RESUMO
To decipher the molecular bases governing seed germination, this study presents the pivotal role of the cap-binding complex (CBC), comprising CBP20 and CBP80, in modulating the inhibitory effects of abscisic acid (ABA) in barley. Using both single and double barley mutants in genes encoding the CBC, we revealed that the double mutant hvcbp20.ab/hvcbp80.b displays ABA insensitivity, in stark contrast to the hypersensitivity observed in single mutants during germination. Our comprehensive transcriptome and metabolome analysis not only identified significant alterations in gene expression and splicing patterns but also underscored the regulatory nexus among CBC, ABA, and brassinosteroid (BR) signaling pathways.
Assuntos
Ácido Abscísico , Regulação da Expressão Gênica de Plantas , Germinação , Hordeum , Proteínas de Plantas , Hordeum/genética , Hordeum/metabolismo , Hordeum/crescimento & desenvolvimento , Germinação/genética , Ácido Abscísico/metabolismo , Ácido Abscísico/farmacologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Splicing de RNA , Mutação , Transdução de Sinais , Transcriptoma , Perfilação da Expressão Gênica , Proteínas de Ligação ao Cap de RNA/metabolismo , Proteínas de Ligação ao Cap de RNA/genética , Sementes/crescimento & desenvolvimento , Sementes/genética , Sementes/metabolismoRESUMO
SignificanceOur results demonstrate the existence of early cellular pathways and network alterations in oligodendrocytes in the alpha-synucleinopathies Parkinson's disease and multiple system atrophy. They further reveal the involvement of an immune component triggered by alpha-synuclein protein, as well as a connection between (epi)genetic changes and immune reactivity in multiple system atrophy. The knowledge generated in this study could be used to devise novel therapeutic approaches to treat synucleinopathies.
Assuntos
Células-Tronco Pluripotentes Induzidas , Atrofia de Múltiplos Sistemas , Doença de Parkinson , Sinucleinopatias , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Atrofia de Múltiplos Sistemas/metabolismo , Oligodendroglia/metabolismo , Doença de Parkinson/genética , Doença de Parkinson/metabolismo , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismoRESUMO
The core abscisic acid (ABA) signaling pathway consists of receptors, phosphatases, kinases and transcription factors, among them ABA INSENSITIVE 5 (ABI5) and ABRE BINDING FACTORs/ABRE-BINDING PROTEINs (ABFs/AREBs), which belong to the BASIC LEUCINE ZIPPER (bZIP) family and control expression of stress-responsive genes. ABI5 is mostly active in seeds and prevents germination and post-germinative growth under unfavorable conditions. The activity of ABI5 is controlled at transcriptional and protein levels, depending on numerous regulators, including components of other phytohormonal pathways. ABFs/AREBs act redundantly in regulating genes that control physiological processes in response to stress during vegetative growth. In this review, we focus on recent reports regarding ABI5 and ABFs/AREBs functions during abiotic stress responses, which seem to be partially overlapping and not restricted to one developmental stage in Arabidopsis and other species. Moreover, we point out that ABI5 and ABFs/AREBs play a crucial role in the core ABA pathway's feedback regulation. In this review, we also discuss increased stress tolerance of transgenic plants overexpressing genes encoding ABA-dependent bZIPs. Taken together, we show that ABI5 and ABFs/AREBs are crucial ABA-dependent transcription factors regulating processes essential for plant adaptation to stress at different developmental stages.
