The continuously prolonged human lifespan is accompanied by increase in neurodegenerative diseases incidence, calling for the development of inexpensive blood-based diagnostics. Analyzing blood cell transcripts by RNA-Seq is a robust means to identify novel biomarkers that rapidly becomes a commonplace. However, there is lack of tools to discover novel exons, junctions and splicing events and to precisely and sensitively assess differential splicing through RNA-Seq data analysis and across RNA-Seq platforms. Here, we present a new and comprehensive computational workflow for whole-transcriptome RNA-Seq analysis, using an updated version of the software AltAnalyze, to identify both known and novel high-confidence alternative splicing events, and to integrate them with both protein-domains and microRNA binding annotations. We applied the novel workflow on RNA-Seq data from Parkinson's disease (PD) patients' leukocytes pre- and post- Deep Brain Stimulation (DBS) treatment and compared to healthy controls. Disease-mediated changes included decreased usage of alternative promoters and N-termini, 5′-end variations and mutually-exclusive exons. The PD regulated FUS and HNRNP A/B included prion-like domains regulated regions. We also present here a workflow to identify and analyze long non-coding RNAs (lncRNAs) via RNA-Seq data. We identified reduced lncRNA expression and selective PD-induced changes in 13 of over 6,000 detected leukocyte lncRNAs, four of which were inversely altered post-DBS. These included the U1 spliceosomal lncRNA and RP11-462G22.1, each entailing sequence complementarity to numerous microRNAs. Analysis of RNA-Seq from PD and unaffected controls brains revealed over 7,000 brain-expressed lncRNAs, of which 3,495 were co-expressed in the leukocytes including U1, which showed both leukocyte and brain increases. Furthermore, qRT-PCR validations confirmed these co-increases in PD leukocytes and two brain regions, the amygdala and substantia-nigra, compared to controls. This novel workflow allows deep multi-level inspection of RNA-Seq datasets and provides a comprehensive new resource for understanding disease transcriptome modifications in PD and other neurodegenerative diseases.

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Circular RNAs (circRNAs) are brain‐abundant RNAs of mostly unknown functions. To seek their roles in Parkinson's disease (PD), we generated an RNA sequencing resource of several brain region tissues from dozens of PD and control donors. In the healthy substantia nigra (SN), circRNAs accumulate in an age‐dependent manner, but in the PD SN this correlation is lost and the total number of circRNAs reduced. In contrast, the levels of circRNAs are increased in the other studied brain regions of PD patients. We also found circSLC8A1 to increase in the SN of PD individuals. CircSLC8A1 carries 7 binding sites for miR‐128 and is strongly bound to the microRNA effector protein Ago2. Indeed, RNA targets of miR‐128 are also increased in PD individuals, suggesting that circSLC8A1 regulates miR‐128 function and/or activity. CircSLC8A1 levels also increased in cultured cells exposed to the oxidative stress‐inducing agent paraquat but were decreased in cells treated with the neuroprotective antioxidant regulator drug Simvastatin. Together, our work links circSLC8A1 to oxidative stress‐related Parkinsonism and suggests further exploration of its molecular function in PD.

Recent reports attribute numerous regulatory functions to the nuclear paraspeckle‐forming long non‐coding RNA, nuclear enriched assembly transcript 1 (NEAT1), but the implications of its involvement in Parkinson's disease (PD) remain controversial. To address this issue, we assessed NEAT1 expression levels and cell type patterns in the substantia nigra (SN) from 53 donors with and without PD, as well as in interference tissue culture tests followed by multiple in‐house and web‐available models of PD. PCR quantification identified elevated levels of NEAT1 expression in the PD SN compared with control brains, an elevation that was reproducible across a multitude of disease models. In situ RNA hybridization supported neuron‐specific formation of NEATl‐based para‐speckles at the SN and demonstrated coincreases of NEAT1 and paraspeckles in cultured cells under paraquat (PQ)‐induced oxidative stress. Furthermore, neuroprotective agents, including fenofibrate and simvastatin, induced NEAT1 up‐regulation, whereas RNA interference—mediated depletion of NEAT1 exacerbated death of PQ‐exposed cells in a leucine‐rich repeat kinase 2‐mediated manner. Our findings highlight a novel protective role for NEAT1 in PD and suggest a previously unknown mechanism for the neuroprotective traits of widely used preventive therapeutics.—Simchovitz, A., Hanan, M., Niederhoffer, N., Madrer, N., Yayon, N., Bennett, E. R., Greenberg, D. S., Kadener, S., Soreq, H. NEAT1 is overexpressed in Parkinson's disease substantia nigra and confers drug‐inducible neuroprotection from oxidative stress. FASEB J. 33, 11223–11234 (2019). http://www.fasebj.org

Recent reports highlight regulatory functions of long noncoding RNAs (lncRNAs) in neurodegeneration and aging, but biomedical implications remain limited. Here, we report an rRNA‐depletion‐based long RNA‐Sequencing Resource of 65 substantia nigra, amygdala, and medial temporal gyrus samples from Parkinson's disease (PD) and matched control brains. Using a lncRNA‐focused analysis approach to identify functionally important transcripts, we discovered and prioritized many lncRNAs dysregulated in PD. Those included pronounced elevation of the P53‐induced noncoding transcript LINC‐PINT in the substantia nigra of PD patients, as well as in additional models of oxidative stress and PD. Intriguingly, we found that LINC‐PINT is a primarily neuronal transcript which showed conspicuous increases in maturing primary culture neurons. LINC‐PINT also accumulated in several brain regions of Alzheimer's and Huntington's disease patients and decreased with healthy brain aging, suggesting a general role in aging and neurodegeneration for this lncRNA. RNAi‐mediated depletion of LINC‐PINT exacerbated the death of cultured N2A and SH‐SY5Y cells exposed to oxidative stress, highlighting a previously undiscovered neuroprotective role for this tumor‐inducible lncRNA in the brains of patients with neurodegenerative disorders.