ABSTRACT
Sleep pressure and sleep depth are key regulators of wake and sleep. Current methods of measuring these parameters in Drosophila melanogaster have low temporal resolution and/or require disrupting sleep. Here we report analysis tools for high-resolution, noninvasive measurement of sleep pressure and depth from movement data. Probability of initiating activity, P(Wake), measures sleep depth while probability of ceasing activity, P(Doze), measures sleep pressure. In vivo and computational analyses show that P(Wake) and P(Doze) are largely independent and control the amount of total sleep. We also develop a Hidden Markov Model that allows visualization of distinct sleep/wake substates. These hidden states have a predictable relationship with P(Doze) and P(Wake), suggesting that the methods capture the same behaviors. Importantly, we demonstrate that both the Doze/Wake probabilities and the sleep/wake substates are tied to specific biological processes. These metrics provide greater mechanistic insight into behavior than measuring the amount of sleep alone.
Subject(s)
Circadian Rhythm/physiology , Drosophila melanogaster/physiology , Sleep/physiology , Wakefulness/physiology , Animals , Behavior, Animal/physiology , Humans , Models, Statistical , Movement/physiologyABSTRACT
Neurodevelopmental disorders arise from single or multiple gene defects. However, the way multiple loci interact to modify phenotypic outcomes remains poorly understood. Here, we studied phenotypes associated with mutations in the schizophrenia susceptibility gene dysbindin (dysb), in isolation or in combination with null alleles in the dysb network component Blos1. In humans, the Blos1 ortholog Bloc1s1 encodes a polypeptide that assembles, with dysbindin, into the octameric BLOC-1 complex. We biochemically confirmed BLOC-1 presence in Drosophila neurons, and measured synaptic output and complex adaptive behavior in response to BLOC-1 perturbation. Homozygous loss-of-function alleles of dysb, Blos1, or compound heterozygotes of these alleles impaired neurotransmitter release, synapse morphology, and homeostatic plasticity at the larval neuromuscular junction, and impaired olfactory habituation. This multiparameter assessment indicated that phenotypes were differentially sensitive to genetic dosages of loss-of-function BLOC-1 alleles. Our findings suggest that modification of a second genetic locus in a defined neurodevelopmental regulatory network does not follow a strict additive genetic inheritance, but rather, precise stoichiometry within the network determines phenotypic outcomes.
Subject(s)
Carrier Proteins/genetics , Drosophila Proteins/genetics , Gene Dosage/physiology , Nerve Net/physiology , Neuronal Plasticity/physiology , Schizophrenia/genetics , Synapses/genetics , Animals , Animals, Genetically Modified , Drosophila , Dysbindin , Dystrophin-Associated Proteins , Female , Nerve Net/ultrastructure , Schizophrenia/physiopathology , Synapses/ultrastructure , Synaptic Transmission/physiologyABSTRACT
Dysbindin is a schizophrenia susceptibility factor and subunit of the biogenesis of lysosome-related organelles complex 1 (BLOC-1) required for lysosome-related organelle biogenesis, and in neurons, synaptic vesicle assembly, neurotransmission, and plasticity. Protein networks, or interactomes, downstream of dysbindin/BLOC-1 remain partially explored despite their potential to illuminate neurodevelopmental disorder mechanisms. Here, we conducted a proteome-wide search for polypeptides whose cellular content is sensitive to dysbindin/BLOC-1 loss of function. We identified components of the vesicle fusion machinery as factors downregulated in dysbindin/BLOC-1 deficiency in neuroectodermal cells and iPSC-derived human neurons, among them the N-ethylmaleimide-sensitive factor (NSF). Human dysbindin/BLOC-1 coprecipitates with NSF and vice versa, and both proteins colocalized in a Drosophila model synapse. To test the hypothesis that NSF and dysbindin/BLOC-1 participate in a pathway-regulating synaptic function, we examined the role for NSF in dysbindin/BLOC-1-dependent synaptic homeostatic plasticity in Drosophila. As previously described, we found that mutations in dysbindin precluded homeostatic synaptic plasticity elicited by acute blockage of postsynaptic receptors. This dysbindin mutant phenotype is fully rescued by presynaptic expression of either dysbindin or Drosophila NSF. However, neither reduction of NSF alone or in combination with dysbindin haploinsufficiency impaired homeostatic synaptic plasticity. Our results demonstrate that dysbindin/BLOC-1 expression defects result in altered cellular content of proteins of the vesicle fusion apparatus and therefore influence synaptic plasticity.
