Proteomic Signaling of Dual-Specificity Phosphatase 4 (DUSP4) in Alzheimer's Disease.

DUSP4 is a member of the DUSP (dual-specificity phosphatase) subfamily that is selective to the mitogen-activated protein kinases (MAPK) and has been implicated in a range of biological processes and functions in Alzheimer's disease (AD). In this study, we utilized the stereotactic delivery of adeno-associated virus (AAV)-DUSP4 to overexpress DUSP4 in the dorsal hippocampus of 5xFAD and wildtype (WT) mice, then used mass spectrometry (MS)-based proteomics along with the label-free quantification to profile the proteome and phosphoproteome in the hippocampus. We identified protein expression and phosphorylation patterns modulated in 5xFAD mice and examined the sex-specific impact of DUSP4 overexpression on the 5xFAD proteome/phosphoproteome. In 5xFAD mice, a substantial number of proteins were up- or down-regulated in both male and female mice in comparison to age and sex-matched WT mice, many of which are involved in AD-related biological processes, such as activated immune response or suppressed synaptic activities. Many proteins in pathways, such as immune response were found to be suppressed in response to DUSP4 overexpression in male 5xFAD mice. In contrast, such a shift was absent in female mice. For the phosphoproteome, we detected an array of phosphorylation sites regulated in 5xFAD compared to WT and modulated via DUSP4 overexpression in each sex. Interestingly, 5xFAD- and DUSP4-associated phosphorylation changes occurred in opposite directions. Strikingly, both the 5xFAD- and DUSP4-associated phosphorylation changes were found to be mostly in neurons and play key roles in neuronal processes and synaptic functions. Site-centric pathway analysis revealed that both the 5xFAD- and DUSP4-associated phosphorylation sites were enriched for a number of kinase sets in females but only a limited number of sets of kinases in male mice. Taken together, our results suggest that male and female 5xFAD mice responded to DUSP4 overexpression via shared and sex-specific molecular mechanisms, which might underly similar reductions in amyloid pathology in both sexes while learning deficits were reduced in only females with DUSP4 overexpression. Finally, we validated our findings with the sex-specific AD-associated proteomes in human cohorts and further developed DUSP4-centric proteomic network models and signaling maps for each sex.

Proteomic signaling of dual specificity phosphatase 4 (DUSP4) in Alzheimer's disease.

DUSP4 is a member of the DUSP (Dual-Specificity Phosphatase) subfamily that is selective to the mitogen-activated protein kinases (MAPK) and has been implicated in a range of biological processes and functions in Alzheimer's disease (AD). In this study, we utilized stereotactic delivery of adeno-associated virus (AAV)-DUSP4 to overexpress DUSP4 in the dorsal hippocampus of 5xFAD and wildtype (WT) mice, then used mass spectrometry (MS)-based proteomics along with label-free quantification to profile the proteome and phosphoproteome in the hippocampus. We identified patterns of protein expression and phosphorylation that are modulated in 5xFAD mice and examined the sex-specific impact of DUSP4 overexpression on the 5xFAD proteome/phosphoproteome. In 5xFAD mice, a substantial number of proteins were up- or down-regulated in both male and female mice in comparison to age and sex-matched WT mice, many of which are involved in AD-related biological processes, such as the activated immune response or suppression of synaptic activities. Upon DUSP4 overexpression, significantly regulated proteins were found in pathways that were suppressed, such as the immune response, in male 5xFAD mice. In contrast, such a shift was absent in female mice. For the phosphoproteome, we detected an array of phosphorylation sites that are regulated in 5xFAD compared to WT, and are modulated by DUSP4 overexpression in each sex. Interestingly, the changes in 5xFAD- and DUSP4-associated phosphorylation occurred in opposite directions. Strikingly, both the 5xFAD- and DUSP4-associated phosphorylation changes were found for the most part in neurons, and play key roles in neuronal processes and synaptic function. Site-centric pathway analysis revealed that both the 5xFAD- and DUSP4-associated phosphorylation sites were enriched for a number of kinase sets in female, but only a limited number of sets of kinases in male mice. Taken together, our results suggest that male and female 5xFAD mice respond to DUSP4 overexpression via shared and sex-specific molecular mechanisms, which might underly similar reductions in amyloid pathology in both sexes, while learning deficits were reduced in only females with DUSP4 overexpression. Finally, we validated our findings with the sex-specific AD-associated proteomes in human cohorts and further developed DUSP4-centric proteomic network models and signaling maps for each sex.

