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https://www.readbyqxmd.com/read/29130196/super-resolution-single-molecule-fish-at-the-drosophila-neuromuscular-junction
#1
Joshua S Titlow, Lu Yang, Richard M Parton, Ana Palanca, Ilan Davis
The lack of an effective, simple, and highly sensitive protocol for fluorescent in situ hybridization (FISH) at the Drosophila larval neuromuscular junction (NMJ) has hampered the study of mRNA biology. Here, we describe our modified single molecule FISH (smFISH) methods that work well in whole mount Drosophila NMJ preparations to quantify primary transcription and count individual cytoplasmic mRNA molecules in specimens while maintaining ultrastructural preservation. The smFISH method is suitable for high-throughput sample processing and 3D image acquisition using any conventional microscopy imaging modality and is compatible with the use of antibody colabeling and transgenic fluorescent protein tags in axons, glia, synapses, and muscle cells...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29114039/neuronal-activity-drives-fmrp-and-hspg-dependent-matrix-metalloproteinase-function-required-for-rapid-synaptogenesis
#2
Mary L Dear, Jarrod Shilts, Kendal Broadie
Matrix metalloproteinase (MMP) functions modulate synapse formation and activity-dependent plasticity. Aberrant MMP activity is implicated in fragile X syndrome (FXS), a disease caused by the loss of the RNA-binding protein FMRP and characterized by neurological dysfunction and intellectual disability. Gene expression studies in Drosophila suggest that Mmps cooperate with the heparan sulfate proteoglycan (HSPG) glypican co-receptor Dally-like protein (Dlp) to restrict trans-synaptic Wnt signaling and that synaptogenic defects in the fly model of FXS are alleviated by either inhibition of Mmp or genetic reduction of Dlp...
November 7, 2017: Science Signaling
https://www.readbyqxmd.com/read/29105522/orb2-as-modulator-of-brat-and-their-role-at-the-neuromuscular-junction
#3
Elena Santana, Sergio Casas-Tintó
How synapses are built and dismantled is a central question in neurobiology. A wide range of proteins and processes from gene transcription to protein degradation are involved. Orb2 regulates mRNA translation depending on its monomeric or oligomeric state to modulate nervous system development and memory. Orb2 is expressed in Drosophila larval brain and neuromuscular junction (NMJ), Orb2 knockdown causes a reduction of synapse number and defects in neuronal morphology. Brain tumor (Brat) is an Orb2 target; it is expressed in larval brain related with cell growth and proliferation...
November 6, 2017: Journal of Neurogenetics
https://www.readbyqxmd.com/read/29037990/roles-for-the-vcp-co-factors-npl4-and-ufd1-in-neuronal-function-in-drosophila-melanogaster
#4
Dwayne J Byrne, Mark J Harmon, Jeremy C Simpson, Craig Blackstone, Niamh C O'Sullivan
The VCP-Ufd1-Npl4 complex regulates proteasomal processing within cells by delivering ubiquitinated proteins to the proteasome for degradation. Mutations in VCP are associated with two neurodegenerative diseases, amyotrophic lateral sclerosis (ALS) and inclusion body myopathy with Paget's disease of the bone and frontotemporal dementia (IBMPFD), and extensive study has revealed crucial functions of VCP within neurons. By contrast, little is known about the functions of Npl4 or Ufd1 in vivo. Using neuronal-specific knockdown of Npl4 or Ufd1 in Drosophila melanogaster, we infer that Npl4 contributes to microtubule organization within developing motor neurons...
October 20, 2017: Journal of Genetics and Genomics, Yi Chuan Xue Bao
https://www.readbyqxmd.com/read/28994789/focal-macropatch-recordings-of-synaptic-currents-from-the-drosophila-larval-neuromuscular-junction
#5
Alexander Vasin, Maria Bykhovskaia
Drosophila neuromuscular junction (NMJ) is an excellent model system to study glutamatergic synaptic transmission. We describe the technique of focal macropatch recordings of synaptic currents from visualized boutons at the Drosophila larval NMJ. This technique requires customized fabrication of recording micropipettes, as well as a compound microscope equipped with a high magnification, long-distance water immersion objective, differential interference contrast (DIC) optics, and a fluorescent attachment. The recording electrode is positioned on the top of a selected synaptic bouton visualized with DIC optics, epi-fluorescence, or both...
