Cross-Regulation between TDP-43 and Paraspeckles Promotes Pluripotency-Differentiation Transition.

RNA-binding proteins (RBPs) and long non-coding RNAs (lncRNAs) are key regulators of gene expression, but their joint functions in coordinating cell fate decisions are poorly understood. Here we show that the expression and activity of the RBP TDP-43 and the long isoform of the lncRNA Neat1, the scaffold of the nuclear compartment "paraspeckles," are reciprocal in pluripotent and differentiated cells because of their cross-regulation. In pluripotent cells, TDP-43 represses the formation of paraspeckles by enhancing the polyadenylated short isoform of Neat1. TDP-43 also promotes pluripotency by regulating alternative polyadenylation of transcripts encoding pluripotency factors, including Sox2, which partially protects its 3' UTR from miR-21-mediated degradation. Conversely, paraspeckles sequester TDP-43 and other RBPs from mRNAs and promote exit from pluripotency and embryonic patterning in the mouse. We demonstrate that cross-regulation between TDP-43 and Neat1 is essential for their efficient regulation of a broad network of genes and, therefore, of pluripotency and differentiation.

Abl kinase-mediated FUS Tyr526 phosphorylation alters nucleocytoplasmic FUS localization in FTLD-FUS.

Nuclear to cytoplasmic mislocalization and aggregation of multiple RNA-binding proteins (RBPs), including FUS, are the main neuropathological features of the majority of cases of amyotrophic lateral sclerosis (ALS) and frontotemporal lobular degeneration (FTLD). In ALS-FUS, these aggregates arise from disease-associated mutations in FUS, whereas in FTLD-FUS, the cytoplasmic inclusions do not contain mutant FUS, suggesting different molecular mechanisms of FUS pathogenesis in FTLD that remain to be investigated. We have previously shown that phosphorylation of the C-terminal Tyr526 of FUS results in increased cytoplasmic retention of FUS due to impaired binding to the nuclear import receptor TNPO1. Inspired by the above notions, in the current study we developed a novel antibody against the C-terminally phosphorylated Tyr526 FUS (FUSp-Y526) that is specifically capable of recognizing phosphorylated cytoplasmic FUS, which is poorly recognized by other commercially available FUS antibodies. Using this FUSp-Y526 antibody, we demonstrated a FUS phosphorylation-specific effect on the cytoplasmic distribution of soluble and insoluble FUSp-Y526 in various cells and confirmed the involvement of the Src kinase family in Tyr526 FUS phosphorylation. In addition, we found that FUSp-Y526 expression pattern correlates with active pSrc/pAbl kinases in specific brain regions of mice, indicating preferential involvement of cAbl in the cytoplasmic mislocalization of FUSp-Y526 in cortical neurons. Finally, the pattern of immunoreactivity of active cAbl kinase and FUSp-Y526 revealed altered cytoplasmic distribution of FUSp-Y526 in cortical neurons of post-mortem frontal cortex tissue from FTLD patients compared with controls. The overlap of FUSp-Y526 and FUS signals was found preferentially in small diffuse inclusions and was absent in mature aggregates, suggesting possible involvement of FUSp-Y526 in the formation of early toxic FUS aggregates in the cytoplasm that are largely undetected by commercially available FUS antibodies. Given the overlapping patterns of cAbl activity and FUSp-Y526 distribution in cortical neurons, and cAbl induced sequestration of FUSp-Y526 into G3BP1 positive granules in stressed cells, we propose that cAbl kinase is actively involved in mediating cytoplasmic mislocalization and promoting toxic aggregation of wild-type FUS in the brains of FTLD patients, as a novel putative underlying mechanism of FTLD-FUS pathophysiology and progression.

Cytoplasmic TDP-43 is involved in cell fate during stress recovery.

Transactive response DNA binding protein 43 (TDP-43) is an RNA processing protein central to the pathogenesis of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Nuclear TDP-43 mislocalizes in patients to the cytoplasm, where it forms ubiquitin-positive inclusions in affected neurons and glia. Physiologically, cytoplasmic TDP-43 is associated with stress granules (SGs). Here, we explored TDP-43 cytoplasmic accumulation and stress granule formation following osmotic and oxidative stress. We show that sorbitol drives TDP-43 redistribution to the cytoplasm, while arsenite induces the recruitment of cytoplasmic TDP-43 to TIA-1 positive SGs. We demonstrate that inducing acute oxidative stress after TDP-43 cytoplasmic relocalization by osmotic shock induces poly (ADP-ribose) polymerase (PARP) cleavage, which triggers cellular toxicity. Recruitment of cytoplasmic TDP-43 to polyribosomes occurs in an SH-SY5Y cellular stress model and is observed in FTD brain lysate. Moreover, the processing body (P-body) marker DCP1a is detected in TDP-43 granules during recovery from stress. Overall, this study supports a central role for cytoplasmic TDP-43 in controlling protein translation in stressed cells.

