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O'Connor-Giles, K. M.

Publications and source records attributed to O'Connor-Giles, K. M..

10 recordsLinked to original sources

Selective regulation of transsynaptic alignment and postsynaptic assembly by a novel NCAM family synaptic adhesion molecule

Synapse formation underlies the organization of neurons into functional circuits during brain development and requires precise alignment and maturation of pre-and postsynaptic compartments. Many synaptogenic adhesion molecules have been identified that drive target recognition and sustained adhesion between appropriate synaptic partners. Yet the degree to which individual molecules serve selective functions in distinct aspects of synapse formation and maturation remains poorly understood. In particular, how exquisite nanoalignment of pre-and postsynaptic specializations flows from micron-scale adhesive interactions between synaptic partners remains a key unanswered question. Here we shed new light on this question by establishing a specialized set of synaptic functions for Epithelial limiter of Fasciclin II function (Elff), an NCAM family member with previously unknown roles in the nervous system. Our structural and functional studies at the glutamatergic Drosophila NMJ indicate that Elff is required for postsynaptic assembly, maturation, and transsynaptic alignment; however, it is not required for presynaptic function, bouton formation, or developmental expansion of the NMJ. Notably, NMJs in elff null mutants display reduced glutamate receptor clustering beginning at the embryonic stage when NMJ synapses first form. These poorly defined postsynaptic specializations are frequently out of register with presynaptic release sites, disrupting neurotransmission. Unexpectedly, the striking defects in elff nulls occur in the context of both normal active zone number and developmental expansion of the NMJ. These findings suggest a surprising degree of specialization among transsynaptic adhesion complexes and demonstrate that Elff-mediated signaling is critical for the transsynaptic nanoarchitecture of glutamatergic synapses.

neuroscience↗

Neurodegenerative and neurodevelopmental roles for bulk lipid transporters VPS13A and BLTP2 in movement disorders

BackgroundBridge-like lipid transfer proteins (BLTPs) mediate bulk lipid transport at membrane contact sites. Mutations in BLTPs are linked to both early-onset neurodevelopmental and later-onset neurodegenerative diseases, including movement disorders. The tissue specificity and temporal requirements of BLTPs in disease pathogenesis remain poorly understood. ObjectivesTo determine the age-of-onset and tissue-specific roles of VPS13A and BLTP2 in movement disorder pathogenesis using Drosophila models. MethodsWe generated tissue-specific knockdowns of the VPS13A ortholog (Vps13) and the BLTP2 ortholog (hobbit) in neurons and muscles of Drosophila. We analyzed age-dependent locomotor behavior, neurodegeneration, and synapse development and function. ResultsNeuron-specific loss of the VPS13A ortholog caused neurodegeneration followed by age- onset movement deficits and reduced lifespan, while muscle-specific loss affected only lifespan, revealing neurodegeneration and myopathy as independent comorbidities in VPS13A disease. In contrast, neuronal loss of the BLTP2 ortholog resulted in severe early-onset locomotor defects without neurodegeneration, while muscle loss impaired synaptogenesis and neurotransmission at the neuromuscular junction (NMJ). ConclusionsVPS13A maintains neuronal survival, while BLTP2 orchestrates synaptic development. VPS13A function in muscle does not play a role in movement defects. The phenotypic specificity of BLTP function provides mechanistic insights into distinct disease trajectories for BLTP-associated movement disorders.

neuroscience↗

TimeFlies: an snRNA-seq aging clock for the fruit fly head sheds light on sex-biased aging

Although multiple high-performing epigenetic aging clocks exist, few are based directly on gene expression. Such transcriptomic aging clocks allow us to identify potential age-associated genes directly. However, most existing transcriptomic clocks model a subset of genes and are limited in their ability to predict novel biomarkers. With the growing application of single-cell sequencing, there is a need for robust single-cell transcriptomic aging clocks. Moreover, aging clocks have yet to be applied to investigate the elusive phenomenon of sex differences in aging. We introduce TimeFlies, a pan-cell-type snRNA-seq aging clock for the Drosophila melanogaster head. TimeFlies uses deep learning to classify the donor age of cells based on genome-wide gene expression profiles. Using explainability methods, we identified key marker genes contributing to the classification, with lncRNAs showing up as highly enriched among predicted biomarkers. lncRNA:roX1 and lncRNA:roX2 are top clock genes across cell types. Both are regulators of X chromosome dosage compensation, a pathway previously found to be significantly affected by aging in the mouse brain. We validated these findings experimentally in Drosophila, showing a decrease in survival when dosage compensation is inhibited in vivo. Furthermore, we trained sex-specific TimeFlies clocks and noted significant differences in model predictions and explanations between male and female clocks, suggesting that different pathways drive aging in males and females.

genomics↗

Synaptic gene expression is coordinated across development by conserved chromatin regulators

