Functional analysis of the toll receptor protein family and their downstream signaling pathways in the central nervous system of Drosophila

Cell number plasticity drives organismal growth, and is coupled in the CNS to the emergence of neural circuits, ensuring appropriate function. In mammals, neurotrophins promote cell survival via Trk and p75$^{NTR}$ receptors or induce cell death via p75$^{NTR}$ and Sortilin. In $Drosophila$, DNTs bind Toll receptors promoting cell survival, but whether they regulate cell death within the CNS remains unknown. I show Toll receptors have distinct and overlapping spatial and temporal expression and functions. Driving RNAi knockdown and overexpression of each Toll, I show that different Toll receptors are required in glia for adult locomotion; in neurons for the regulation of VNC size; and to induce cell survival or death in distinct contexts. I focused on the signalling mechanisms downstream of Toll-6. My data show DNT-Toll-6 signalling switches between promoting cell survival or death via NFkB, ERK, or JNK signalling. These outcomes depend on the cleavage state of the DNT, time and available downstream adaptors. Toll-6 induces cell survival via MyD88 and cell death via dSarm, and these alternative outcomes depend on Weckle. Altogether, my data contribute to showing that the Toll receptors, DNTs and downstream signalling adaptors constitute a novel mechanism of cell number plasticity within the CNS

Toll and Toll-like receptor signalling in development

The membrane receptor Toll and the related Toll-like receptors (TLRs) are best known for their universal function in innate immunity. However, Toll/TLRs were initially discovered in a developmental context, and recent studies have revealed that Toll/TLRs carry out previously unanticipated functions in development, regulating cell fate, cell number, neural circuit connectivity and synaptogenesis. Furthermore, knowledge of their molecular mechanisms of action is expanding and has highlighted that Toll/TLRs function beyond the canonical NF-kappa B pathway to regulate cell-to-cell communication and signalling at the synapse. Here, we provide an overview of Toll/TLR signalling and discuss how this signalling pathway regulates various aspects of development across species

Three-tier regulation of cell number plasticity by neurotrophins and Tolls in Drosophila

Cell number plasticity is coupled to circuitry in the nervous system, adjusting cell mass to functional requirements. In mammals, this is achieved by neurotrophin (NT) ligands, which promote cell survival via their Trk and p75NTR receptors and cell death via p75NTR and Sortilin. Drosophila NTs (DNTs) bind Toll receptors instead to promote neuronal survival, but whether they can also regulate cell death is unknown. In this study, we show that DNTs and Tolls can switch from promoting cell survival to death in the central nervous system (CNS) via a three-tier mechanism. First, DNT cleavage patterns result in alternative signaling outcomes. Second, different Tolls can preferentially promote cell survival or death. Third, distinct adaptors downstream of Tolls can drive either apoptosis or cell survival. Toll-6 promotes cell survival via MyD88-NF-kappa B and cell death via Wek-Sarm-JNK. The distribution of adaptors changes in space and time and may segregate to distinct neural circuits. This novel mechanism for CNS cell plasticity may operate in wider contexts

Three-tier regulation of cell number plasticity by neurotrophins and Tolls in Drosophila

Cell number plasticity is coupled to circuitry in the nervous system, adjusting cell mass to functional requirements. In mammals, this is achieved by neurotrophin (NT) ligands, which promote cell survival via their Trk and p75 receptors and cell death via p75 and Sortilin. NTs (DNTs) bind Toll receptors instead to promote neuronal survival, but whether they can also regulate cell death is unknown. In this study, we show that DNTs and Tolls can switch from promoting cell survival to death in the central nervous system (CNS) via a three-tier mechanism. First, DNT cleavage patterns result in alternative signaling outcomes. Second, different Tolls can preferentially promote cell survival or death. Third, distinct adaptors downstream of Tolls can drive either apoptosis or cell survival. Toll-6 promotes cell survival via MyD88-NF-κB and cell death via Wek-Sarm-JNK. The distribution of adaptors changes in space and time and may segregate to distinct neural circuits. This novel mechanism for CNS cell plasticity may operate in wider contexts

Experimentally induced active and quiet sleep engage non-overlapping transcriptional programs in Drosophila

Sleep in mammals can be broadly classified into two different physiological categories: rapid eye movement (REM) sleep and slow-wave sleep (SWS), and accordingly REM and SWS are thought to achieve a different set of functions. The fruit fly Drosophila melanogaster is increasingly being used as a model to understand sleep functions, although it remains unclear if the fly brain also engages in different kinds of sleep as well. Here, we compare two commonly used approaches for studying sleep experimentally in Drosophila: optogenetic activation of sleep-promoting neurons and provision of a sleep-promoting drug, gaboxadol. We find that these different sleep-induction methods have similar effects on increasing sleep duration, but divergent effects on brain activity. Transcriptomic analysis reveals that drug-induced deep sleep (‘quiet’ sleep) mostly downregulates metabolism genes, whereas optogenetic ‘active’ sleep upregulates a wide range of genes relevant to normal waking functions. This suggests that optogenetics and pharmacological induction of sleep in Drosophila promote different features of sleep, which engage different sets of genes to achieve their respective functions

The neuronal ceroid lipofuscinosis protein Cln7 functions in the postsynaptic cell to regulate synapse development

The neuronal ceroid lipofuscinoses (NCLs) are a group of fatal, monogenic neurodegenerative disorders with an early onset in infancy or childhood. Despite identification of the genes disrupted in each form of the disease, their normal cellular role and how their deficits lead to disease pathology is not fully understood. Cln7, a major facilitator superfamily domain-containing protein, is affected in a late infantile-onset form of NCL. Cln7 is conserved across species suggesting a common function. Here we demonstrate that Cln7 is required for the normal growth of synapses at the Drosophila larval neuromuscular junction. In a Cln7 mutant, synapses fail to develop fully leading to reduced function and behavioral changes with dysregulation of TOR activity. Cln7 expression is restricted to the post-synaptic cell and the protein localizes to vesicles immediately adjacent to the post-synaptic membrane. Our data suggest an involvement for Cln7 in regulating trans-synaptic communication necessary for normal synapse development

A Toll-receptor map underlies structural brain plasticity

Experience alters brain structure, but the underlying mechanism remained unknown. Structural plasticity reveals that brain function is encoded in generative changes to cells that compete with destructive processes driving neurodegeneration. At an adult critical period, experience increases fiber number and brain size in . Here, we asked if Toll receptors are involved. Tolls demarcate a map of brain anatomical domains. Focusing on loss of function caused apoptosis, neurite atrophy and impaired behaviour. Toll-2 gain of function and neuronal activity at the critical period increased cell number. Toll-2 induced cycling of adult progenitor cells via a novel pathway, that antagonized MyD88-dependent quiescence, and engaged Weckle and Yorkie downstream. Constant knock-down of multiple synergistically reduced brain size. Conditional over-expression of and at the adult critical period increased brain size. Through their topographic distribution, Toll receptors regulate neuronal number and brain size, modulating structural plasticity in the adult brain