Proteomic signaling of dual specificity phosphatase 4 (DUSP4) in Alzheimer's disease.

DUSP4 is a member of the DUSP (Dual-Specificity Phosphatase) subfamily that is selective to the mitogen-activated protein kinases (MAPK) and has been implicated in a range of biological processes and functions in Alzheimer's disease (AD). In this study, we utilized stereotactic delivery of adeno-associated virus (AAV)-DUSP4 to overexpress DUSP4 in the dorsal hippocampus of 5×FAD and wildtype (WT) mice, then used mass spectrometry (MS)-based proteomics along with label-free quantification to profile the proteome and phosphoproteome in the hippocampus. We identified patterns of protein expression and phosphorylation that are modulated in 5×FAD mice and examined the sex-specific impact of DUSP4 overexpression on the 5×FAD proteome/phosphoproteome. In 5×FAD mice, a substantial number of proteins were up- or down-regulated in both male and female mice in comparison to age and sex-matched WT mice, many of which are involved in AD-related biological processes, such as the activated immune response or suppression of synaptic activities. Upon DUSP4 overexpression, significantly regulated proteins were found in pathways that were suppressed, such as the immune response, in male 5×FAD mice. In contrast, such a shift was absent in female mice. For the phosphoproteome, we detected an array of phosphorylation sites that are regulated in 5×FAD compared to WT, and are modulated by DUSP4 overexpression in each sex. Interestingly, the changes in 5×FAD- and DUSP4-associated phosphorylation occurred in opposite directions. Strikingly, both the 5×FAD- and DUSP4-associated phosphorylation changes were found for the most part in neurons, and play key roles in neuronal processes and synaptic function. Site-centric pathway analysis revealed that both the 5×FAD- and DUSP4-associated phosphorylation sites were enriched for a number of kinase sets in female, but only a limited number of sets of kinases in male mice. Taken together, our results suggest that male and female 5×FAD mice respond to DUSP4 overexpression via shared and sex-specific molecular mechanisms, which might underly similar reductions in amyloid pathology in both sexes, while learning deficits were reduced in only females with DUSP4 overexpression. Finally, we validated our findings with the sex-specific AD-associated proteomes in human cohorts and further developed DUSP4-centric proteomic network models and signaling maps for each sex.

miR155, TREM2, INPP5D: Disease stage and cell type are essential considerations when targeting clinical interventions based on mouse models of Alzheimer's amyloidopathy.

Studies of microglial gene manipulation in mouse models of Alzheimer's disease (AD) amyloidopathy can cause unpredictable effects on various key endpoints, including amyloidosis, inflammation, neuritic dystrophy, neurodegeneration, and learning behavior. In this Correspondence, we discuss three examples, microRNA 155 (miR155), TREM2, and INPP5D, in which observed results have been difficult to reconcile with predicted results based on precedent, because these six key endpoints do not reliably track together. The pathogenesis of AD involves multiple cell types and complex events that may change with disease stage. We propose that cell-type targeting and timing of intervention are responsible for the sometimes impossibility of predicting whether any prospective therapeutic intervention should aim at increasing or decreasing the level or activity of a particular molecular target.

Molecular and long-term behavioral consequences of neonatal opioid exposure and withdrawal in mice.