Synapses are the primary sites of communication in the nervous system. Synapse formation requires the simultaneous expression of hundreds of proteins across cells. Through genomic and functional studies in Drosophila, we investigated the regulatory mechanisms underlying the spatiotemporal coordination of synaptic gene expression during neurodevelopment. We find that chromatin opening is coordinated across hundreds of synaptic genes immediately preceding transcription, whereas repression of these genes outside developmental periods of synaptogenesis is regulated by distinct mechanisms. We identify the conserved DEAF1 transcription factor (TF) as a direct repressor of synaptic genes. Disruption of DEAF1 or pioneer TF CLAMP, which promotes DEAF1 binding, increases synaptic gene expression across neuronal subtypes, leading to excess synapse formation. DEAF1, which is linked to syndromic intellectual disability, is also sufficient to block synaptogenesis. These findings reveal the importance of coordinated synaptic gene repression in sculpting neuronal connectivity and broaden our understanding of gene regulation in neurodevelopment.

neuroscience↗

tRNA modification enzyme-dependent redox homeostasis regulates synapse formation and memory

Post-transcriptional modification of RNA regulates gene expression at multiple levels. ALKBH8 is a tRNA modifying enzyme that methylates wobble uridines in specific tRNAs to modulate translation. Through methylation of tRNA-selenocysteine, ALKBH8 promotes selenoprotein synthesis and regulates redox homeostasis. Pathogenic variants in ALKBH8 have been linked to intellectual disability disorders in the human population, but the role of ALKBH8 in the nervous system is unknown. Through in vivo studies in Drosophila, we show that ALKBH8 controls oxidative stress in the brain to restrain synaptic growth and support learning and memory. ALKBH8 null animals lack wobble uridine methylation and exhibit a global reduction in protein synthesis, including a specific decrease in selenoprotein levels. Loss of ALKBH8 or independent disruption of selenoprotein synthesis results in ectopic synapse formation. Genetic expression of antioxidant enzymes fully suppresses synaptic overgrowth in ALKBH8 null animals, confirming oxidative stress as the underlying cause of dysregulation. ALKBH8 animals also exhibit associative learning and memory impairments that are reversed by pharmacological antioxidant treatment. Together, these findings demonstrate the critical role of tRNA modification in redox homeostasis in the nervous system and reveal antioxidants as a potential therapy for ALKBH8-associated intellectual disability. Significance StatementtRNA modifying enzymes are emerging as important regulators of nervous system development and function due to their growing links to neurological disorders. Yet, their roles in the nervous system remain largely elusive. Through in vivo studies in Drosophila, we link tRNA methyltransferase-regulated selenoprotein synthesis to synapse development and associative memory. These findings demonstrate the key role of tRNA modifiers in redox homeostasis during nervous system development and highlight the potential therapeutic benefit of antioxidant-based therapies for cognitive disorders linked to dysregulation of tRNA modification.

cell biology↗

PDZD8 promotes autophagy at ER-Lysosome contact sites to regulate synaptogenesis

Building synaptic connections, which are often far from the soma, requires coordinating a host of cellular activities from transcription to protein turnover, placing a high demand on intracellular communication. Membrane contact sites (MCSs) formed between cellular organelles have emerged as key signaling hubs for coordinating an array of cellular activities. We have found that the endoplasmic reticulum (ER) MCS tethering protein PDZD8 is required for activity-dependent synaptogenesis. PDZD8 is sufficient to drive ectopic synaptic bouton formation through an autophagy-dependent mechanism and required for basal synapse formation when autophagy biogenesis is limited. PDZD8 functions at ER-late endosome/lysosome (LEL) MCSs to promote lysosome maturation and accelerate autophagic flux. Mutational analysis of PDZD8s SMP domain further suggests a role for lipid transfer at ER-LEL MCSs. We propose that PDZD8-dependent lipid transfer from ER to LELs promotes lysosome maturation to increase autophagic flux during periods of high demand, including activity-dependent synapse formation. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=170 SRC="FIGDIR/small/564828v2_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@1c6b752org.highwire.dtl.DTLVardef@12d77c3org.highwire.dtl.DTLVardef@11d50corg.highwire.dtl.DTLVardef@ba404f_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

Molecular and organizational diversity intersect to generate functional synaptic heterogeneity within and between excitatory neuronal subtypes