September 25, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28978466/post-transcriptional-inhibition-of-hsc70-4-hspa8-expression-leads-to-synaptic-vesicle-cycling-defects-in-multiple-models-of-als
#6
Alyssa N Coyne, Ileana Lorenzini, Ching-Chieh Chou, Meaghan Torvund, Robert S Rogers, Alexander Starr, Benjamin L Zaepfel, Jennifer Levy, Jeffrey Johannesmeyer, Jacob C Schwartz, Hiroshi Nishimune, Konrad Zinsmaier, Wilfried Rossoll, Rita Sattler, Daniela C Zarnescu
Amyotrophic lateral sclerosis (ALS) is a synaptopathy accompanied by the presence of cytoplasmic aggregates containing TDP-43, an RNA-binding protein linked to ∼97% of ALS cases. Using a Drosophila model of ALS, we show that TDP-43 overexpression (OE) in motor neurons results in decreased expression of the Hsc70-4 chaperone at the neuromuscular junction (NMJ). Mechanistically, mutant TDP-43 sequesters hsc70-4 mRNA and impairs its translation. Expression of the Hsc70-4 ortholog, HSPA8, is also reduced in primary motor neurons and NMJs of mice expressing mutant TDP-43...
October 3, 2017: Cell Reports
https://www.readbyqxmd.com/read/28973139/homeostatic-plasticity-can-be-induced-and-expressed-to-restore-synaptic-strength-at-neuromuscular-junctions-undergoing-als-related-degeneration
#7
Sarah Perry, Yifu Han, Anushka Das, Dion Dickman
Amyotrophic lateral sclerosis (ALS) is debilitating neurodegenerative disease characterized by motor neuron dysfunction and progressive weakening of the neuromuscular junction (NMJ). Hereditary ALS is strongly associated with variants in the human C9orf72 gene. We have characterized C9orf72 pathology at the Drosophila NMJ and utilized several approaches to restore synaptic strength in this model. First, we demonstrate a dramatic reduction in synaptic arborization and active zone number at NMJs following C9orf72 transgenic expression in motor neurons...
November 1, 2017: Human Molecular Genetics
https://www.readbyqxmd.com/read/28912273/neuroligin-4-regulates-synaptic-growth-via-the-bone-morphogenetic-protein-bmp-signaling-pathway-at-the-drosophila-neuromuscular-junction
#8
Xinwang Zhang, Menglong Rui, Guangmin Gan, Cong Huang, Jukang Yi, Huihui Lv, Wei Xie
The neuroligin (Nlg) family of neural cell adhesion molecules is thought to be required for synapse formation and development and has been linked to the development of autism spectrum disorders in humans. In Drosophila melanogaster, mutations in the neuroligin 1-3 genes have been reported to induce synapse developmental defects at neuromuscular junctions (NMJs), but the role of neuroligin 4 (dnlg4) in synapse development has not been determined. Here, we report that the Drosophila neuroligin 4 (DNlg4) is different from DNlg1-3 in that it presynaptically regulates NMJ synapse development...
November 3, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/28860114/notum-coordinates-synapse-development-via-extracellular-regulation-of-wingless-trans-synaptic-signaling
#9
Danielle L Kopke, Sofia C Lima, Cyrille Alexandre, Kendal Broadie
Synaptogenesis requires orchestrated communication between pre- and postsynaptic cells via coordinated trans-synaptic signaling across the extracellular synaptomatrix. The first Wnt signaling ligand discovered, Drosophila Wingless (Wg; Wnt1 in mammals), plays crucial roles in synaptic development, regulating synapse architecture as well as functional differentiation. Here, we investigate synaptogenic functions of the secreted extracellular deacylase Notum, which restricts Wg signaling by cleaving an essential palmitoleate moiety...
October 1, 2017: Development
https://www.readbyqxmd.com/read/28846707/kek-6-a-truncated-trk-like-receptor-for-drosophila-neurotrophin-2-regulates-structural-synaptic-plasticity
#10
Suzana Ulian-Benitez, Simon Bishop, Istvan Foldi, Jill Wentzell, Chinenye Okenwa, Manuel G Forero, Bangfu Zhu, Marta Moreira, Mark Phizacklea, Graham McIlroy, Guiyi Li, Nicholas J Gay, Alicia Hidalgo
Neurotrophism, structural plasticity, learning and long-term memory in mammals critically depend on neurotrophins binding Trk receptors to activate tyrosine kinase (TyrK) signaling, but Drosophila lacks full-length Trks, raising the question of how these processes occur in the fly. Paradoxically, truncated Trk isoforms lacking the TyrK predominate in the adult human brain, but whether they have neuronal functions independently of full-length Trks is unknown. Drosophila has TyrK-less Trk-family receptors, encoded by the kekkon (kek) genes, suggesting that evolutionarily conserved functions for this receptor class may exist...