Introduction: Infants exposed to opioids in utero are at high risk of exhibiting Neonatal Opioid Withdrawal Syndrome (NOWS), a combination of somatic withdrawal symptoms including high pitched crying, sleeplessness, irritability, gastrointestinal distress, and in the worst cases, seizures. The heterogeneity of in utero opioid exposure, particularly exposure to polypharmacy, makes it difficult to investigate the underlying molecular mechanisms that could inform early diagnosis and treatment of NOWS, and challenging to investigate consequences later in life. Methods: To address these issues, we developed a mouse model of NOWS that includes gestational and post-natal morphine exposure that encompasses the developmental equivalent of all three human trimesters and assessed both behavior and transcriptome alterations. Results: Opioid exposure throughout all three human equivalent trimesters delayed developmental milestones and produced acute withdrawal phenotypes in mice reminiscent of those observed in infants. We also uncovered different patterns of gene expression depending on the duration and timing of opioid exposure (3-trimesters, in utero only, or the last trimester equivalent only). Opioid exposure and subsequent withdrawal affected social behavior and sleep in adulthood in a sex-dependent manner but did not affect adult behaviors related to anxiety, depression, or opioid response. Discussion: Despite marked withdrawal and delays in development, long-term deficits in behaviors typically associated with substance use disorders were modest. Remarkably, transcriptomic analysis revealed an enrichment for genes with altered expression in published datasets for Autism Spectrum Disorders, which correlate well with the deficits in social affiliation seen in our model. The number of differentially expressed genes between the NOWS and saline groups varied markedly based on exposure protocol and sex, but common pathways included synapse development, the GABAergic and myelin systems, and mitochondrial function.

Microglial INPP5D limits plaque formation and glial reactivity in the PSAPP mouse model of Alzheimer's disease.

INTRODUCTION: The inositol polyphosphate-5-phosphatase D (INPP5D) gene encodes a dual-specificity phosphatase that can dephosphorylate both phospholipids and phosphoproteins. Single nucleotide polymorphisms in INPP5D impact risk for developing late onset sporadic Alzheimer's disease (LOAD). METHODS: To assess the consequences of inducible Inpp5d knockdown in microglia of APPKM670/671NL /PSEN1Δexon9 (PSAPP) mice, we injected 3-month-old Inpp5dfl/fl /Cx3cr1CreER/+ and PSAPP/Inpp5dfl/fl /Cx3cr1CreER/+ mice with either tamoxifen (TAM) or corn oil (CO) to induce recombination. RESULTS: At age 6 months, we found that the percent area of 6E10+ deposits and plaque-associated microglia in Inpp5d knockdown mice were increased compared to controls. Spatial transcriptomics identified a plaque-specific expression profile that was extensively altered by Inpp5d knockdown. DISCUSSION: These results demonstrate that conditional Inpp5d downregulation in the PSAPP mouse increases plaque burden and recruitment of microglia to plaques. Spatial transcriptomics highlighted an extended gene expression signature associated with plaques and identified CST7 (cystatin F) as a novel marker of plaques. HIGHLIGHTS: Inpp5d knockdown increases plaque burden and plaque-associated microglia number. Spatial transcriptomics identifies an expanded plaque-specific gene expression profile. Plaque-induced gene expression is altered by Inpp5d knockdown in microglia. Our plaque-associated gene signature overlaps with human Alzheimer's disease gene networks.

Dual-Specificity Protein Phosphatase 4 (DUSP4) Overexpression Improves Learning Behavior Selectively in Female 5xFAD Mice, and Reduces ß-Amyloid Load in Males and Females.

Recent multiscale network analyses of banked brains from subjects who died of late-onset sporadic Alzheimer's disease converged on VGF (non-acronymic) as a key hub or driver. Within this computational VGF network, we identified the dual-specificity protein phosphatase 4 (DUSP4) [also known as mitogen-activated protein kinase (MAPK) phosphatase 2] as an important node. Importantly, DUSP4 gene expression, like that of VGF, is downregulated in postmortem Alzheimer's disease (AD) brains. We investigated the roles that this VGF/DUSP4 network plays in the development of learning behavior impairment and neuropathology in the 5xFAD amyloidopathy mouse model. We found reductions in DUSP4 expression in the hippocampi of male AD subjects, correlating with increased CDR scores, and in 4-month-old female and 12-18-month-old male 5xFAD hippocampi. Adeno-associated virus (AAV5)-mediated overexpression of DUSP4 in 5xFAD mouse dorsal hippocampi (dHc) rescued impaired Barnes maze performance in females but not in males, while amyloid loads were reduced in both females and males. Bulk RNA sequencing of the dHc from 5-month-old mice overexpressing DUSP4, and Ingenuity Pathway and Enrichr analyses of differentially expressed genes (DEGs), revealed that DUSP4 reduced gene expression in female 5xFAD mice in neuroinflammatory, interferon-gamma (IFNγ), programmed cell death protein-ligand 1/programmed cell death protein 1 (PD-L1/PD-1), and extracellular signal-regulated kinase (ERK)/MAPK pathways, via which DUSP4 may modulate AD phenotype with gender-specificity.