Subject(s)
Drosophila Proteins/metabolism , Dystrophin-Associated Proteins/metabolism , Gene Expression Regulation/physiology , N-Ethylmaleimide-Sensitive Proteins/metabolism , Neuronal Plasticity/physiology , Synapses/physiology , Animals , Animals, Genetically Modified , Cell Line, Tumor , Drosophila , Drosophila Proteins/genetics , Dysbindin , Dystrophin-Associated Proteins/genetics , Gene Expression Regulation/genetics , Humans , Melanoma/pathology , N-Ethylmaleimide-Sensitive Proteins/genetics , Nerve Net/physiology , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Neuroblastoma/pathology , Neuromuscular Junction/genetics , Neuromuscular Junction/metabolism , Neuronal Plasticity/drug effects , Neuronal Plasticity/genetics , SNARE Proteins/metabolism , Synapses/genetics , Synaptic Vesicles/genetics , Synaptic Vesicles/metabolismABSTRACT
Neural activity has profound effects on the development of dendritic structure. Mechanisms that link neural activity to nuclear gene expression include activity-regulated factors, such as CREB, Crest or Mef2, as well as activity-regulated immediate-early genes, such as fos and jun. This study investigates the role of the transcriptional regulator AP-1, a Fos-Jun heterodimer, in activity-dependent dendritic structure development. We combine genetic manipulation, imaging and quantitative dendritic architecture analysis in a Drosophila single neuron model, the individually identified motoneuron MN5. First, Dα7 nicotinic acetylcholine receptors (nAChRs) and AP-1 are required for normal MN5 dendritic growth. Second, AP-1 functions downstream of activity during MN5 dendritic growth. Third, using a newly engineered AP-1 reporter we demonstrate that AP-1 transcriptional activity is downstream of Dα7 nAChRs and Calcium/calmodulin-dependent protein kinase II (CaMKII) signaling. Fourth, AP-1 can have opposite effects on dendritic development, depending on the timing of activation. Enhancing excitability or AP-1 activity after MN5 cholinergic synapses and primary dendrites have formed causes dendritic branching, whereas premature AP-1 expression or induced activity prior to excitatory synapse formation disrupts dendritic growth. Finally, AP-1 transcriptional activity and dendritic growth are affected by MN5 firing only during development but not in the adult. Our results highlight the importance of timing in the growth and plasticity of neuronal dendrites by defining a developmental period of activity-dependent AP-1 induction that is temporally locked to cholinergic synapse formation and dendritic refinement, thus significantly refining prior models derived from chronic expression studies.
Subject(s)
Dendrites/metabolism , Drosophila melanogaster/metabolism , Gene Expression Regulation, Developmental , Transcription Factor AP-1/metabolism , Transcription, Genetic , Animals , Animals, Genetically Modified , Calcium-Calmodulin-Dependent Protein Kinase Type 2/genetics , Calcium-Calmodulin-Dependent Protein Kinase Type 2/metabolism , Cell Nucleus/genetics , Cell Nucleus/metabolism , Cholinergic Neurons/metabolism , Dendrites/genetics , Drosophila Proteins/genetics , Drosophila Proteins/metabolism , Drosophila melanogaster/embryology , Drosophila melanogaster/genetics , Embryo, Nonmammalian/cytology , Embryo, Nonmammalian/embryology , Embryo, Nonmammalian/metabolism , Enzyme Activation , Genes, Reporter , Immunohistochemistry/methods , Microscopy, Confocal/methods , Microscopy, Fluorescence , Receptors, Nicotinic/genetics , Receptors, Nicotinic/metabolism , Signal Transduction , Synaptic Transmission , Time Factors , Transcription Factor AP-1/genetics , Transcriptional ActivationABSTRACT
Dr. K. S. Krishnan was on the faculty of the Division of Biological Sciences at the Tata Institute of Fundamental Research (TIFR) in Mumbai, India, and later emeritus professor at the National Center for Biological Sciences (NCBS) in Bangalore, India. His research using fruit flies has contributed richly to our understanding of synaptic function and mechanisms of anesthetic action. Dr. Krishnan passed away suddenly of a heart attack on the 24th of May, 2014. Below a few of his students fondly recall how it was to work in his group.