August 2017: PLoS Genetics
https://www.readbyqxmd.com/read/28790398/shedding-light-on-alzheimer-s-%C3%AE-amyloidosis-photosensitized-methylene-blue-inhibits-self-assembly-of-%C3%AE-amyloid-peptides-and-disintegrates-their-aggregates
#11
Byung Il Lee, Yoon Seok Suh, You Jung Chung, Kweon Yu, Chan Beum Park
Abnormal aggregation of β-amyloid (Aβ) peptides is a major hallmark of Alzheimer's disease (AD). In spite of numerous attempts to prevent the β-amyloidosis, no effective drugs for treating AD have been developed to date. Among many candidate chemicals, methylene blue (MB) has proved its therapeutic potential for AD in a number of in vitro and in vivo studies; but the result of recent clinical trials performed with MB and its derivative was negative. Here, with the aid of multiple photochemical analyses, we first report that photoexcited MB molecules can block Aβ42 aggregation in vitro...
August 8, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28745645/a-simple-neuronal-mechanical-injury-methodology-to-study-drosophila-motor-neuron-degeneration
#12
Erika B Danella, Lani C Keller
The degeneration of neurons occurs during normal development and in response to injury, stress, and disease. The cellular hallmarks of neuronal degeneration are remarkably similar in humans and invertebrates as are the molecular mechanisms that drive these processes. The fruit fly, Drosophila melanogaster, provides a powerful yet simple genetic model organism to study the cellular complexities of neurodegenerative diseases. In fact, approximately 70% of disease-associated human genes have a Drosophila homolog and a plethora of tools and assays have been described using flies to study human neurodegenerative diseases...
July 19, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28576972/secreted-tissue-inhibitor-of-matrix-metalloproteinase-restricts-trans-synaptic-signaling-to-coordinate-synaptogenesis
#13
Jarrod Shilts, Kendal Broadie
Synaptogenesis is coordinated by trans-synaptic signals that traverse the specialized synaptomatrix between presynaptic and postsynaptic cells. Matrix metalloproteinase (Mmp) activity sculpts this environment, balanced by secreted tissue inhibitors of Mmp (Timp). Here, we use the simplified Drosophila melanogaster matrix metalloproteome to test the consequences of eliminating all Timp regulatory control of Mmp activity at the neuromuscular junction (NMJ). Using in situ zymography, we find Timp limits Mmp activity at the NMJ terminal and shapes extracellular proteolytic dynamics surrounding individual synaptic boutons...
July 15, 2017: Journal of Cell Science
https://www.readbyqxmd.com/read/28544556/electrophysiological-analysis-of-synaptic-transmission-in-drosophila
#14
REVIEW
Maria Bykhovskaia, Alexander Vasin
Synaptic transmission is dynamic, plastic, and highly regulated. Drosophila is an advantageous model system for genetic and molecular studies of presynaptic and postsynaptic mechanisms and plasticity. Electrical recordings of synaptic responses represent a wide-spread approach to study neuronal signaling and synaptic transmission. We discuss experimental techniques that allow monitoring synaptic transmission in Drosophila neuromuscular and central systems. Recordings of synaptic potentials or currents at the larval neuromuscular junction (NMJ) are most common and provide numerous technical advantages due to robustness of the preparation, large and identifiable muscles, and synaptic boutons which can be readily visualized...
September 2017: Wiley Interdisciplinary Reviews. Developmental Biology
https://www.readbyqxmd.com/read/28520784/presynaptic-a%C3%AE-40-prevents-synapse-addition-in-the-adult-drosophila-neuromuscular-junction
#15
Begoña López-Arias, Enrique Turiégano, Ignacio Monedero, Inmaculada Canal, Laura Torroja
Complexity in the processing of the Amyloid Precursor Protein, which generates a mixture of βamyloid peptides, lies beneath the difficulty in understanding the etiology of Alzheimer's disease. Moreover, whether Aβ peptides have any physiological role in neurons is an unresolved question. By expressing single, defined Aβ peptides in Drosophila, specific effects can be discriminated in vivo. Here, we show that in the adult neuromuscular junction (NMJ), presynaptic expression of Aβ40 hinders the synaptic addition that normally occurs in adults, yielding NMJs with an invariable number of active zones at all ages tested...