Intranasal Peptide Therapeutics: A Promising Avenue for Overcoming the Challenges of Traditional CNS Drug Development.

The central nervous system (CNS) has, among all organ systems in the human body, the highest failure rate of traditional small-molecule drug development, ranging from 80-100% depending on the area of disease research. This has led to widespread abandonment by the pharmaceutical industry of research and development for CNS disorders, despite increased diagnoses of neurodegenerative disorders and the continued lack of adequate treatment options for brain injuries, stroke, neurodevelopmental disorders, and neuropsychiatric illness. However, new approaches, concurrent with the development of sophisticated bioinformatic and genomic tools, are being used to explore peptide-based therapeutics to manipulate endogenous pathways and targets, including "undruggable" intracellular protein-protein interactions (PPIs). The development of peptide-based therapeutics was previously rejected due to systemic off-target effects and poor bioavailability arising from traditional oral and systemic delivery methods. However, targeted nose-to-brain, or intranasal (IN), approaches have begun to emerge that allow CNS-specific delivery of therapeutics via the trigeminal and olfactory nerve pathways, laying the foundation for improved alternatives to systemic drug delivery. Here we review a dozen promising IN peptide therapeutics in preclinical and clinical development for neurodegenerative (Alzheimer's, Parkinson's), neuropsychiatric (depression, PTSD, schizophrenia), and neurodevelopmental disorders (autism), with insulin, NAP (davunetide), IGF-1, PACAP, NPY, oxytocin, and GLP-1 agonists prominent among them.

Microglial TYROBP/DAP12 in Alzheimer's disease: Transduction of physiological and pathological signals across TREM2.

TYROBP (also known as DAP12 or KARAP) is a transmembrane adaptor protein initially described as a receptor-activating subunit component of natural killer (NK) cells. TYROBP is expressed in numerous cell types, including peripheral blood monocytes, macrophages, dendritic cells, and osteoclasts, but a key point of recent interest is related to the critical role played by TYROBP in the function of many receptors expressed on the plasma membrane of microglia. TYROBP is the downstream adaptor and putative signaling partner for several receptors implicated in Alzheimer's disease (AD), including SIRP1ß, CD33, CR3, and TREM2. TYROBP has received much of its current notoriety because of its importance in brain homeostasis by signal transduction across those receptors. In this review, we provide an overview of evidence indicating that the biology of TYROBP extends beyond its interaction with these four ligand-binding ectodomain-intramembranous domain molecules. In addition to reviewing the structure and localization of TYROBP, we discuss our recent progress using mouse models of either cerebral amyloidosis or tauopathy that were engineered to be TYROBP-deficient or TYROBP-overexpressing. Remarkably, constitutively TYROBP-deficient mice provided a model of genetic resilience to either of the defining proteinopathies of AD. Learning behavior and synaptic electrophysiological function were preserved at normal physiological levels even in the face of robust cerebral amyloidosis (in APP/PSEN1;Tyrobp-/- mice) or tauopathy (in MAPTP301S;Tyrobp-/- mice). A fundamental underpinning of the functional synaptic dysfunction associated with each proteotype was an accumulation of complement C1q. TYROBP deficiency prevented C1q accumulation associated with either proteinopathy. Based on these data, we speculate that TYROBP plays a key role in the microglial sensome and the emergence of the disease-associated microglia (DAM) phenotype. TYROBP may also play a key role in the loss of markers of synaptic integrity (e.g., synaptophysin-like immunoreactivity) that has long been held to be the feature of human AD molecular neuropathology that most closely correlates with concurrent clinical cognitive function.