Subject(s)
Electrophysiology/history , Molecular Biology/history , Research/history , Academies and Institutes/history , Aged , History, 20th Century , History, 21st Century , Humans , India , MaleABSTRACT
Memory deficits in Drosophila nalyot mutants suggest that the Myb family transcription factor Adf-1 is an important regulator of developmental plasticity in the brain. However, the cellular functions for this transcription factor in neurons or molecular mechanisms by which it regulates plasticity remain unknown. Here, we use in vivo 3D reconstruction of identifiable larval motor neuron dendrites to show that Adf-1 is required cell autonomously for dendritic development and activity-dependent plasticity of motor neurons downstream of CaMKII. Adf-1 inhibition reduces dendrite growth and neuronal excitability, and results in motor deficits and altered transcriptional profiles. Surprisingly, analysis by comparative chromatin immunoprecipitation followed by sequencing (ChIP-Seq) of Adf-1, RNA Polymerase II (Pol II), and histone modifications in Kc cells shows that Adf-1 binding correlates positively with high Pol II-pausing indices and negatively with active chromatin marks such as H3K4me3 and H3K27ac. Consistently, the expression of Adf-1 targets Staufen and Fasciclin II (FasII), identified through larval brain ChIP-Seq for Adf-1, is negatively regulated by Adf-1, and manipulations of these genes predictably modify dendrite growth. Our results imply mechanistic interactions between transcriptional and local translational machinery in neurons as well as conserved neuronal growth mechanisms mediated by cell adhesion molecules, and suggest that CaMKII, Adf-1, FasII, and Staufen influence crucial aspects of dendrite development and plasticity with potential implications for memory formation. Further, our experiments reveal molecular details underlying transcriptional regulation by Adf-1, and indicate active interaction between Adf-1 and epigenetic regulators of gene expression during activity-dependent neuronal plasticity.
Subject(s)
Calcium-Calmodulin-Dependent Protein Kinase Type 2/metabolism , Cell Adhesion Molecules, Neuronal/metabolism , Dendrites/physiology , Drosophila Proteins/metabolism , Neurons/metabolism , RNA-Binding Proteins/metabolism , Transcription Factors/metabolism , Animals , Behavior, Animal/physiology , Cell Adhesion Molecules, Neuronal/genetics , Dendrites/genetics , Dendrites/metabolism , Drosophila , Drosophila Proteins/genetics , Gene Expression Regulation, Developmental , Larva/genetics , Larva/metabolism , Neuronal Plasticity/physiology , RNA-Binding Proteins/genetics , Transcription Factors/geneticsABSTRACT
Despite its ubiquity and significance, behavioral habituation is poorly understood in terms of the underlying neural circuit mechanisms. Here, we present evidence that habituation arises from potentiation of inhibitory transmission within a circuit motif commonly repeated in the nervous system. In Drosophila, prior odorant exposure results in a selective reduction of response to this odorant. Both short-term (STH) and long-term (LTH) forms of olfactory habituation require function of the rutabaga-encoded adenylate cyclase in multiglomerular local interneurons (LNs) that mediate GABAergic inhibition in the antennal lobe; LTH additionally requires function of the cAMP response element-binding protein (CREB2) transcription factor in LNs. The odorant selectivity of STH and LTH is mirrored by requirement for NMDA receptors and GABA(A) receptors in odorant-selective, glomerulus-specific projection neurons(PNs). The need for the vesicular glutamate transporter in LNs indicates that a subset of these GABAergic neurons also releases glutamate. LTH is associated with a reduction of odorant-evoked calcium fluxes in PNs as well as growth of the respective odorant-responsive glomeruli. These cellular changes use similar mechanisms to those required for behavioral habituation. Taken together with the observation that enhancement of GABAergic transmission is sufficient to attenuate olfactory behavior, these data indicate that habituation arises from glomerulus-selective potentiation of inhibitory synapses in the antennal lobe. We suggest that similar circuit mechanisms may operate in other species and sensory systems.