Assuntos
Ácido Abscísico/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Desenvolvimento Vegetal , Plantas Geneticamente Modificadas/metabolismo , Estresse Fisiológico , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/genética , Fatores de Transcrição de Zíper de Leucina Básica/genética , Flores/genética , Flores/crescimento & desenvolvimento , Flores/metabolismo , Frutas/genética , Frutas/crescimento & desenvolvimento , Frutas/metabolismo , Regulação da Expressão Gênica de Plantas , Germinação , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Sementes/genética , Sementes/crescimento & desenvolvimento , Sementes/metabolismo , Transdução de SinaisRESUMO
Here, we examine the cellular changes triggered by tumor necrosis factor alpha (TNF-α) and different alpha-synuclein (αSYN) species in astrocytes derived from induced pluripotent stem cells. Human astrocytes treated with TNF-α display a strong reactive pro-inflammatory phenotype with upregulation of pro-inflammatory gene networks, activation of the nuclear factor κB (NF-κB) pathway, and release of pro-inflammatory cytokines, whereas those treated with high-molecular-weight αSYN fibrils acquire a reactive antigen (cross)-presenting phenotype with upregulation of major histocompatibility complex (MHC) genes and increased human leukocyte antigen (HLA) molecules at the cell surface. Surprisingly, the cell surface location of MHC proteins is abrogated by larger F110 fibrillar polymorphs, despite the upregulation of MHC genes. Interestingly, TNF-α and αSYN fibrils compete to drive the astrocyte immune reactive response. The astrocyte immune responses are accompanied by an impaired mitochondrial respiration, which is exacerbated in Parkinson's disease (PD) astrocytes. Our data provide evidence for astrocytic involvement in PD pathogenesis and reveal their complex immune reactive responses to exogenous stressors.
Assuntos
Astrócitos/imunologia , Mitocôndrias/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , alfa-Sinucleína/metabolismo , Trifosfato de Adenosina/metabolismo , Sequência de Aminoácidos , Apresentação de Antígeno , Astrócitos/metabolismo , Membrana Celular/metabolismo , Respiração Celular , Citocinas/metabolismo , Cadeias HLA-DRB1/química , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Mediadores da Inflamação/metabolismo , Peso Molecular , Doença de Parkinson/patologia , Peptídeos/química , Peptídeos/metabolismo , Fenótipo , Ubiquitina-Proteína Ligases/metabolismoRESUMO
ABA INSENSITIVE 5 (ABI5) is a basic leucine zipper (bZIP) transcription factor which acts in the abscisic acid (ABA) network and is activated in response to abiotic stresses. However, the precise role of barley (Hordeum vulgare) ABI5 in ABA signaling and its function under stress remains elusive. Here, we show that HvABI5 is involved in ABA-dependent regulation of barley response to drought stress. We identified barley TILLING mutants carrying different alleles in the HvABI5 gene and we studied in detail the physiological and molecular response to drought and ABA for one of them. The hvabi5.d mutant, carrying G1751A transition, was insensitive to ABA during seed germination, yet it showed the ability to store more water than its parent cv. "Sebastian" (WT) in response to drought stress. The drought-tolerant phenotype of hvabi5.d was associated with better membrane protection, higher flavonoid content, and faster stomatal closure in the mutant under stress compared to the WT. The microarray transcriptome analysis revealed up-regulation of genes associated with cell protection mechanisms in the mutant. Furthermore, HvABI5 target genes: HVA1 and HVA22 showed higher activity after drought, which may imply better adaptation of hvabi5.d to stress. On the other hand, chlorophyll content in hvabi5.d was lower than in WT, which was associated with decreased photosynthesis efficiency observed in the mutant after drought treatment. To verify that HvABI5 acts in the ABA-dependent manner we analyzed expression of selected genes related to ABA pathway in hvabi5.d and its WT parent after drought and ABA treatments. The expression of key genes involved in ABA metabolism and signaling differed in the mutant and the WT under stress. Drought-induced increase of expression of HvNCED1, HvBG8, HvSnRK2.1, and HvPP2C4 genes was 2-20 times higher in hvabi5.d compared to "Sebastian". We also observed a faster stomatal closure in hvabi5.d and much higher induction of HvNCED1 and HvSnRK2.1 genes after ABA treatment. Together, these findings demonstrate that HvABI5 plays a role in regulation of drought response in barley and suggest that HvABI5 might be engaged in the fine tuning of ABA signaling by a feedback regulation between biosynthetic and signaling events. In addition, they point to different mechanisms of HvABI5 action in regulating drought response and seed germination in barley.