Striatal Cholinergic Dysregulation after Neonatal Decrease in X-Linked Dystonia Parkinsonism-Related TAF1 Isoforms.

BACKGROUND: X-linked dystonia parkinsonism is a generalized, progressive dystonia followed by parkinsonism with onset in adulthood and accompanied by striatal neurodegeneration. Causative mutations are located in a noncoding region of the TATA-box binding protein-associated factor 1 (TAF1) gene and result in aberrant splicing. There are 2 major TAF1 isoforms that may be decreased in symptomatic patients, including the ubiquitously expressed canonical cTAF1 and the neuronal-specific nTAF1. OBJECTIVE: The objective of this study was to determine the behavioral and transcriptomic effects of decreased cTAF1 and/or nTAF1 in vivo. METHODS: We generated adeno-associated viral (AAV) vectors encoding microRNAs targeting Taf1 in a splice-isoform selective manner. We performed intracerebroventricular viral injections in newborn mice and rats and intrastriatal infusions in 3-week-old rats. The effects of Taf1 knockdown were assayed at 4 months of age with evaluation of motor function, histology, and RNA sequencing of the striatum, followed by its validation. RESULTS: We report motor deficits in all cohorts, more pronounced in animals injected at P0, in which we also identified transcriptomic alterations in multiple neuronal pathways, including the cholinergic synapse. In both species, we show a reduced number of striatal cholinergic interneurons and their marker mRNAs after Taf1 knockdown in the newborn. CONCLUSION: This study provides novel information regarding the requirement for TAF1 in the postnatal maintenance of striatal cholinergic neurons, the dysfunction of which is involved in other inherited forms of dystonia. © 2021 International Parkinson and Movement Disorder Society.

Neuronal intranuclear inclusion disease: Polyglycine protein is the culprit.

In this issue of Neuron, Boivin et al. (2021) show that a polyglycine-expanded protein, uN2CpolyG, is translated from an expansion of GGC repeats in the 5' UTR of the NOTCH2NLC (Notch homolog 2 N-terminal-like C) gene, defining a new pathological mechanism for neuronal intranuclear inclusion diseases (NIID).

Striatal Dopamine Induced ERK Phosphorylation Is Altered in Mouse Models of Monogenic Dystonia.

BACKGROUND: Similar to some monogenic forms of dystonia, levodopa-induced dyskinesia is a hyperkinetic movement disorder with abnormal nigrostriatal dopaminergic neurotransmission. Molecularly, it is characterized by hyper-induction of phosphorylation of extracellular signal-related kinase in response to dopamine in medium spiny neurons of the direct pathway. OBJECTIVES: The objective of this study was to determine if mouse models of monogenic dystonia exhibit molecular features of levodopa-induced dyskinesia. METHODS: Western blotting and quantitative immunofluorescence was used to assay baseline and/or dopamine-induced levels of the phosphorylated kinase in the striatum in mouse models of DYT1, DYT6, and DYT25 expressing a reporter in dopamine D1 receptor-expressing projection neurons. Cyclic adenosine monophosphate (cAMP) immunoassay and adenylyl cyclase activity assays were also performed. RESULTS: In DYT1 and DYT6 models, blocking dopamine reuptake with cocaine leads to enhanced extracellular signal-related kinase phosphorylation in dorsomedial striatal medium spiny neurons in the direct pathway, which is abolished by pretreatment with the N-methyl-d-aspartate antagonist MK-801. Phosphorylation is decreased in a model of DYT25. Levels of basal and stimulated cAMP and adenylyl cyclase activity were normal in the DYT1 and DYT6 mice and decreased in the DYT25 mice. Oxotremorine induced increased abnormal movements in the DYT1 knock-in mice. CONCLUSIONS: The increased dopamine induction of extracellular signal-related kinase phosphorylation in 2 genetic types of dystonia, similar to what occurs in levodopa-induced dyskinesia, and its decrease in a third, suggests that abnormal signal transduction in response to dopamine in the postsynaptic nigrostriatal pathway might be a point of convergence for dystonia and other hyperkinetic movement disorders, potentially offering common therapeutic targets. © 2021 International Parkinson and Movement Disorder Society.

