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Epigenetic alterations are a fundamental pathological hallmark of Alzheimer’s disease (AD). Herein, we uncover the unknown G9a modulation pathways involved in AD, showing the upregulation of G9a and H3K9me2 in the brains of AD patients. Likewise, treatment with a G9a inhibitor in SAMP8 mice reversed the high levels of H3K9me2 and rescued the cognitive decline. Interestingly, a transcriptional profile analysis revealed induction of neuronal plasticity and a reduction of oxidative stress and neuroinflammation; the latter being also validated in cell cultures. Furthermore, an exploratory H3K9me2 ChIP-seq analysis demonstrated that during G9a inhibition treatment, the H3K9me2 mark is enriched at the promoter of genes associated with neural functions. Lastly, we showed in Caenorhabditis elegans (C. elegans) AD transgenic strains, similar epigenetic modifications and modulated pathways were altered with increased β-amyloid levels, which were reverted by the set-25 (in C. elegans is similar to the mammalian G9a protein) knockout, including the cognitive impairment. Therefore, our findings confirm that RNAi suppression of set-25 or pharmacological G9a inhibition promotes a positive outcome in AD, being a promising therapeutic strategy.

International audience; The phenotypic transformation of astrocytes in Alzheimer’s disease (AD) is still not well understood. Recentanalyses based on single-nucleus RNA sequencing of postmortem Alzheimer’s disease (AD) samples are limited bythe low number of sequenced astrocytes, small cohort sizes, and low number of differentially expressed genesdetected. To optimize the detection of astrocytic genes, we employed a novel strategy consisting of the localization of pre-determined astrocyte and neuronal gene clusters in publicly available whole-brain transcriptomes.Specifically, we used cortical transcriptomes from 766 individuals, including cognitively normal subjects(Controls), and people diagnosed with mild cognitive impairment (MCI) or dementia due to AD. Samples camefrom three independent cohorts organized by the Mount Sinai Hospital, the Mayo Clinic, and the Religious OrderStudy/Memory and Aging Project (ROSMAP). Astrocyte- and neuron-specific gene clusters were generated fromhuman brain cell-type specific RNAseq data using hierarchical clustering and cell-type enrichment scoring. Genesfrom each cluster were manually annotated according to cell-type specific functional Categories. Gene SetVariation Analysis (GSVA) and Principal Component Analysis (PCA) were used to establish changes in thesefunctional categories among clinical cohorts. We highlight three novel findings of the study. First, individualswith the same clinical diagnosis were molecularly heterogeneous. Particularly in the Mayo Clinic and ROSMAPcohorts, over 50% of Controls presented down-regulation of genes encoding synaptic proteins typical of AD,whereas 30% of patients diagnosed with dementia due to AD presented Control-like transcriptomic profiles.Second, down-regulation of neuronal genes related to synaptic proteins coincided, in astrocytes, with upregulation of genes related to perisynaptic astrocytic processes (PAP) and down-regulation of genes encoding endolysosomal and mitochondrial proteins. Third, down-regulation of astrocytic mitochondrial genes inverselycorrelated with the disease stages defined by Braak and CERAD scoring. Finally, we interpreted these changes asmaladaptive or adaptive from the point of view of astrocyte biology in a model of the phenotypical transformation of astrocytes in AD. The main prediction is that early malfunction of the astrocytic endolysosomal system, associated with progressive mitochondrial dysfunction, contribute to Alzheimer’s disease. If this prediction is correct, therapies preventing organelle dysfunction in astrocytes may be beneficial in preclinical and clinical AD.