SFPQ regulates the accumulation of RNA foci and dipeptide repeat proteins from the expanded repeat mutation in C9orf72.

The expanded GGGGCC repeat mutation in the C9orf72 gene is the most common genetic cause of the neurodegenerative diseases amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). The expansion is transcribed to sense and antisense RNA, which form RNA foci and bind cellular proteins. This mechanism of action is considered cytotoxic. Translation of the expanded RNA transcripts also leads to the accumulation of toxic dipeptide repeat proteins (DPRs). The RNA-binding protein splicing factor proline and glutamine rich (SFPQ), which is being increasingly associated with ALS and FTD pathology, binds to sense RNA foci. Here, we show that SFPQ plays an important role in the C9orf72 mutation. Overexpression of SFPQ resulted in higher numbers of both sense and antisense RNA foci and DPRs in transfected human embryonic kidney (HEK) cells. Conversely, reduced SPFQ levels resulted in lower numbers of RNA foci and DPRs in both transfected HEK cells and C9orf72 mutation-positive patient-derived fibroblasts and lymphoblasts. Therefore, we have revealed a role of SFPQ in regulating the C9orf72 mutation that has implications for understanding and developing novel therapeutic targets for ALS and FTD.This article has an associated First Person interview with the first author of the paper.

Interactome screening of C9orf72 dipeptide repeats reveals VCP sequestration and functional impairment by polyGA.

Repeat expansions in the C9orf72 gene are a common cause of amyotrophic lateral sclerosis and frontotemporal lobar degeneration, two devastating neurodegenerative disorders. One of the proposed mechanisms of GGGGCC repeat expansion is their translation into non-canonical dipeptide repeats, which can then accumulate as aggregates and contribute to these pathologies. There are five different dipeptide repeat proteins (polyGA, polyGR, polyPR, polyPA and polyGP), some of which are known to be neurotoxic. In the present study, we used BioID2 proximity labelling to identify the interactomes of all five dipeptide repeat proteins consisting of 125 repeats each. We identified 113 interacting partners for polyGR, 90 for polyGA, 106 for polyPR, 25 for polyPA and 27 for polyGP. Gene Ontology enrichment analysis of the proteomic data revealed that these target interaction partners are involved in a variety of functions, including protein translation, signal transduction pathways, protein catabolic processes, amide metabolic processes and RNA-binding. Using autopsy brain tissue from patients with C9orf72 expansion complemented with cell culture analysis, we evaluated the interactions between polyGA and valosin containing protein (VCP). Functional analysis of this interaction revealed sequestration of VCP with polyGA aggregates, altering levels of soluble valosin-containing protein. VCP also functions in autophagy processes, and consistent with this, we observed altered autophagy in cells expressing polyGA. We also observed altered co-localization of polyGA aggregates and p62 in cells depleted of VCP. All together, these data suggest that sequestration of VCP with polyGA aggregates contributes to the loss of VCP function, and consequently to alterations in autophagy processes in C9orf72 expansion disorders.

Karyopherin abnormalities in neurodegenerative proteinopathies.