Reactive or transgenic increase in microglial TYROBP reveals a TREM2-independent TYROBP-APOE link in wild-type and Alzheimer's-related mice.

INTRODUCTION: Microglial TYROBP (DAP12) is a network hub and driver in sporadic late-onset Alzheimer's disease (AD). TYROBP is a cytoplasmic adaptor for TREM2 and other receptors, but little is known about its roles and actions in AD. Herein, we demonstrate that endogenous Tyrobp transcription is specifically increased in recruited microglia. METHODS: Using a novel transgenic mouse overexpressing TYROBP in microglia, we observed a decrease of the amyloid burden and an increase of TAU phosphorylation stoichiometry when crossed with APP/PSEN1 or MAPTP301S mice, respectively. Characterization of these mice revealed Tyrobp-related modulation of apolipoprotein E (Apoe) transcription. We also showed that Tyrobp and Apoe mRNAs were increased in Trem2-null microglia recruited around either amyloid beta deposits or a cortical stab injury. Conversely, microglial Apoe transcription was dramatically diminished when Tyrobp was absent. CONCLUSIONS: Our results provide evidence that TYROBP-APOE signaling does not require TREM2 and could be an initiating step in establishment of the disease-associated microglia (DAM) phenotype.

VGF-derived peptide TLQP-21 modulates microglial function through C3aR1 signaling pathways and reduces neuropathology in 5xFAD mice.

BACKGROUND: Multiomic studies by several groups in the NIH Accelerating Medicines Partnership for Alzheimer's Disease (AMP-AD) identified VGF as a major driver of Alzheimer's disease (AD), also finding that reduced VGF levels correlate with mean amyloid plaque density, Clinical Dementia Rating (CDR) and Braak scores. VGF-derived peptide TLQP-21 activates the complement C3a receptor-1 (C3aR1), predominantly expressed in the brain on microglia. However, it is unclear how mouse or human TLQP-21, which are not identical, modulate microglial function and/or AD progression. METHODS: We performed phagocytic/migration assays and RNA sequencing on BV2 microglial cells and primary microglia isolated from wild-type or C3aR1-null mice following treatment with TLQP-21 or C3a super agonist (C3aSA). Effects of intracerebroventricular TLQP-21 delivery were evaluated in 5xFAD mice, a mouse amyloidosis model of AD. Finally, the human HMC3 microglial cell line was treated with human TLQP-21 to determine whether specific peptide functions are conserved from mouse to human. RESULTS: We demonstrate that TLQP-21 increases motility and phagocytic capacity in murine BV2 microglial cells, and in primary wild-type but not in C3aR1-null murine microglia, which under basal conditions have impaired phagocytic function compared to wild-type. RNA sequencing of primary microglia revealed overlapping transcriptomic changes induced by treatment with TLQP-21 or C3a super agonist (C3aSA). There were no transcriptomic changes in C3aR1-null or wild-type microglia exposed to the mutant peptide TLQP-R21A, which does not activate C3aR1. Most of the C3aSA- and TLQP-21-induced differentially expressed genes were linked to cell migration and proliferation. Intracerebroventricular TLQP-21 administration for 28 days via implanted osmotic pump resulted in a reduction of amyloid plaques and associated dystrophic neurites and restored expression of subsets of Alzheimer-associated microglial genes. Finally, we found that human TLQP-21 activates human microglia in a fashion similar to activation of murine microglia by mouse TLQP-21. CONCLUSIONS: These data provide molecular and functional evidence suggesting that mouse and human TLQP-21 modulate microglial function, with potential implications for the progression of AD-related neuropathology.

Diverse Mechanisms Lead to Common Dysfunction of Striatal Cholinergic Interneurons in Distinct Genetic Mouse Models of Dystonia.