Subject(s)
Habituation, Psychophysiologic/physiology , Neuronal Plasticity/physiology , Neurons/metabolism , Smell/physiology , Adenylyl Cyclases/genetics , Adenylyl Cyclases/metabolism , Animals , Cyclic AMP Response Element-Binding Protein/genetics , Cyclic AMP Response Element-Binding Protein/metabolism , Drosophila Proteins/genetics , Drosophila Proteins/metabolism , Drosophila melanogaster , Receptors, GABA-A/genetics , Receptors, GABA-A/metabolism , Receptors, N-Methyl-D-Aspartate/genetics , Receptors, N-Methyl-D-Aspartate/metabolism , Receptors, Odorant/genetics , Receptors, Odorant/metabolism , Trans-Activators/genetics , Trans-Activators/metabolismABSTRACT
Here we report a human intellectual disability disease locus on chromosome 14q31.3 corresponding to mutation of the ZC3H14 gene that encodes a conserved polyadenosine RNA binding protein. We identify ZC3H14 mRNA transcripts in the human central nervous system, and we find that rodent ZC3H14 protein is expressed in hippocampal neurons and colocalizes with poly(A) RNA in neuronal cell bodies. A Drosophila melanogaster model of this disease created by mutation of the gene encoding the ZC3H14 ortholog dNab2, which also binds polyadenosine RNA, reveals that dNab2 is essential for development and required in neurons for normal locomotion and flight. Biochemical and genetic data indicate that dNab2 restricts bulk poly(A) tail length in vivo, suggesting that this function may underlie its role in development and disease. These studies reveal a conserved requirement for ZC3H14/dNab2 in the metazoan nervous system and identify a poly(A) RNA binding protein associated with a human brain disorder.
Subject(s)
Drosophila Proteins/genetics , Drosophila Proteins/physiology , Intellectual Disability/genetics , Mutation , Nuclear Proteins/genetics , Nuclear Proteins/physiology , RNA-Binding Proteins/genetics , RNA-Binding Proteins/physiology , Adolescent , Adult , Amino Acid Sequence , Animals , Central Nervous System/physiology , Chromosome Mapping , Chromosomes, Human, Pair 14/genetics , Cohort Studies , Consanguinity , Conserved Sequence , Drosophila melanogaster/genetics , Drosophila melanogaster/physiology , Evolution, Molecular , Female , Flight, Animal/physiology , Gene Knockdown Techniques , Genes, Recessive , Hippocampus/metabolism , Humans , Iran , Male , Models, Animal , Molecular Sequence Data , Pedigree , Poly(A)-Binding Proteins , RNA, Messenger/genetics , RNA, Messenger/metabolism , Sequence Homology, Amino Acid , Young Adult , Zinc Fingers/geneticsABSTRACT
AIM: Characterize Growth Differentiation Factor 15 (GDF15) as a secreted biomarker of the integrated stress response (ISR) within the central nervous system (CNS). METHODS: We determined GDF15 levels utilizing in vitro and in vivo neuronal systems wherein the ISR was activated. Primarily, we used the murine model of vanishing white matter disease (VWMD), a neurological disease driven by persistent ISR in the CNS, to establish a link between levels of GDF15 in the cerebrospinal fluid (CSF) and ISR gene expression signature in the CNS. GDF15 was also determined in the CSF of VWM patients. RESULTS: GDF15 expression was increased concomitant to ISR activation in stress-induced primary astrocytes as well as in retinal ganglion cells following optic nerve crush, while treatment with 2Bact, a specific eIF2B activator, suppressed both the ISR and GDF15. In the VWMD model, CSF GDF15 levels corresponded with the magnitude of the ISR and were reduced by 2BAct. In VWM patients, mean CSF GDF15 was elevated >20-fold as compared to healthy controls, whereas plasma GDF15 was undifferentiated. CONCLUSIONS: These data suggest that CSF GDF15 is a dynamic marker of ISR activation in the CNS and may serve as a pharmacodynamic biomarker for ISR-modulating therapies.
