SAK3 confers neuroprotection in the neurodegeneration model of X-linked Dystonia-Parkinsonism.

Background X-linked Dystonia-Parkinsonism(XDP) is an adult-onset neurodegenerative disorder that results in the loss of striatal medium spiny neurons (MSNs). XDP is associated with disease-specific mutations in and around the TAF1 gene. This study highlights the utility of directly reprogrammed MSNs from fibroblasts of affected XDP individuals as a platform that captures cellular and epigenetic phenotypes associated with XDP-related neurodegeneration. In addition, the current study demonstrates the neuroprotective effect of SAK3 currently tested in other neurodegenerative diseases. Methods XDP fibroblasts from three independent patients as well as age- and sex-matched control fibroblasts were used to generate MSNs by direct neuronal reprogramming using miRNA-9/9*-124 and thetranscription factors CTIP2 , DLX1 -P2A- DLX2 , and MYT1L . Neuronal death, DNA damage, and mitochondrial health assays were carried out to assess the neurodegenerative state of directly reprogrammed MSNs from XDP patients (XDP-MSNs). RNA sequencing and ATAC sequencing were performed to infer changes in the transcriptomic and chromatin landscapesof XDP-MSNs compared to those of control MSNs (Ctrl-MSNs). Results Our results show that XDP patient fibroblasts can be successfully reprogrammed into MSNs and XDP-MSNs display several degenerative phenotypes, including neuronal death, DNA damage, and mitochondrial dysfunction, compared to Ctrl-MSNs reprogrammed from age- and sex-matched control individuals' fibroblasts. In addition, XDP-MSNs showed increased vulnerability to TNFα -toxicity compared to Ctrl-MSNs. To dissect the altered cellular state in XDP-MSNs, we conducted transcriptomic and chromatin accessibility analyses using RNA- and ATAC-seq. Our results indicate that pathways related to neuronal function, calcium signaling, and genes related to other neurodegenerative diseases are commonly altered in XDP-MSNs from multiple patients. Interestingly, we found that SAK3, a T-type calcium channel activator, that may have therapeutic values in other neurodegenerative disorders, protected XDP-MSNs from neuronal death. Notably, we found that SAK3-mediated alleviation of neurodegeneration in XDP-MSNs was accompanied by gene expression changes toward Ctrl-MSNs.

Generation of Human Neurons by microRNA-Mediated Direct Conversion of Dermal Fibroblasts.

MicroRNAs (miRNAs), miR-9/9*, and miR-124 (miR-9/9*-124) display fate-reprogramming activities when ectopically expressed in human fibroblasts by erasing the fibroblast identity and evoking a pan-neuronal state. In contrast to induced pluripotent stem cell-derived neurons, miRNA-induced neurons (miNs) retain the biological age of the starting fibroblasts through direct fate conversion and thus provide a human neuron-based platform to study cellular properties inherent in aged neurons and model adult-onset neurodegenerative disorders using patient-derived cells. Furthermore, expression of neuronal subtype-specific transcription factors in conjunction with miR-9/9*-124 guides the miNs to distinct neuronal fates, a feature critical for modeling disorders that affect specific neuronal subtypes. Here, we describe the miR-9/9*-124-based neuronal reprogramming protocols for the generation of several disease-relevant neuronal subtypes: striatal medium spiny neurons, cortical neurons, and spinal cord motor neurons.

TRRAP is a central regulator of human multiciliated cell formation.

The multiciliated cell (MCC) is an evolutionarily conserved cell type, which in vertebrates functions to promote directional fluid flow across epithelial tissues. In the conducting airway, MCCs are generated by basal stem/progenitor cells and act in concert with secretory cells to perform mucociliary clearance to expel pathogens from the lung. Studies in multiple systems, including Xenopus laevis epidermis, murine trachea, and zebrafish kidney, have uncovered a transcriptional network that regulates multiple steps of multiciliogenesis, ultimately leading to an MCC with hundreds of motile cilia extended from their apical surface, which beat in a coordinated fashion. Here, we used a pool-based short hairpin RNA screening approach and identified TRRAP, an essential component of multiple histone acetyltransferase complexes, as a central regulator of MCC formation. Using a combination of immunofluorescence, signaling pathway modulation, and genomic approaches, we show that (a) TRRAP acts downstream of the Notch2-mediated basal progenitor cell fate decision and upstream of Multicilin to control MCC differentiation; and (b) TRRAP binds to the promoters and regulates the expression of a network of genes involved in MCC differentiation and function, including several genes associated with human ciliopathies.