BackgroundOur understanding of the impact of astrocytes in Alzheimer’s disease (AD) is hindered by the lack of astrocyte-specific omics data from patients diagnosed with dementia due to AD. Studies aiming to profile human AD astrocytes—including single-nucleus RNA sequencing—were limited by the low number of differentially expressed genes detected, and by the small size of cohorts. We improved on prior studies with a novel systems-biology-based approach.Methods Human astrocytic and neuronal gene clusters were generated from RNA sequencing data from isolated healthy human brain cells using a cell-type enrichment score and clustering. The cell-specific gene clusters were localized in 766 subjects from three AD whole-brain transcriptomes generated by the Mount Sinai Hospital, the Mayo Clinic, and the Religious Order Study/Memory and Aging Project (ROSMAP), which also contains subjects with mild cognitive impairment (MCI). Gene clusters were organized into functional categories and subcategories using manual curation. Functional changes among subject groups were determined by gene set variation analysis (GSVA) and principal component analysis (PCA).Results Hierarchical clustering of transcriptomic data revealed molecular heterogeneity in individuals with the same clinical diagnosis. Particularly in the Mayo Clinic and ROSMAP cohorts, over 50% of Controls presented massive down-regulation of genes encoding for synaptic proteins, as widely documented in AD, suggesting that these subjects might have been at a preclinical stage at the time of death. Conversely, approximately 30% of AD patients showed preservation of neuronal genes as if they were non demented subjects, suggesting that they were resilient to AD pathology (present, according to CERAD and Braak scoring), but developed dementia due to comorbidities. The astrocytic gene profiles in AD patients presenting down-regulation of neuronal genes were termed ‘AD astrocytes’. According to GSVA and PCA, AD astrocytes showed down-regulation of genes encoding for mitochondrial and endolysosomal proteins, and up-regulation of genes related to perisynaptic astrocytic processes (PAP), and survival and stress responses.Discussion Astrocytes undergo a profound transcriptional change in a remarkable percentage of Control, MCI and AD subjects, affecting organelles and astrocyte-neuron interactions. We argue that therapies preventing organelle dysfunction in astrocytes may protect neural circuits in preclinical and clinical AD.

Mfn2 is a mitochondrial fusion protein with bioenergetic functions implicated in the pathophysiology of neuronal and metabolic disorders. Understanding the bioenergetic mechanism of Mfn2 may aid in designing therapeutic approaches for these disorders. Here we show using endoplasmic reticulum (ER) or mitochondria‐targeted Mfn2 that Mfn2 stimulation of the mitochondrial metabolism requires its localization in the ER, which is independent of its fusion function. ER‐located Mfn2 interacts with mitochondrial Mfn1/2 to tether the ER and mitochondria together, allowing Ca2+ transfer from the ER to mitochondria to enhance mitochondrial bioenergetics. The physiological relevance of these findings is shown during neurite outgrowth, when there is an increase in Mfn2‐dependent ER‐mitochondria contact that is necessary for correct neuronal arbor growth. Reduced neuritic growth in Mfn2 KO neurons is recovered by the expression of ER‐targeted Mfn2 or an artificial ER‐mitochondria tether, indicating that manipulation of ER‐mitochondria contacts could be used to treat pathologic conditions involving Mfn2., The bioenergetic role of Mfn2 relies on its localization in the ER. Mfn2 localization to mitochondria is dispensable in the presence of mitochondrial Mfn1, which tethers ER and mitochondria, and facilitates Ca2+ transfer.

Altres ajuts: La Marató de TV3 (TV3-20141430) Astrocytic excitability relies on cytosolic calcium increases as a key mechanism, whereby astrocytes contribute to synaptic transmission and hence learning and memory. While it is a cornerstone of neurosciences that experiences are remembered, because transmitters activate gene expression in neurons, long-term adaptive astrocyte plasticity has not been described. Here, we investigated whether the transcription factor CREB mediates adaptive plasticity-like phenomena in astrocytes. We found that activation of CREB-dependent transcription reduced the calcium responses induced by ATP, noradrenaline, or endothelin-1. As to the mechanism, expression of VP16-CREB, a constitutively active CREB mutant, had no effect on basal cytosolic calcium levels, extracellular calcium entry, or calcium mobilization from lysosomal-related acidic stores. Rather, VP16-CREB upregulated sigma-1 receptor expression thereby increasing the release of calcium from the endoplasmic reticulum and its uptake by mitochondria. Sigma-1 receptor was also upregulated in vivo upon VP16-CREB expression in astrocytes. We conclude that CREB decreases astrocyte responsiveness by increasing calcium signalling at the endoplasmic reticulum-mitochondria interface, which might be an astrocyte-based form of long-term depression.