Neurodegenerative proteinopathies are characterized by progressive cell loss that is preceded by the mislocalization and aberrant accumulation of proteins prone to aggregation. Despite their different physiological functions, disease-related proteins like tau, α-synuclein, TAR DNA binding protein-43, fused in sarcoma and mutant huntingtin, all share low complexity regions that can mediate their liquid-liquid phase transitions. The proteins' phase transitions can range from native monomers to soluble oligomers, liquid droplets and further to irreversible, often-mislocalized aggregates that characterize the stages and severity of neurodegenerative diseases. Recent advances into the underlying pathogenic mechanisms have associated mislocalization and aberrant accumulation of disease-related proteins with defective nucleocytoplasmic transport and its mediators called karyopherins. These studies identify karyopherin abnormalities in amyotrophic lateral sclerosis, frontotemporal dementia, Alzheimer's disease, and synucleinopathies including Parkinson's disease and dementia with Lewy bodies, that range from altered expression levels to the subcellular mislocalization and aggregation of karyopherin α and ß proteins. The reported findings reveal that in addition to their classical function in nuclear import and export, karyopherins can also act as chaperones by shielding aggregation-prone proteins against misfolding, accumulation and irreversible phase-transition into insoluble aggregates. Karyopherin abnormalities can, therefore, be both the cause and consequence of protein mislocalization and aggregate formation in degenerative proteinopathies. The resulting vicious feedback cycle of karyopherin pathology and proteinopathy identifies karyopherin abnormalities as a common denominator of onset and progression of neurodegenerative disease. Pharmacological targeting of karyopherins, already in clinical trials as therapeutic intervention targeting cancers such as glioblastoma and viral infections like COVID-19, may therefore represent a promising new avenue for disease-modifying treatments in neurodegenerative proteinopathies.

Functional diversity of small nucleolar RNAs.

Small nucleolar RNAs (snoRNAs) are short non-protein-coding RNAs with a long-recognized role in tuning ribosomal and spliceosomal function by guiding ribose methylation and pseudouridylation at targeted nucleotide residues of ribosomal and small nuclear RNAs, respectively. SnoRNAs are increasingly being implicated in regulation of new types of post-transcriptional processes, for example rRNA acetylation, modulation of splicing patterns, control of mRNA abundance and translational efficiency, or they themselves are processed to shorter stable RNA species that seem to be the principal or alternative bioactive isoform. Intriguingly, some display unusual cellular localization under exogenous stimuli, or tissue-specific distribution. Here, we discuss the new and unforeseen roles attributed to snoRNAs, focusing on the presumed mechanisms of action. Furthermore, we review the experimental approaches to study snoRNA function, including high resolution RNA:protein and RNA:RNA interaction mapping, techniques for analyzing modifications on targeted RNAs, and cellular and animal models used in snoRNA biology research.

Nuclear RNA foci from C9ORF72 expansion mutation form paraspeckle-like bodies.

The GGGGCC (G4C2) repeat expansion mutation in the C9ORF72 gene is the most common genetic cause of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). Transcription of the repeat and formation of nuclear RNA foci, which sequester specific RNA-binding proteins, is one of the possible pathological mechanisms. Here, we show that (G4C2) n repeat RNA predominantly associates with essential paraspeckle proteins SFPQ, NONO, RBM14, FUS and hnRNPH and colocalizes with known paraspeckle-associated RNA hLinc-p21. As formation of paraspeckles in motor neurons has been associated with early phases of ALS, we investigated the extent of similarity between paraspeckles and (G4C2) n RNA foci. Overexpression of (G4C2)72 RNA results in their increased number and colocalization with SFPQ-stained nuclear bodies. These paraspeckle-like (G4C2)72 RNA foci form independently of the known paraspeckle scaffold, the long non-coding RNA NEAT1 Moreover, the knockdown of SFPQ protein in C9ORF72 expansion mutation-positive fibroblasts significantly reduces the number of (G4C2) n RNA foci. In conclusion, (G4C2) n RNA foci have characteristics of paraspeckles, which suggests that both RNA foci and paraspeckles play roles in FTD and ALS, and implies approaches for regulation of their formation.

Intracellular Responses Triggered by Cold Atmospheric Plasma and Plasma-Activated Media in Cancer Cells.

Cold atmospheric plasma (CAP), an ionized gas operating at room temperature, has been increasingly studied with respect to its potential use in medicine, where its beneficial effects on tumor reduction in oncology have been demonstrated. This review discusses the cellular changes appearing in cell membranes, cytoplasm, various organelles, and DNA content upon cells' direct or indirect exposure to CAP or CAP-activated media/solutions (PAM), respectively. In addition, the CAP/PAM impact on the main cellular processes of proliferation, migration, protein degradation and various forms of cell death is addressed, especially in light of CAP use in the oncology field of plasma medicine.

Exon-specific U1 snRNAs improve ELP1 exon 20 definition and rescue ELP1 protein expression in a familial dysautonomia mouse model.