Subject(s)
Growth Differentiation Factor 15 , Leukoencephalopathies , Humans , Mice , Animals , Growth Differentiation Factor 15/genetics , Leukoencephalopathies/genetics , Central Nervous System/metabolism , Eukaryotic Initiation Factor-2B/genetics , Eukaryotic Initiation Factor-2B/metabolism , BiomarkersABSTRACT
By the middle of the 20th century, development of powerful genetic approaches had ensured that the fruit fly would remain a model organism of choice for genetic and developmental studies. But in the 1970s, a few pioneering groups turned their attention to the prospect of using the fly for neurophysiological experiments. They proposed that in a poikilothermic organism such as Drosophila, temperature-sensitive or "ts" mutations in proteins that controlled nerve function would translate to a "ts" paralytic phenotype. This was by no means an obvious or even a likely assumption. However, following directed screens these groups soon reported dramatic demonstrations of reversible ts paralysis in fly mutants. Resultantly, these "simple" experiments led to the isolation of a number of conditional mutations including shibire, paralytic, and comatose. All have since been cloned and have enabled deep mechanistic insights into synaptic transmission and nerve conduction. comatose (comt) mutations, for example, were found to map to missense changes in dNSF1, a neuron-specific fly homolog of mammalian NSF (N-ethylmaleimide-sensitive fusion factor). Studies on comt were also some of the first to discriminate between nuanced models of NSF function during presynaptic transmitter release that have since been borne out by experiments in multiple preparations. Here, the authors present an overview of NSF function as it is understood today, with an emphasis on contributions from Drosophila beginning with experiments carried out by Obaid Siddiqi in the Benzer laboratory. The authors also outline initial results from a genetic screen for phenotypic modifiers of comt that hold the promise of further elucidating NSF function at the synapse. Over the years, the neuromuscular system of Drosophila has served as a uniquely accessible model to unravel mechanisms underlying synaptic transmission. To this day, ts paralysis remains one of the most emphatic demonstrations of nerve function in an intact organism.
Subject(s)
Mutation/genetics , N-Ethylmaleimide-Sensitive Proteins/genetics , N-Ethylmaleimide-Sensitive Proteins/physiology , Neurons/metabolism , Animals , Animals, Genetically Modified , Drosophila/genetics , Drosophila Proteins , Electroretinography , Ethyl Methanesulfonate/pharmacology , Exocytosis/drug effects , Exocytosis/genetics , Genetic Testing , History, 20th Century , Mutagenesis/drug effects , Mutagenesis/physiology , N-Ethylmaleimide-Sensitive Proteins/history , Paralysis/genetics , Paralysis/therapy , Protein Transport/drug effects , Protein Transport/genetics , Repressor Proteins/genetics , Repressor Proteins/metabolism , Synapses/drug effects , Synapses/genetics , Synaptic Vesicles/metabolism , Time FactorsABSTRACT
The calcium-regulated transcription factor NFAT is emerging as a key regulator of neuronal development and plasticity but precise cellular consequences of NFAT function remain poorly understood. Here, we report that the single Drosophila NFAT homolog is widely expressed in the nervous system including motor neurons and unexpectedly controls neural excitability. Likely due to this effect on excitability, NFAT regulates overall larval locomotion and both chronic and acute forms of activity-dependent plasticity at the larval glutamatergic neuro-muscular synapse. Specifically, NFAT-dependent synaptic phenotypes include changes in the number of pre-synaptic boutons, stable modifications in synaptic microtubule architecture and pre-synaptic transmitter release, while no evidence is found for synaptic retraction or alterations in the level of the synaptic cell adhesion molecule FasII. We propose that NFAT regulates pre-synaptic development and constrains long-term plasticity by dampening neuronal excitability.