RESUMO
The hippocampus is important for memory formation and is severely affected in the brain with Alzheimer disease (AD). Our understanding of early pathogenic processes occurring in hippocampi in AD is limited due to tissue unavailability. Here, we report a chemical approach to rapidly generate free-floating hippocampal spheroids (HSs), from human induced pluripotent stem cells. When used to model AD, both APP and atypical PS1 variant HSs displayed increased Aß42/Aß40 peptide ratios and decreased synaptic protein levels, which are common features of AD. However, the two variants differed in tau hyperphosphorylation, protein aggregation, and protein network alterations. NeuroD1-mediated gene therapy in HSs-derived progenitors resulted in modulation of expression of numerous genes, including those involved in synaptic transmission. Thus, HSs can be harnessed to unravel the mechanisms underlying early pathogenic changes in the hippocampi of AD patients, and provide a robust platform for the development of therapeutic strategies targeting early stage AD.
Assuntos
Doença de Alzheimer/patologia , Hipocampo/patologia , Células-Tronco Pluripotentes Induzidas/patologia , Esferoides Celulares/patologia , Doença de Alzheimer/genética , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Estudos de Casos e Controles , Terapia Genética , Humanos , Neurônios/patologia , Fenótipo , Presenilina-1/genética , Presenilina-1/metabolismo , Agregados Proteicos , Proteoma/metabolismo , Proteômica , Transcrição GênicaRESUMO
Strigolactones (SLs) are a group of plant hormones involved in many aspects of plant development and stress adaptation. Here, we investigated the drought response of a barley (Hordeum vulgare L.) mutant carrying a missense mutation in the gene encoding the SL-specific receptor HvD14. Our results clearly showed that hvd14.d mutant is hyper-sensitive to drought stress. This was illustrated by a lower leaf relative water content (RWC), impaired photosynthesis, disorganization of chloroplast structure, altered stomatal density and slower closure of stomata in response to drought in the mutant compared to the wild type parent cultivar Sebastian. Although the content of abscisic acid (ABA) and its derivatives remained unchanged in the mutant, significant differences in expression of genes related to ABA biosynthesis were observed. Moreover, hvd14.d was insensitive to ABA during seed germination. Analysis of Arabidopsis thaliana mutant atd14-1 also demonstrated that mutation in the SL receptor resulted in increased sensitivity to drought. Our results indicate that the drought-sensitive phenotype of barley SL mutant might be caused by a disturbed ABA metabolism and/or signalling pathways. These results together uncovered a link between SL signalling and ABA-dependent drought stress response in barley.
Assuntos
Ácido Abscísico/metabolismo , Arabidopsis/fisiologia , Compostos Heterocíclicos com 3 Anéis/metabolismo , Hordeum/fisiologia , Lactonas/metabolismo , Proteínas de Plantas/genética , Ácido Abscísico/farmacologia , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Desidratação , Secas , Regulação da Expressão Gênica de Plantas , Germinação/efeitos dos fármacos , Germinação/fisiologia , Hordeum/efeitos dos fármacos , Mutação , Complexo de Proteína do Fotossistema II/metabolismo , Proteínas de Plantas/metabolismo , Estômatos de Plantas/fisiologia , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Sementes/efeitos dos fármacos , Sementes/fisiologia , Transdução de Sinais/genéticaRESUMO
Here, we describe the generation of an induced pluripotent stem cell (iPSC) line, from a male patient diagnosed with Parkinson's disease (PD). The patient carries a heterozygous variation p.A53T in the SNCA gene. Skin fibroblasts were reprogrammed using the non-integrating Sendai virus technology to deliver OCT3/4, SOX2, c-MYC and KLF4 factors. The generated iPSC line (CSC-32) preserved the mutation, displayed expression of common pluripotency markers, differentiated into derivatives of the three germ layers, and exhibited a normal karyotype. The clone CSC-32B is presented thereafter; it can be used to study the mechanisms underlying PD pathogenesis.
