Periodic Reporting for period 4 - NEUROFLIES (Drosophila sechellia: a novel model to investigate nervous system and behavioral evolution)
Periodo di rendicontazione: 2024-04-01 al 2025-03-31
How animals’ extraordinarily diverse behaviours have evolved remains largely mysterious. Relating species-specific behaviours to differences in the anatomy or function of neural circuits offers a powerful approach to understand how nervous systems develop, function and change. The overall goal of this project was to establish a new model neurogenetic system for comparative neurobiology and behaviour, Drosophila sechellia. This species is closely related to the well-studied laboratory model D. melanogaster, and these species are consequently genetically and morphologically very similar. However, while D. melanogaster is a cosmopolitan, ecological generalist, D. sechellia is endemic to the Seychelles archipelago, where it has adapted to a highly-specialised niche, using the toxic Morinda citrifolia “noni” fruit as a sole host for feeding and breeding. As such, D. sechellia represents an exceptional model in which ecologically-relevant behavioural (and metabolic) adaptations can be studied, while taking advantage of the immense (neuro)biological knowledge and experimental toolkit of D. melanogaster.
The project had three main aims. Aim 1: to generate genetic tools in D. sechellia to manipulate the genes and neurons of this species to relate differences in molecular and cellular properties of the nervous system to its unique behaviours. Aim 2: to systematically compare the behaviours, the neuronal circuits and the neural gene expression patterns of D. sechellia with its close drosophilid cousins to identify both similarities and differences. Aim 3: to explore the genetic basis and functional significance of a previously-identified “simple” neuronal adaptation of D. sechellia: the increase in number of olfactory sensory neurons (OSNs) that sense an important host fruit odour.
As described below, the project firmly established a powerful genetically-tractable model system for evolutionary neuroscience (and many other fields) and provided several fundamental insights into the origins and mechanisms of nervous system and behavioural diversification. Knowing how brains and behaviours have been sculpted by random genetic mutation and natural selection in the past may enable future directed manipulation of the connectivity and activity of neural circuits, to enhance our ability to repair them.
The project had three main aims. Aim 1: to generate genetic tools in D. sechellia to manipulate the genes and neurons of this species to relate differences in molecular and cellular properties of the nervous system to its unique behaviours. Aim 2: to systematically compare the behaviours, the neuronal circuits and the neural gene expression patterns of D. sechellia with its close drosophilid cousins to identify both similarities and differences. Aim 3: to explore the genetic basis and functional significance of a previously-identified “simple” neuronal adaptation of D. sechellia: the increase in number of olfactory sensory neurons (OSNs) that sense an important host fruit odour.
As described below, the project firmly established a powerful genetically-tractable model system for evolutionary neuroscience (and many other fields) and provided several fundamental insights into the origins and mechanisms of nervous system and behavioural diversification. Knowing how brains and behaviours have been sculpted by random genetic mutation and natural selection in the past may enable future directed manipulation of the connectivity and activity of neural circuits, to enhance our ability to repair them.
original plans following unanticipated experimental challenges, new technological developments and/or serendipitous scientific observations. Aim 1. We made and validated a variety of tools in D. sechellia for labelling specific populations of neurons, for tracing neuronal architecture with fluorescent reporters, for physiological measurements of neuronal activity using calcium sensors, and for artificial neuronal activation with light-sensitive ion channels. We also generated and validated more general genetic tools for efficient integration of new transgenes into the genome. Aim 2. We surveyed a variety of behaviours of D. sechellia (e.g. chemosensory, visual, mechanosensory, oviposition, circadian as well as interactions with parasitoids and microbes) in comparison with those of D. melanogaster – and another close cousin, D. simulans – leading us to identify both conservation and divergence between these species. Notably, our study of circadian behaviours led us to discover a loss in plasticity of D. sechellia, in that it cannot adapt its circadian rhythms to changing day-length, which we linked to regulatory differences in a key circadian neuropeptide gene. Neuronal circuit comparisons focussed principally on the olfactory system, where functional significance can be more easily interpreted and investigated, and we characterised changes in cell number and projection patterns at several levels of this sensory system. The original proposal to survey neuronal gene expression patterns across these drosophilids by bulk RNA-sequencing of nervous system tissues was superseded by single cell-resolution transcriptomics approaches, allowing us to generate the first cross-species atlases for the whole central brain in any species; comparisons of these atlases pointed to multiple examples of D. sechellia-specific cell type representation and gene expression patterns, notably in glial populations. Aim 3. our efforts to map the genetic basis of OSN population expansions in D. sechellia revealed a surprisingly complex genetic architecture underlying these phenotypic differences, which unfortunately prevents pinpointing of the responsible genes. Greater progress was made in our understanding of the functional significance of these neuronal population increases, through comparative neurophysiological analyses in D. sechellia and D. melanogaster. We found that increased sensory pooling of OSNs results in reduced post-synaptic depression in their partner projection neurons in D. sechellia compared to D. melanogaster, enabling more persistent transmission of repeated or prolonged olfactory signals by these interneurons to higher brain centres. This phenomenon reveals a striking synergy with the changes in sensory sensitivity conferred by olfactory receptor tuning evolution. Taken together, this project has generated tools, datasets and results to reveal many new insights into how and why nervous systems have evolved. Moreover, it lays the foundation for a wealth of future research on other divergent behaviours and brain regions (as well as other phenotypes) of these drosophilid species, defining principles and mechanisms of relevance for all animals.
The project has gone beyond the state of the art technically or intellectually in three main ways. First, in our studies on the evolution of D. sechellia’s olfactory behaviours, we have provided the first causal genetic evidence explaining how changes in peripheral olfactory receptor tuning underlies species-specific odour preferences. Second, we have exploited new single-nuclear RNA sequencing technology to generate the first cross-species “whole” brain cellular atlases for D. sechellia (both on standard and noni diet), D. melanogaster and D. simulans to reveal changes in cellular composition and gene expression patterns of these species. This resource will be essential for our and other researchers’ further exploration of the neurobiology of known behavioural differences of these drosophilids, as well as provide a starting point for analysis of additional neuroanatomical, physiological and behavioural traits that have diverged between these species. Third, through our studies on circadian behavioural changes of D. sechellia we have found that this equatorial species has lost the ability to adapt to different photoperiods, which now reveals the promise of this species as a model for investigating the neurobiology and genetics of the evolution of “behavioural plasticity” (i.e. the ability to adapt to changing/unpredictable environmental selection pressures). Through these and other lines of investigation, we expect consolidation of D. sechellia as an exceptional model in evolutionary neuroscience, seeding many new concepts and technical/data resources for exploitation by research groups worldwide.