Subject(s)
Drosophila melanogaster/physiology , NFATC Transcription Factors/metabolism , Neurogenesis , Neuronal Plasticity/physiology , Presynaptic Terminals/metabolism , Animals , Blotting, Western , Larva , Motor Neurons/metabolism , Neuromuscular Junction/growth & development , Neuromuscular Junction/metabolism , Neurotransmitter Agents/metabolism , Patch-Clamp TechniquesABSTRACT
Recent work indicates a role for RING finger protein 11 (RNF11) in Parkinson disease (PD) pathology, which involves the loss of dopaminergic neurons. However, the role of RNF11 in regulating dopamine neurotransmission has not been studied. In this work, we tested the effect of RNF11 RNAi knockdown or overexpression on stimulated dopamine release in the larval Drosophila central nervous system. Dopamine release was stimulated using optogenetics and monitored in real-time using fast-scan cyclic voltammetry at an electrode implanted in an isolated ventral nerve cord. RNF11 knockdown doubled dopamine release, but there was no decrease in dopamine from RNF11 overexpression. RNF11 knockdown did not significantly increase stimulated serotonin or octopamine release, indicating the effect is dopamine specific. Dopamine clearance was also changed, as RNF11 RNAi flies had a higher Vmax and RNF11 overexpressing flies had a lower Vmax than control flies. RNF11 RNAi flies had increased mRNA levels of dopamine transporter (DAT) in RNF11, confirming changes in DAT. In RNF11 RNAi flies, release was maintained better for stimulations repeated at short intervals, indicating increases in the recycled releasable pool of dopamine. Nisoxetine, a DAT inhibitor, and flupenthixol, a D2 antagonist, did not affect RNF11 RNAi or overexpressing flies differently than control. Thus, RNF11 knockdown causes early changes in dopamine neurotransmission, and this is the first work to demonstrate that RNF11 affects both dopamine release and uptake. RNF11 expression decreases in human dopaminergic neurons during PD, and that decrease may be protective by increasing dopamine neurotransmission in the surviving dopaminergic neurons.
Subject(s)
Dopamine Plasma Membrane Transport Proteins/genetics , Dopamine , Drosophila Proteins/genetics , Drosophila , Animals , Central Nervous System , DNA-Binding Proteins , Dopaminergic Neurons , Humans , SerotoninABSTRACT
As organisms age, they often accumulate protein aggregates that are thought to be toxic, potentially leading to age-related diseases. This accumulation of protein aggregates is partially attributed to a failure to maintain protein homeostasis. A variety of genetic factors have been linked to longevity, but how these factors also contribute to protein homeostasis is not completely understood. In order to understand the relationship between aging and protein aggregation, we tested how a gene that regulates lifespan and age-dependent locomotor behaviors, p38 MAPK (p38Kb), influences protein homeostasis as an organism ages. We find that p38Kb regulates age-dependent protein aggregation through an interaction with starvin, a regulator of muscle protein homeostasis. Furthermore, we have identified Lamin as an age-dependent target of p38Kb and starvin.
Subject(s)
Aging/metabolism , Drosophila Proteins/metabolism , Drosophila melanogaster/metabolism , Longevity/genetics , MAP Kinase Signaling System/genetics , Proteostasis/genetics , p38 Mitogen-Activated Protein Kinases/metabolism , Aging/genetics , Animals , Animals, Genetically Modified , Drosophila Proteins/genetics , Drosophila melanogaster/genetics , Female , Gene Deletion , Lamins/metabolism , Locomotion/genetics , Macroautophagy/genetics , Muscles/metabolism , Oxidative Stress/genetics , Phenotype , Proteolysis , RNA Interference , p38 Mitogen-Activated Protein Kinases/geneticsABSTRACT
Since its description more than fifteen years ago, the GAL4-UAS system of heterologous transgenic expression has found universal and widespread use in Drosophila research , making it a uniquely powerful analytical tool. Several hundreds of enhancer-trap GAL4 "driver" lines have since been used to express proteins of interest in specific spatio-temporal domains. However, the identities of enhancer elements that regulate GAL4 expression in vivo are often not known. Here, I report the mapping of three GAL4 lines commonly used as motor neuron drivers. Sequencing of genomic DNA flanking these three P-element transposon insertion, C380, (BG380), OK6, and D42, shows that these insertions lie upstream of the futsch, Rapgap 1 and toll-6 gene, respectively . Of the three, OK6-GAL4 (Rapgap 1) expression is most restricted to motor neurons, while C380-GAL4 and D42-GAL4 also show prominent expression in the peripheral nervous system, including body wall sensory nervous system. Albeit with clear differences. Finally, I test if the highly restricted expression pattern of Futsch is maintained in six other species of Drosophilids (D. yakuba, D. ananssae, D.pseudoobscura, D. dimulars, D. willistoni and D. virilis). My results suggest conserved control of Futsch expression across species, most likely through upstream cis-acting elements. A comparative anatomy of the laval central nervous systems and peripheral innervation in these Frosophilids species as revealed by contemporary immunohistochemical markers is also presented.