2017: PloS One
https://www.readbyqxmd.com/read/28518121/two-algorithms-for-high-throughput-and-multi-parametric-quantification-of-drosophila-neuromuscular-junction-morphology
#16
Anna Castells-Nobau, Bonnie Nijhof, Ilse Eidhof, Louis Wolf, Jolanda M Scheffer-de Gooyert, Ignacio Monedero, Laura Torroja, Jeroen A W M van der Laak, Annette Schenck
Synaptic morphology is tightly related to synaptic efficacy, and in many cases morphological synapse defects ultimately lead to synaptic malfunction. The Drosophila larval neuromuscular junction (NMJ), a well-established model for glutamatergic synapses, has been extensively studied for decades. Identification of mutations causing NMJ morphological defects revealed a repertoire of genes that regulate synapse development and function. Many of these were identified in large-scale studies that focused on qualitative approaches to detect morphological abnormalities of the Drosophila NMJ...
May 3, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28500055/rnai-mediated-reverse-genetic-screen-identified-drosophila-chaperones-regulating-eye-and-neuromuscular-junction-morphology
#17
Sandeep Raut, Bhagaban Mallik, Arpan Parichha, Valsakumar Amrutha, Chandan Sahi, Vimlesh Kumar
Accumulation of toxic proteins in neurons has been linked with the onset of neurodegenerative diseases, which in many cases are characterized by altered neuronal function and synapse loss. Molecular chaperones help protein folding and the resolubilization of unfolded proteins, thereby reducing the protein aggregation stress. While most of the chaperones are expressed in neurons, their functional relevance remains largely unknown. Here, using bioinformatics analysis, we identified 95 Drosophila chaperones and classified them into seven different classes...
July 5, 2017: G3: Genes—Genomes—Genetics
https://www.readbyqxmd.com/read/28466185/tissue-specific-transcription-of-the-neuronal-gene-lim3-affects-drosophila-melanogaster-lifespan-and-locomotion
#18
Olga Y Rybina, Svetlana V Sarantseva, Ekaterina R Veselkina, Olga I Bolschakova, Alexander V Symonenko, Anna V Krementsova, Elena V Ryabova, Natalia V Roshina, Elena G Pasyukova
The identity of neuronal cell types is established and maintained by the expression of neuronal genes coding for ion channels, neurotransmitters, and neuropeptides, among others. Some of these genes have been shown to affect lifespan; however, their role in lifespan control remains largely unclear. The Drosophila melanogaster gene Lim3 encodes a transcription factor involved in complicated motor neuron specification networks. We previously identified Lim3 as a candidate gene affecting lifespan. To obtain direct evidence of the involvement of Lim3 in lifespan control, Lim3 overexpression and RNAi knockdown were induced in the nervous system and muscles of Drosophila using the GAL4-UAS binary system...
October 2017: Biogerontology
https://www.readbyqxmd.com/read/28455372/regulation-of-neuromuscular-junction-organization-by-rab2-and-its-effector-ica69-in-drosophila
#19
Bhagaban Mallik, Manish Kumar Dwivedi, Zeeshan Mushtaq, Manisha Kumari, Praveen Kumar Verma, Vimlesh Kumar
The mechanisms underlying synaptic differentiation, which involves neuronal membrane and cytoskeletal remodeling, are not completely understood. We performed a targeted RNAi-mediated screen of Drosophila BAR-domain proteins and identified islet cell autoantigen 69 kDa (ICA69) as one of the key regulators of morphological differentiation of the larval neuromuscular junction (NMJ). We show that Drosophila ICA69 colocalizes with α-Spectrin at the NMJ. The conserved N-BAR domain of ICA69 deforms liposomes in vitro Full-length ICA69 and the ICAC but not the N-BAR domain of ICA69 induce filopodia in cultured cells...
June 1, 2017: Development
https://www.readbyqxmd.com/read/28379367/a-drosophila-model-of-als-reveals-a-partial-loss-of-function-of-causative-human-pfn1-mutants
#20
Chi-Hong Wu, Anthony Giampetruzzi, Helene Tran, Claudia Fallini, Fen-Biao Gao, John E Landers
Mutations in the profilin 1 (PFN1) gene are causative for familial amyotrophic lateral sclerosis (fALS). However, it is still not fully understood how these mutations lead to neurodegeneration. To address this question, we generated a novel Drosophila model expressing human wild-type and ALS-causative PFN1 mutants. We show that at larval neuromuscular junctions (NMJ), motor neuron expression of wild-type human PFN1 increases the number of ghost boutons, active zone density, F-actin content, and the formation of filopodia...
June 1, 2017: Human Molecular Genetics
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