Assuntos
Células-Tronco Pluripotentes Induzidas/metabolismo , Doença de Parkinson/genética , alfa-Sinucleína/genética , Diferenciação Celular , Células Cultivadas , Heterozigoto , Humanos , Fator 4 Semelhante a Kruppel , Masculino , Pessoa de Meia-IdadeRESUMO
Variations in the POLG1 gene encoding the catalytic subunit of the mitochondrial DNA polymerase gamma, have recently been associated with Parkinson's disease (PD), especially in patients diagnosed with progressive external ophthalmoplegia (PEO). However, the majority of the studies reporting this association mainly focused on the genetic identification of the variation in POLG1 in PD patient primary cells, and determination of mitochondrial DNA copy number, providing little information about the cellular alterations existing in patient brain cells, in particular dopaminergic neurons. Therefore, through the use of induced pluripotent stem cells (iPSCs), we assessed cellular alterations in novel p.Q811R POLG1 (POLG1Q811R) variant midbrain dopaminergic neuron-containing spheroids (MDNS) from a female patient who developed early-onset PD, and compared them to cultures derived from a healthy control of the same gender. Both POLG1 variant and control MDNS contained functional midbrain regionalized TH/FOXA2-positive dopaminergic neurons, capable of releasing dopamine. Western blot analysis identified the presence of high molecular weight oligomeric alpha-synuclein in POLG1Q811R MDNS compared to control cultures. In order to assess POLG1Q811R-related cellular alterations within the MDNS, we applied mass-spectrometry based quantitative proteomic analysis. In total, 6749 proteins were identified, with 61 significantly differentially expressed between POLG1Q811R and control samples. Pro- and anti-inflammatory signaling and pathways involved in energy metabolism were altered. Notably, increased glycolysis in POLG1Q811R MDNS was suggested by the increase in PFKM and LDHA levels and confirmed using functional analysis of glycolytic rate and oxygen consumption levels. Our results validate the use of iPSCs to assess cellular alterations in relation to PD pathogenesis, in a unique PD patient carrying a novel p.Q811R variation in POLG1, and identify several altered pathways that may be relevant to PD pathogenesis.
Assuntos
DNA Polimerase gama/genética , Variação Genética/genética , Oftalmoplegia Externa Progressiva Crônica/genética , Transtornos Parkinsonianos/genética , Células-Tronco Pluripotentes/fisiologia , Esferoides Celulares/fisiologia , Adulto , Feminino , Humanos , Mesencéfalo/patologia , Mesencéfalo/fisiologia , Oftalmoplegia Externa Progressiva Crônica/complicações , Oftalmoplegia Externa Progressiva Crônica/diagnóstico , Transtornos Parkinsonianos/complicações , Transtornos Parkinsonianos/diagnóstico , Células-Tronco Pluripotentes/patologia , Proteômica/métodos , Esferoides Celulares/patologiaRESUMO
Mutations in the glucocerebrosidase (GBA) gene have been associated with the development of Parkinson's disease (PD). An induced pluripotent stem cell (iPSC) line was generated from a 60-year old patient diagnosed with PD and carrying a new mutation variant p.R301C in GBA. Using non-integrating Sendai virus-based technology, we utilized OCT3/4, SOX2, c-MYC and KLF4 transcription factors to reprogram skin fibroblasts into iPSCs. The generated iPSC line retained the mutation, displayed expression of common pluripotency markers, differentiated into the three germ layers, and exhibited normal karyotype. The iPSC line can be further used for studying PD pathogenesis.