Synaptic heterogeneity is a hallmark of nervous systems that enables complex and adaptable communication in neural circuits. To understand circuit function, it is thus critical to determine the factors that contribute to the functional diversity of synapses. We investigated the contributions of voltage-gated calcium channel (VGCC) abundance, spatial organization, and subunit composition to synapse diversity among and between synapses formed by two closely related Drosophila glutamatergic motor neurons with distinct neurotransmitter release probabilities (Pr). Surprisingly, VGCC levels are highly predictive of heterogeneous Pr among individual synapses of either low- or high-Pr inputs, but not between inputs. We find that the same number of VGCCs are more densely organized at high-Pr synapses, consistent with tighter VGCC-synaptic vesicle coupling. We generated endogenously tagged lines to investigate VGCC subunits in vivo and found that the 2{delta}-3 subunit Straightjacket along with the CAST/ELKS active zone (AZ) protein Bruchpilot, both key regulators of VGCCs, are less abundant at high-Pr inputs, yet positively correlate with Pr among synapses formed by either input. Consistently, both Straightjacket and Bruchpilot levels are dynamically increased across AZs of both inputs when neurotransmitter release is potentiated to maintain stable communication following glutamate receptor inhibition. Together, these findings suggest a model in which VGCC and AZ protein abundance intersects with input-specific spatial and molecular organization to shape the functional diversity of synapses.

neuroscience↗

Expanded tRNA methyltransferase family member TRMT9B regulates synaptic growth and function

Nervous system function relies on the formation and function of synaptic connections between neurons. Through a genetic screen in Drosophila for new conserved synaptic genes, we identified CG42261/Fid/ TRMT9B as a negative regulator of synaptogenesis. TRMT9B has been studied for its role as a tumor suppressor in multiple carcinomas and is one of two metazoan homologs of yeast tRNA methyltransferase 9 (Trm9), which methylates tRNA wobble uridines. Members of the expanded family of tRNA methyltransferases are increasingly being associated with neurological disorders and new biochemical functions. Interestingly, whereas Trm9 homolog ALKBH8/CG17807 is ubiquitously expressed, we find that TRMT9B is enriched in the nervous system, including at synapses. However, in the absence of animal models the role of TRMT9B in the nervous system has remained unknown. Here, we generated null alleles of TRMT9B and ALKBH8, and through liquid chromatography-mass spectrometry find that ALKBH8 is responsible for canonical tRNA wobble uridine methylation under basal conditions. In the nervous system, we find that TRMT9B negatively regulates synaptogenesis through a methyltransferase-dependent mechanism in agreement with our modeling studies. Finally, we find that neurotransmitter release is impaired in TRMT9B mutants. Our findings reveal a role for TRMT9B in regulating synapse formation and function, and highlight the importance of the expanded family of tRNA methyltransferases in the nervous system.

neuroscience↗

The calcium channel subunit α2δ-3 organizes synapses via a novel activity-dependent, autocrine BMP signaling pathway

Synapses are highly specialized for neurotransmitter signaling, yet activity-dependent growth factor release also plays critical roles at synapses. While efficient neurotransmitter signaling is known to rely on precise apposition of release sites and neurotransmitter receptors, molecular mechanisms enabling high-fidelity growth factor signaling within the synaptic microenvironment remain obscure. Here we show that the auxiliary calcium channel subunit 2{delta}-3 promotes the function of a novel activity-dependent autocrine BMP signaling pathway at the Drosophila NMJ. 2{delta} proteins have conserved synaptogenic activity, although how they execute this function has remained elusive. We find that 2{delta}-3 provides an extracellular scaffold for autocrine BMP signaling, suggesting a new mechanistic framework for understanding 2{delta}s conserved role in synapse organization. We further establish a transcriptional requirement for activity-dependent, autocrine BMP signaling in determining synapse density, structure, and function. We propose that activity-dependent, autocrine signals provide neurons with continuous feedback on their activity state and are thus well poised to modulate synapse structure and function.

neuroscience↗

Calcium channel levels at single synapses predict release probability and are upregulated in homeostatic potentiation

Neurons communicate through Ca2+-dependent neurotransmitter release at presynaptic active zones (AZs). Neurotransmitter release properties play a key role in defining information flow in circuits and are tuned during multiple forms of plasticity. Despite their central role in determining neurotransmitter release properties, little is known about how Ca2+ channel levels are modulated to calibrate synaptic function. We used CRISPR to tag the Drosophila CaV2 Ca2+ channel Cacophony (Cac) and investigated the regulation of endogenous Ca2+ channels during homeostatic plasticity in males in which all endogenous Cac channels are tagged. We found that heterogeneously distributed Cac is highly predictive of neurotransmitter release probability at individual AZs and differentially regulated during opposing forms of presynaptic homeostatic plasticity. Specifically, Cac levels at AZ are increased during chronic and acute presynaptic homeostatic potentiation (PHP), and live imaging during acute expression of PHP reveals proportional Ca2+ channel accumulation across heterogeneous AZs. In contrast, endogenous Cac levels do not change during presynaptic homeostatic depression (PHD), implying that the reported reduction in Ca2+ influx during PHD is achieved through functional adaptions to pre-existing Ca2+ channels. Thus, distinct mechanisms bi-directionally modulate presynaptic Ca2+ levels to maintain stable synaptic strength in response to diverse challenges, with Ca2+ channel abundance providing a rapidly tunable substrate for potentiating neurotransmitter release over both acute and chronic timescales.

neuroscience↗