uniform conditions used in conventional laboratory experiments. However, the robust functions of microRNAs can be revealed when environmental or genetic variation otherwise has an impact on developmental outcomes.

Sensory neuron numbers and positions are precisely organized to accurately map environmental signals in the brain. This precision emerges from biochemical processes within and between cells that are inherently stochastic. We investigated impact of stochastic gene expression on pattern formation, focusing on senseless (sens), a key determinant of sensory fate in Drosophila. Perturbing microRNA regulation or genomic location of sens produced distinct noise signatures. Noise was greatly enhanced when both sens alleles were present in homologous loci such that each allele was regulated in trans by the other allele. This led to disordered patterning. In contrast, loss of microRNA repression of sens increased protein abundance but not sensory pattern disorder. This suggests that gene expression stochasticity is a critical feature that must be constrained during development to allow rapid yet accurate cell fate resolution.