Familial dysautonomia (FD) is a rare genetic disease with no treatment, caused by an intronic point mutation (c.2204+6T>C) that negatively affects the definition of exon 20 in the elongator complex protein 1 gene (ELP1 also known as IKBKAP). This substitution modifies the 5' splice site and, in combination with regulatory splicing factors, induces different levels of exon 20 skipping, in various tissues. Here, we evaluated the therapeutic potential of a novel class of U1 snRNA molecules, exon-specific U1s (ExSpeU1s), in correcting ELP1 exon 20 recognition. Lentivirus-mediated expression of ELP1-ExSpeU1 in FD fibroblasts improved ELP1 splicing and protein levels. We next focused on a transgenic mouse model that recapitulates the same tissue-specific mis-splicing seen in FD patients. Intraperitoneal delivery of ELP1-ExSpeU1s-adeno-associated virus particles successfully increased the production of full-length human ELP1 transcript and protein. This splice-switching class of molecules is the first to specifically correct the ELP1 exon 20 splicing defect. Our data provide proof of principle of ExSpeU1s-adeno-associated virus particles as a novel therapeutic strategy for FD.

Structural Diversity of Sense and Antisense RNA Hexanucleotide Repeats Associated with ALS and FTLD.

The hexanucleotide expansion GGGGCC located in C9orf72 gene represents the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar dementia (FTLD). Since the discovery one of the non-exclusive mechanisms of expanded hexanucleotide G4C2 repeats involved in ALS and FTLD is RNA toxicity, which involves accumulation of pathological sense and antisense RNA transcripts. Formed RNA foci sequester RNA-binding proteins, causing their mislocalization and, thus, diminishing their biological function. Therefore, structures adopted by pathological RNA transcripts could have a key role in pathogenesis of ALS and FTLD. Utilizing NMR spectroscopy and complementary methods, we examined structures adopted by both guanine-rich sense and cytosine-rich antisense RNA oligonucleotides with four hexanucleotide repeats. While both oligonucleotides tend to form dimers and hairpins, the equilibrium of these structures differs with antisense oligonucleotide being more sensitive to changes in pH and sense oligonucleotide to temperature. In the presence of K+ ions, guanine-rich sense RNA oligonucleotide also adopts secondary structures called G-quadruplexes. Here, we also observed, for the first time, that antisense RNA oligonucleotide forms i-motifs under specific conditions. Moreover, simultaneous presence of sense and antisense RNA oligonucleotides promotes formation of heterodimer. Studied structural diversity of sense and antisense RNA transcripts not only further depicts the complex nature of neurodegenerative diseases but also reveals potential targets for drug design in treatment of ALS and FTLD.

C9orf72 poly GA RAN-translated protein plays a key role in amyotrophic lateral sclerosis via aggregation and toxicity.

An intronic GGGGCC (G4C2) hexanucleotide repeat expansion inC9orf72 is the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia (C9ALS/FTD). Repeat-associated non-AUG (RAN) translation of G4C2 RNA can result in five different dipeptide repeat proteins (DPR: poly GA, poly GP, poly GR, poly PA, and poly PR), which aggregate into neuronal cytoplasmic and nuclear inclusions in affected patients, however their contribution to disease pathogenesis remains controversial. We show that among the DPR proteins, expression of poly GA in a cell culture model activates programmed cell death and TDP-43 cleavage in a dose-dependent manner. Dual expression of poly GA together with other DPRs revealed that poly GP and poly PA are sequestered by poly GA, whereas poly GR and poly PR are rarely co-localised with poly GA. Dual expression of poly GA and poly PA ameliorated poly GA toxicity by inhibiting poly GA aggregation both in vitro and in vivo in the chick embryonic spinal cord. Expression of alternative codon-derived DPRs in chick embryonic spinal cord confirmed in vitro data, revealing that each of the dipeptides caused toxicity, with poly GA being the most toxic. Further, in vivo expression of G4C2 repeats of varying length caused apoptotic cell death, but failed to generate DPRs. Together, these data demonstrate that C9-related toxicity can be mediated by either RNA or DPRs. Moreover, our findings provide evidence that poly GA is a key mediator of cytotoxicity and that cross-talk between DPR proteins likely modifies their pathogenic status in C9ALS/FTD.

A feedback loop between dipeptide-repeat protein, TDP-43 and karyopherin-α mediates C9orf72-related neurodegeneration.