Assuntos
Técnicas de Cultura de Células/métodos , Glucosilceramidase/genética , Células-Tronco Pluripotentes Induzidas/patologia , Mutação/genética , Doença de Parkinson/genética , Doença de Parkinson/patologia , Animais , Linhagem Celular , Humanos , Fator 4 Semelhante a Kruppel , Masculino , Camundongos , Pessoa de Meia-IdadeRESUMO
Balanced photosynthesis under drought is essential for better survival and for agricultural benefits in terms of biomass and yield. Given the current attempts to improve the photosynthetic efficiency for greater crop yield, the explanation of the genetic basis of that process, together with the phenotypic analysis, is significant in terms of both basic studies and potential agricultural application. Therefore, the main objective of this study was to uncover the molecular basis of the photosynthesis process under drought stress in barley. To address that goal, we conducted transcriptomic examination together with detailed photosynthesis analysis using the JIP-test. Using this approach, we indicated that photosynthesis is a process that is very early affected in barley seedlings treated with severe drought stress. Rather than focusing on individual genes, our strategy was pointed to the identification of groups of genes with similar expression patterns. As such, we identified and annotated almost 150 barley genes as crucial core-components of photosystems, electron transport components, and Calvin cycle enzymes. Moreover, we designated 17 possible regulatory interactions between photosynthesis-related genes and transcription factors in barley. Summarizing, our results provide a list of candidate genes for future genetic research and improvement of barley drought tolerance by targeting photosynthesis.
Assuntos
Secas , Hordeum/genética , Hordeum/fisiologia , Fotossíntese/genética , Estresse Fisiológico/genética , Ácido Abscísico/farmacologia , Clorofila A/metabolismo , Desidratação , Transporte de Elétrons/efeitos dos fármacos , Fluorescência , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Genes de Plantas , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Hordeum/efeitos dos fármacos , Oxigênio/metabolismo , Fotossíntese/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Plântula/efeitos dos fármacos , Plântula/fisiologia , Estresse Fisiológico/efeitos dos fármacos , Fatores de Transcrição/metabolismo , Transcriptoma/genéticaRESUMO
The leucine-rich repeat kinase 2 (LRRK2) p.G2019S mutation is the most common genetic cause of Parkinson's disease (PD). An induced pluripotent stem cell (iPSC) line CSC-41 was generated from a 75-year old patient diagnosed with PD caused by a p.G2019S mutation in LRRK2. Skin fibroblasts were reprogrammed using a non-integrating Sendai virus-based technology to deliver OCT3/4, SOX2, c-MYC and KLF4 transcription factors. The generated iPSC line exhibits expression of common pluripotency markers, differentiates into the three germ layers and has a normal karyotype. The iPSC line can be used to explore the association between LRRK2 mutation and PD.
Assuntos
Técnicas de Cultura de Células/métodos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Mutação/genética , Doença de Parkinson/genética , Doença de Parkinson/patologia , Idoso , Animais , Linhagem Celular , Feminino , Humanos , Células-Tronco Pluripotentes Induzidas , Fator 4 Semelhante a Kruppel , CamundongosRESUMO
Skin fibroblasts were collected from a 44-year-old patient with sporadic case of Parkinson's disease (PD). The non-integrating Sendai virus vector encoding OCT3/4, SOX2, c-MYC and KLF4 was used to reprogram fibroblasts into induced pluripotent stem cells (iPSCs). Generated iPSCs had normal karyotypes, expressed common stem cell markers, and were capable of differentiating into all three germ layers. Generated line could be used for PD modeling to understand the mechanisms that influence the disorder.
Assuntos
Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Doença de Parkinson/metabolismo , Adulto , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Células Cultivadas , Feminino , Fibroblastos/citologia , Fibroblastos/metabolismo , Camadas Germinativas/citologia , Camadas Germinativas/metabolismo , Humanos , Cariótipo , Fator 4 Semelhante a KruppelRESUMO
An induced pluripotent stem cell (iPSC) line was generated from a 36-year-old patient with sporadic Parkinson's disease (PD). Skin fibroblasts were reprogrammed using the non-integrating Sendai virus technology to deliver OCT3/4, SOX2, c-MYC and KLF4 factors. The generated cell line (CSC-43) exhibits expression of common pluripotency markers, in vitro differentiation into three germ layers and normal karyotype. This iPSC line can be used to study the mechanisms underlying the development of PD.