Subject(s)
Drosophila/genetics , Gene Expression Profiling/methods , Genomics/methods , Animals , Animals, Genetically Modified , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Drosophila/classification , Drosophila/metabolism , Drosophila Proteins/genetics , Drosophila Proteins/metabolism , ELAV Proteins/genetics , ELAV Proteins/metabolism , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Immunohistochemistry , Microscopy, Confocal , Microtubule-Associated Proteins/genetics , Microtubule-Associated Proteins/metabolism , Polymerase Chain Reaction , Repressor Proteins/genetics , Repressor Proteins/metabolism , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae Proteins/metabolism , Species Specificity , Transcription Factors/genetics , Transcription Factors/metabolismABSTRACT
The increase in human life expectancy is accompanied by age-related cognitive and motor disability, thus raising the demand for strategies toward healthy aging. This requires understanding the biology of normal aging and late-life functional phenotypes. Genetic model organisms, such as Drosophila melanogaster, can help identifying evolutionary conserved mechanisms underlying aging. Longitudinal assessment of motor performance of more than 1000 individual flies revealed age-related motor performance decline and specific late-life motor disabilities. This allows defining heath- and ill-span and scoring late-life quality of individual flies. As in mammals, including humans, onset, duration, severity, and progression dynamics of decline are heterogenic and characterized by both, progressive worsening and sudden late-life events. Flies either become increasingly incapacitated by accumulating disability over multiple days prior to death, or they escape disability until few hours prior to death. Both late-life trajectories converge into a terminal stage characterized by stereotypical signs of functional collapse and death within 3 hours. Drosophila can now be used to evaluate life prolonging manipulations in the context of late-life quality. High sugar diet increases lifespan and late-life quality, whereas lifespan prolonging antioxidant supplementation has either no, or negative effects on late-life quality, depending on base diet and gender.
Subject(s)
Aging/physiology , Longevity , Physical Functional Performance , Animals , Drosophila melanogaster , Female , Longitudinal Studies , Male , Models, AnimalABSTRACT
In this issue of Neuron, Sweeney and Davis present a beautiful characterization of Drosophila mutants in a gene named spinster. The results indicate a function of the endocytic pathway in regulating transforming growth factor-beta (TGF-beta) signaling at the Drosophila motor synapse. This study provides important new information at an intersection of several disciplines, including membrane traffic, lipid organization, synaptic signaling, and neurodegenerative lysosomal storage disease.
Subject(s)
Drosophila Proteins/metabolism , Drosophila melanogaster/growth & development , Membrane Proteins/metabolism , Mutation/genetics , Neuromuscular Junction/growth & development , Synaptic Transmission/genetics , Transforming Growth Factor beta/metabolism , Animals , Drosophila Proteins/genetics , Drosophila melanogaster/metabolism , Endocytosis/genetics , Humans , Lysosomal Storage Diseases, Nervous System/genetics , Lysosomal Storage Diseases, Nervous System/metabolism , Lysosomal Storage Diseases, Nervous System/physiopathology , Membrane Proteins/genetics , Neuromuscular Junction/metabolism , Signal Transduction/genetics , Transforming Growth Factor beta/geneticsABSTRACT
Retrograde signals influence neuronal survival, differentiation, synaptogenesis, and plasticity. Several recent papers describe novel roles for the well-studied TGF-beta pathway in retrograde synaptic signaling. While each dissects spatial and molecular aspects of TGF-beta signaling in a specific synaptic context, together these studies demonstrate that a specific retrograde signal may be interpreted in diverse, neuron-specific ways. Thus, a neuron's intrinsic properties and its other extrinsic signaling inputs determine its cellular and genomic response to TGF-beta.
Subject(s)
Axonal Transport/physiology , Central Nervous System/metabolism , Motor Neurons/metabolism , Signal Transduction/physiology , Transforming Growth Factor beta/metabolism , Animals , Central Nervous System/cytology , Homeostasis/physiology , Humans , Motor Neurons/cytology , Neuromuscular Junction/metabolism , Synaptic Transmission/physiologyABSTRACT
Translational repression by Drosophila Pumilio (Pum) protein controls posterior patterning during embryonic development. Here, we show that Pum is an important mediator of synaptic growth and plasticity at the neuromuscular junction (NMJ). Pum is localized to the postsynaptic side of the NMJ in third instar larvae and is also expressed in larval neurons. Neuronal Pum regulates synaptic growth. In its absence, NMJ boutons are larger and fewer in number, while Pum overexpression increases bouton number and decreases bouton size. Postsynaptic Pum negatively regulates expression of the translation factor eIF-4E at the NMJ, and Pum binds selectively to the 3'UTR of eIF-4E mRNA. The GluRIIa glutamate receptor is upregulated in pum mutants. These results, together with genetic epistasis studies, suggest that postsynaptic Pum modulates synaptic function via direct control of eIF-4E expression.