Accumulation and aggregation of TDP-43 is a major pathological hallmark of amyotrophic lateral sclerosis and frontotemporal dementia. TDP-43 inclusions also characterize patients with GGGGCC (G4C2) hexanucleotide repeat expansion in C9orf72 that causes the most common genetic form of amyotrophic lateral sclerosis and frontotemporal dementia (C9ALS/FTD). Functional studies in cell and animal models have identified pathogenic mechanisms including repeat-induced RNA toxicity and accumulation of G4C2-derived dipeptide-repeat proteins. The role of TDP-43 dysfunction in C9ALS/FTD, however, remains elusive. We found G4C2-derived dipeptide-repeat protein but not G4C2-RNA accumulation caused TDP-43 proteinopathy that triggered onset and progression of disease in Drosophila models of C9ALS/FTD. Timing and extent of TDP-43 dysfunction was dependent on levels and identity of dipeptide-repeat proteins produced, with poly-GR causing early and poly-GA/poly-GP causing late onset of disease. Accumulating cytosolic, but not insoluble aggregated TDP-43 caused karyopherin-α2/4 (KPNA2/4) pathology, increased levels of dipeptide-repeat proteins and enhanced G4C2-related toxicity. Comparable KPNA4 pathology was observed in both sporadic frontotemporal dementia and C9ALS/FTD patient brains characterized by its nuclear depletion and cytosolic accumulation, irrespective of TDP-43 or dipeptide-repeat protein aggregates. These findings identify a vicious feedback cycle for dipeptide-repeat protein-mediated TDP-43 and subsequent KPNA pathology, which becomes self-sufficient of the initiating trigger and causes C9-related neurodegeneration.

Nuclear trafficking in amyotrophic lateral sclerosis and frontotemporal lobar degeneration.

Amyotrophic lateral sclerosis and frontotemporal lobar degeneration are two ends of a phenotypic spectrum of disabling, relentlessly progressive and ultimately fatal diseases. A key characteristic of both conditions is the presence of TDP-43 (encoded by TARDBP) or FUS immunoreactive cytoplasmic inclusions in neuronal and glial cells. This cytoplasmic mislocalization of otherwise predominantly nuclear RNA binding proteins implies a perturbation of the nucleocytoplasmic shuttling as a possible event in the pathogenesis. Compromised nucleocytoplasmic shuttling has recently also been associated with a hexanucleotide repeat expansion mutation in C9orf72, which is the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal lobar degeneration, and leads to accumulation of cytoplasmic TDP-43 inclusions. Mutation in C9orf72 may disrupt nucleocytoplasmic shuttling on the level of C9ORF72 protein, the transcribed hexanucleotide repeat RNA, and/or dipeptide repeat proteins translated form the hexanucleotide repeat RNA. These defects of nucleocytoplasmic shuttling may therefore, constitute the common ground of the underlying disease mechanisms in different molecular subtypes of amyotrophic lateral sclerosis and frontotemporal lobar degeneration.

Phosphorylation of C-terminal tyrosine residue 526 in FUS impairs its nuclear import.

Aberrant cytoplasmic aggregation of FUS, which is caused by mutations primarily in the C-terminal nuclear localisation signal, is associated with 3% of cases of familial amyotrophic lateral sclerosis (ALS). FUS aggregates are also pathognomonic for 10% of all frontotemporal lobar degeneration (FTLD) cases; however, these cases are not associated with mutations in the gene encoding FUS. This suggests that there are differences in the mechanisms that drive inclusion formation of FUS in ALS and FTLD. Here, we show that the C-terminal tyrosine residue at position 526 of FUS is crucial for normal nuclear import. This tyrosine is subjected to phosphorylation, which reduces interaction with transportin 1 and might consequentially affect the transport of FUS into the nucleus. Furthermore, we show that this phosphorylation can occur through the activity of the Src family of kinases. Our study implicates phosphorylation as an additional mechanism by which nuclear transport of FUS might be regulated and potentially perturbed in ALS and FTLD.

Differential roles of the ubiquitin proteasome system and autophagy in the clearance of soluble and aggregated TDP-43 species.