The Differences in Local Translatome across Distinct Neuron Types Is Mediated by Both Baseline Cellular Differences and Post-transcriptional Mechanisms.

Local translation in neurites is a phenomenon that enhances the spatial segregation of proteins and their functions away from the cell body, yet it is unclear how local translation varies across neuronal cell types. Further, it is unclear whether differences in local translation across cell types simply reflect differences in transcription or whether there is also a cell type-specific post-transcriptional regulation of the location and translation of specific mRNAs. Most of the mRNAs discovered as being locally translated have been identified from hippocampal neurons because their laminar organization facilitates neurite-specific dissection and microscopy methods. Given the diversity of neurons across the brain, studies have not yet analyzed how locally translated mRNAs differ across cell types. Here, we used the SynapTRAP method to harvest two broad cell types in the mouse forebrain: GABAergic neurons and layer 5 projection neurons. While some transcripts overlap, the majority of the local translatome is not shared across these cell types. In addition to differences driven by baseline expression levels, some transcripts also exhibit cell type-specific post-transcriptional regulation. Finally, we provide evidence that GABAergic neurons specifically localize mRNAs for peptide neurotransmitters, including somatostatin and cortistatin, suggesting localized production of these key signaling molecules in the neurites of GABAergic neurons. Overall, this work suggests that differences in local translation in neurites across neuronal cell types are poised to contribute substantially to the heterogeneity in neuronal phenotypes.

A candidate regulatory variant at the TREM gene cluster associates with decreased Alzheimer's disease risk and increased TREML1 and TREM2 brain gene expression.

INTRODUCTION: We hypothesized that common Alzheimer's disease (AD)-associated variants within the triggering receptor expressed on myeloid (TREM) gene cluster influence disease through gene expression. METHODS: Expression microarrays on temporal cortex and cerebellum from ∼400 neuropathologically diagnosed subjects and two independent RNAseq replication cohorts were used for expression quantitative trait locus analysis. RESULTS: A variant within a DNase hypersensitive site 5' of TREM2, rs9357347-C, associates with reduced AD risk and increased TREML1 and TREM2 levels (uncorrected P = 6.3 × 10-3 and 4.6 × 10-2, respectively). Meta-analysis on expression quantitative trait locus results from three independent data sets (n = 1006) confirmed these associations (uncorrected P = 3.4 × 10-2 and 3.5 × 10-3, Bonferroni-corrected P = 6.7 × 10-2 and 7.1 × 10-3, respectively). DISCUSSION: Our findings point to rs9357347 as a functional regulatory variant that contributes to a protective effect observed at the TREM locus in the International Genomics of Alzheimer's Project genome-wide association study meta-analysis and suggest concomitant increase in TREML1 and TREM2 brain levels as a potential mechanism for protection from AD.

Failure to upregulate Agrp and Orexin in response to activity based anorexia in weight loss vulnerable rats characterized by passive stress coping and prenatal stress experience.

We hypothesize that anorexia nervosa (AN) poses a physiological stress. Therefore, the way an individual copes with stress may affect AN vulnerability. Since prenatal stress (PNS) exposure alters stress responsivity in offspring this may increase their risk of developing AN. We tested this hypothesis using the activity based anorexia (ABA) rat model in control and PNS rats that were characterized by either proactive or passive stress-coping behavior. We found that PNS passively coping rats ate less and lost more weight during the ABA paradigm. Exposure to ABA resulted in higher baseline corticosterone and lower insulin levels in all groups. However, leptin levels were only decreased in rats with a proactive stress-coping style. Similarly, ghrelin levels were increased only in proactively coping ABA rats. Neuropeptide Y (Npy) expression was increased and proopiomelanocortin (Pomc) expression was decreased in all rats exposed to ABA. In contrast, agouti-related peptide (Agrp) and orexin (Hctr) expression were increased in all but the PNS passively coping ABA rats. Furthermore, DNA methylation of the orexin gene was increased after ABA in proactive coping rats and not in passive coping rats. Overall our study suggests that passive PNS rats have innate impairments in leptin and ghrelin in responses to starvation combined with prenatal stress associated impairments in Agrp and orexin expression in response to starvation. These impairments may underlie decreased food intake and associated heightened body weight loss during ABA in the passively coping PNS rats.