Subject(s)
Drosophila Proteins/metabolism , Eukaryotic Initiation Factor-4E/metabolism , Neuromuscular Junction/embryology , Neuronal Plasticity/physiology , Synapses/metabolism , Animals , Base Sequence , Drosophila , Electrophoretic Mobility Shift Assay , Immunohistochemistry , Molecular Sequence Data , RNA-Binding Proteins , Receptors, AMPA/metabolismABSTRACT
Restless legs syndrome (RLS) is a complex disorder that involves sensory and motor systems. The major pathophysiology of RLS is low iron concentration in the substantia nigra containing the cell bodies of dopamine neurons that project to the striatum, an area that is crucial for modulating movement. People who have RLS often present with normal iron values outside the brain; recent studies implicate several genes are involved in the syndrome. Like most complex diseases, animal models usually do not faithfully capture the full phenotypic spectrum of "disease," which is a uniquely human construct. Nonetheless, animal models have proven useful in helping to unravel the complex pathophysiology of diseases such as RLS and suggesting novel treatment paradigms. For example, hypothesis-independent genome-wide association studies (GWAS) have identified several genes as increasing the risk for RLS, including BTBD9. Independently, the murine homolog Btbd9 was identified as a candidate gene for iron regulation in the midbrain in mice. The relevance of the phenotype of another of the GWAS identified genes, MEIS1, has also been explored. The role of Btbd9 in iron regulation and RLS-like behaviors has been further evaluated in mice carrying a null mutation of the gene and in fruit flies when the BTBD9 protein is degraded. The BTBD9 and MEIS1 stories originate from human GWAS research, supported by work in a genetic reference population of mice (forward genetics) and further verified in mice, fish flies, and worms. Finally, the role of genetics is further supported by an inbred mouse strain that displays many of the phenotypic characteristics of RLS. The role of animal models of RLS phenotypes is also extended to include periodic limb movements.
Subject(s)
Disease Models, Animal , Restless Legs Syndrome/genetics , Restless Legs Syndrome/physiopathology , Animals , Humans , PhenotypeABSTRACT
The dicistronic Drosophila stoned mRNA produces two proteins, stonedA and stonedB, that are localized at nerve terminals. While the stoned locus is required for synaptic-vesicle cycling in neurons, distinct or overlapping synaptic functions of stonedA and stonedB have not been clearly identified. Potential functions of stoned products in nonneuronal cells remain entirely unexplored in vivo. Transgene-based analyses presented here demonstrate that exclusively neuronal expression of a dicistronic stoned cDNA is sufficient for rescue of defects observed in lethal and viable stoned mutants. Significantly, expression of a monocistronic stonedB trangene is sufficient for rescuing various phenotypic deficits of stoned mutants, including those in organismal viability, evoked transmitter release, and synaptotagmin retrieval from the plasma membrane. In contrast, a stonedA transgene does not alleviate any stoned mutant phenotype. Novel phenotypic analyses demonstrate that, in addition to regulation of presynaptic function, stoned is required for regulating normal growth and morphology of the motor terminal; however, this developmental function is also provided by a stonedB transgene. Our data, although most consistent with a hypothesis in which stonedA is a dispensable protein, are limited by the absence of a true null allele for stoned due to partial restoration of presynaptic stonedA by transgenically provided stonedB. Careful analysis of the effects of the monocistronic transgenes together and in isolation clearly reveals that the presence of presynaptic stonedA is dependent on stonedB. Together, our findings improve understanding of the functional relationship between stonedA and stonedB and elaborate significantly on the in vivo functions of stonins, recently discovered phylogenetically conserved stonedB homologs that represent a new family of "orphan" medium (mu) chains of adaptor complexes involved in vesicle formation. Data presented here also provide new insight into potential mechanisms that underlie translation and evolution of the dicistronic stoned mRNA.