Clinical and experimental data indicate striatal cholinergic dysfunction in dystonia, a movement disorder typically resulting in twisted postures via abnormal muscle contraction. Three forms of isolated human dystonia result from mutations in the TOR1A (DYT1), THAP1 (DYT6), and GNAL (DYT25) genes. Experimental models carrying these mutations facilitate identification of possible shared cellular mechanisms. Recently, we reported elevated extracellular striatal acetylcholine by in vivo microdialysis and paradoxical excitation of cholinergic interneurons (ChIs) by dopamine D2 receptor (D2R) agonism using ex vivo slice electrophysiology in Dyt1ΔGAG/+ mice. The paradoxical excitation was caused by overactive muscarinic receptors (mAChRs), leading to a switch in D2R coupling from canonical Gi/o to noncanonical ß-arrestin signaling. We sought to determine whether these mechanisms in Dyt1ΔGAG/+ mice are shared with Thap1C54Y/+ knock-in and Gnal+/- knock-out dystonia models and to determine the impact of sex. We found Thap1C54Y/+ mice of both sexes have elevated extracellular striatal acetylcholine and D2R-induced paradoxical ChI excitation, which was reversed by mAChR inhibition. Elevated extracellular acetylcholine was absent in male and female Gnal+/- mice, but the paradoxical D2R-mediated ChI excitation was retained and only reversed by inhibition of adenosine A2ARs. The Gi/o-preferring D2R agonist failed to increase ChI excitability, suggesting a possible switch in coupling of D2Rs to ß-arrestin, as seen previously in a DYT1 model. These data show that, whereas elevated extracellular acetylcholine levels are not always detected across these genetic models of human dystonia, the D2R-mediated paradoxical excitation of ChIs is shared and is caused by altered function of distinct G-protein-coupled receptors.SIGNIFICANCE STATEMENT Dystonia is a common and often disabling movement disorder. The usual medical treatment of dystonia is pharmacotherapy with nonselective antagonists of muscarinic acetylcholine receptors, which have many undesirable side effects. Development of new therapeutics is a top priority for dystonia research. The current findings, considered in context with our previous investigations, establish a role for cholinergic dysfunction across three mouse models of human genetic dystonia: DYT1, DYT6, and DYT25. The commonality of cholinergic dysfunction in these models arising from diverse molecular etiologies points the way to new approaches for cholinergic modulation that may be broadly applicable in dystonia.

Integrative approach to sporadic Alzheimer's disease: deficiency of TYROBP in cerebral Aß amyloidosis mouse normalizes clinical phenotype and complement subnetwork molecular pathology without reducing Aß burden.

Integrative gene network approaches enable new avenues of exploration that implicate causal genes in sporadic late-onset Alzheimer's disease (LOAD) pathogenesis, thereby offering novel insights for drug-discovery programs. We previously constructed a probabilistic causal network model of sporadic LOAD and identified TYROBP/DAP12, encoding a microglial transmembrane signaling polypeptide and direct adapter of TREM2, as the most robust key driver gene in the network. Here, we show that absence of TYROBP/DAP12 in a mouse model of AD-type cerebral Aß amyloidosis (APPKM670/671NL/PSEN1Δexon9) recapitulates the expected network characteristics by normalizing the transcriptome of APP/PSEN1 mice and repressing the induction of genes involved in the switch from homeostatic microglia to disease-associated microglia (DAM), including Trem2, complement (C1qa, C1qb, C1qc, and Itgax), Clec7a and Cst7. Importantly, we show that constitutive absence of TYROBP/DAP12 in the amyloidosis mouse model prevented appearance of the electrophysiological and learning behavior alterations associated with the phenotype of APPKM670/671NL/PSEN1Δexon9 mice. Our results suggest that TYROBP/DAP12 could represent a novel therapeutic target to slow, arrest, or prevent the development of sporadic LOAD. These data establish that the network pathology observed in postmortem human LOAD brain can be faithfully recapitulated in the brain of a genetically manipulated mouse. These data also validate our multiscale gene networks by demonstrating how the networks intersect with the standard neuropathological features of LOAD.