TAR DNA-binding protein (TDP-43, also known as TARDBP) is the major pathological protein in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Large TDP-43 aggregates that are decorated with degradation adaptor proteins are seen in the cytoplasm of remaining neurons in ALS and FTD patients post mortem. TDP-43 accumulation and ALS-linked mutations within degradation pathways implicate failed TDP-43 clearance as a primary disease mechanism. Here, we report the differing roles of the ubiquitin proteasome system (UPS) and autophagy in the clearance of TDP-43. We have investigated the effects of inhibitors of the UPS and autophagy on the degradation, localisation and mobility of soluble and insoluble TDP-43. We find that soluble TDP-43 is degraded primarily by the UPS, whereas the clearance of aggregated TDP-43 requires autophagy. Cellular macroaggregates, which recapitulate many of the pathological features of the aggregates in patients, are reversible when both the UPS and autophagy are functional. Their clearance involves the autophagic removal of oligomeric TDP-43. We speculate that, in addition to an age-related decline in pathway activity, a second hit in either the UPS or the autophagy pathway drives the accumulation of TDP-43 in ALS and FTD. Therapies for clearing excess TDP-43 should therefore target a combination of these pathways.

The many faces of small nucleolar RNAs.

Small nucleolar RNAs (snoRNAs) are a class of evolutionally conserved non-coding RNAs traditionally associated with nucleotide modifications in other RNA species. Acting as guides pairing with ribosomal (rRNA) and small nuclear RNAs (snRNAs), snoRNAs direct partner enzymes to specific sites for uridine isomerization or ribose methylation, thereby influencing stability, folding and protein-interacting properties of target RNAs. In recent years, however, numerous non-canonical functions have also been ascribed to certain members of the snoRNA group, ranging from regulation of mRNA editing and/or alternative splicing to posttranscriptional gene silencing by a yet poorly understood pathway that may involve microRNA-like mechanisms. While some of these intriguing snoRNAs (the so-called orphan snoRNAs) have no sequence complementarity to rRNA or snRNA, others apparently display dual functionality, performing both traditional and newly elucidated functions. Here, we review the effects elicited by non-canonical snoRNA activities.

ALS mutant FUS disrupts nuclear localization and sequesters wild-type FUS within cytoplasmic stress granules.

Mutations in the gene encoding Fused in Sarcoma (FUS) cause amyotrophic lateral sclerosis (ALS), a fatal neurodegenerative disorder. FUS is a predominantly nuclear DNA- and RNA-binding protein that is involved in RNA processing. Large FUS-immunoreactive inclusions fill the perikaryon of surviving motor neurons of ALS patients carrying mutations at post-mortem. This sequestration of FUS is predicted to disrupt RNA processing and initiate neurodegeneration. Here, we demonstrate that C-terminal ALS mutations disrupt the nuclear localizing signal (NLS) of FUS resulting in cytoplasmic accumulation in transfected cells and patient fibroblasts. FUS mislocalization is rescued by the addition of the wild-type FUS NLS to mutant proteins. We also show that oxidative stress recruits mutant FUS to cytoplasmic stress granules where it is able to bind and sequester wild-type FUS. While FUS interacts with itself directly by protein-protein interaction, the recruitment of FUS to stress granules and interaction with PABP are RNA dependent. These findings support a two-hit hypothesis, whereby cytoplasmic mislocalization of FUS protein, followed by cellular stress, contributes to the formation of cytoplasmic aggregates that may sequester FUS, disrupt RNA processing and initiate motor neuron degeneration.

Analysis of alternative splicing associated with aging and neurodegeneration in the human brain.

Age is the most important risk factor for neurodegeneration; however, the effects of aging and neurodegeneration on gene expression in the human brain have most often been studied separately. Here, we analyzed changes in transcript levels and alternative splicing in the temporal cortex of individuals of different ages who were cognitively normal, affected by frontotemporal lobar degeneration (FTLD), or affected by Alzheimer's disease (AD). We identified age-related splicing changes in cognitively normal individuals and found that these were present also in 95% of individuals with FTLD or AD, independent of their age. These changes were consistent with increased polypyrimidine tract binding protein (PTB)-dependent splicing activity. We also identified disease-specific splicing changes that were present in individuals with FTLD or AD, but not in cognitively normal individuals. These changes were consistent with the decreased neuro-oncological ventral antigen (NOVA)-dependent splicing regulation, and the decreased nuclear abundance of NOVA proteins. As expected, a dramatic down-regulation of neuronal genes was associated with disease, whereas a modest down-regulation of glial and neuronal genes was associated with aging. Whereas our data indicated that the age-related splicing changes are regulated independently of transcript-level changes, these two regulatory mechanisms affected expression of genes with similar functions, including metabolism and DNA repair. In conclusion, the alternative splicing changes identified in this study provide a new link between aging and neurodegeneration.