Correction: Integrative approach to sporadic Alzheimer's disease: deficiency of TYROBP in cerebral Aß amyloidosis mouse normalizes clinical phenotype and complement subnetwork molecular pathology without reducing Aß burden.

This article was originally published under standard licence, but has now been made available under a CC BY 4.0 license. The PDF and HTML versions of the paper have been modified accordingly.

Efficient RNA interference-based knockdown of mutant torsinA reveals reversibility of PERK-eIF2α pathway dysregulation in DYT1 transgenic rats in vivo.

DYT1 dystonia is a neurological disease caused by a dominant mutation that results in the loss of a glutamic acid in the endoplasmic reticulum-resident protein torsinA. Currently, treatments are symptomatic and only provide partial relief. Multiple reports support the hypothesis that selectively reducing expression of mutant torsinA without affecting levels of the wild type protein should be beneficial. Published cell-based studies support this hypothesis. It is unclear, however, if phenotypes are reversible by targeting the molecular defect once established in vivo. Here, we generated adeno-associated virus encoding artificial microRNA targeting human mutant torsinA and delivered them to the striatum of symptomatic transgenic rats that express the full human TOR1A mutant gene. We achieved efficient suppression of human mutant torsinA expression in DYT1 transgenic rats, partly reversing its accumulation in the nuclear envelope. This intervention rescued PERK-eIF2α pathway dysregulation in striatal projection neurons but not behavioral abnormalities. Moreover, we found abnormal expression of components of dopaminergic neurotransmission in DYT1 rat striatum, which were not normalized by suppressing mutant torsinA expression. Our findings demonstrate the reversibility of translational dysregulation in DYT1 neurons and confirm the presence of abnormal dopaminergic neurotransmission in DYT1 dystonia.

Multiscale Analysis of Independent Alzheimer's Cohorts Finds Disruption of Molecular, Genetic, and Clinical Networks by Human Herpesvirus.

Investigators have long suspected that pathogenic microbes might contribute to the onset and progression of Alzheimer's disease (AD) although definitive evidence has not been presented. Whether such findings represent a causal contribution, or reflect opportunistic passengers of neurodegeneration, is also difficult to resolve. We constructed multiscale networks of the late-onset AD-associated virome, integrating genomic, transcriptomic, proteomic, and histopathological data across four brain regions from human post-mortem tissue. We observed increased human herpesvirus 6A (HHV-6A) and human herpesvirus 7 (HHV-7) from subjects with AD compared with controls. These results were replicated in two additional, independent and geographically dispersed cohorts. We observed regulatory relationships linking viral abundance and modulators of APP metabolism, including induction of APBB2, APPBP2, BIN1, BACE1, CLU, PICALM, and PSEN1 by HHV-6A. This study elucidates networks linking molecular, clinical, and neuropathological features with viral activity and is consistent with viral activity constituting a general feature of AD.

Integrated biology approach reveals molecular and pathological interactions among Alzheimer's Aß42, Tau, TREM2, and TYROBP in Drosophila models.

BACKGROUND: Cerebral amyloidosis, neuroinflammation, and tauopathy are key features of Alzheimer's disease (AD), but interactions among these features remain poorly understood. Our previous multiscale molecular network models of AD revealed TYROBP as a key driver of an immune- and microglia-specific network that was robustly associated with AD pathophysiology. Recent genetic studies of AD further identified pathogenic mutations in both TREM2 and TYROBP. METHODS: In this study, we systematically examined molecular and pathological interactions among Aß, tau, TREM2, and TYROBP by integrating signatures from transgenic Drosophila models of AD and transcriptome-wide gene co-expression networks from two human AD cohorts. RESULTS: Glial expression of TREM2/TYROBP exacerbated tau-mediated neurodegeneration and synergistically affected pathways underlying late-onset AD pathology, while neuronal Aß42 and glial TREM2/TYROBP synergistically altered expression of the genes in synaptic function and immune modules in AD. CONCLUSIONS: The comprehensive pathological and molecular data generated through this study strongly validate the causal role of TREM2/TYROBP in driving molecular networks in AD and AD-related phenotypes in flies.