Assuntos
Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Doença de Parkinson/metabolismo , Adulto , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Linhagem Celular , Reprogramação Celular/genética , Reprogramação Celular/fisiologia , Humanos , Fator 4 Semelhante a Kruppel , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Masculino , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismoRESUMO
Parkinson's disease (PD) is a neurodegenerative disease with unknown etiology. Here we show the generation of an induced pluripotent stem cell (iPSC) line, named CSC-40, from dermal fibroblasts obtained from a 59-year-old male patient with a homozygous p.Q456X mutation in the PTEN-induced putative kinase 1 (PINK/PARK6) gene and a confirmed diagnosis of PD, which could be used to model familial PD. A non-integrating Sendai virus-based delivery of the reprogramming factors OCT3/4, SOX2, c-MYC and KLF4 was employed. The CSC-40 cell line showed normal karyotyping and fingerprinting following transduction as well as sustained expression of several pluripotency markers and the ability to differentiate into all three germ layers.
Assuntos
Células-Tronco Pluripotentes Induzidas/metabolismo , Doença de Parkinson/genética , Proteínas Quinases/genética , Linhagem Celular , Células Cultivadas , Humanos , Fator 4 Semelhante a Kruppel , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Masculino , Pessoa de Meia-Idade , Mutação/genética , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Doença de Parkinson/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismoRESUMO
Mutations in the PARK2 gene, which encodes PARKIN, are the most frequent cause of autosomal recessive Parkinson's disease (PD). We report the generation of an induced pluripotent stem cell (iPSC) line from a 78-year-old patient carrying a compound heterozygous mutation (c.823C>T and EX6del) in the PARK2 gene. Skin fibroblasts were reprogrammed using the non-integrating Sendai virus technology to deliver OCT3/4, SOX2, c-MYC and KLF4 factors. The generated cell line CSC-44 exhibits expression of common pluripotency markers, in vitro differentiation into the three germ layers and normal karyotype. This iPSC line can be used to explore the association between PARK2 mutations and PD.
Assuntos
Diferenciação Celular/fisiologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Doença de Parkinson/genética , Ubiquitina-Proteína Ligases/genética , Adulto , Idoso , Diferenciação Celular/genética , Células Cultivadas , Reprogramação Celular/genética , Reprogramação Celular/fisiologia , Feminino , Heterozigoto , Humanos , Fator 4 Semelhante a Kruppel , Mutação/genéticaRESUMO
Induced pluripotent stem cells (iPSCs) are becoming an important source of pre-clinical models for research focusing on neurodegeneration. They offer the possibility for better understanding of common and divergent pathogenic mechanisms of brain diseases. Moreover, iPSCs provide a unique opportunity to develop personalized therapeutic strategies, as well as explore early pathogenic mechanisms, since they rely on the use of patients' own cells that are otherwise accessible only post-mortem, when neuronal death-related cellular pathways and processes are advanced and adaptive. Neurodegenerative diseases are in majority of unknown cause, but mutations in specific genes can lead to familial forms of these diseases. For example, mutations in the superoxide dismutase 1 gene lead to the motor neuron disease amyotrophic lateral sclerosis (ALS), while mutations in the SNCA gene encoding for alpha-synuclein protein lead to familial Parkinson's disease (PD). The generations of libraries of familial human ALS iPSC lines have been described, and the iPSCs rapidly became useful models for studying cell autonomous and non-cell autonomous mechanisms of the disease. Here we report the generation of a comprehensive library of iPSC lines of familial PD and an associated synucleinopathy, multiple system atrophy (MSA). In addition, we provide examples of relevant neural cell types these iPSC can be differentiated into, and which could be used to further explore early disease mechanisms. These human cellular models will be a valuable resource for identifying common and divergent mechanisms leading to neurodegeneration in PD and MSA.
RESUMO
Solitary fibrous tumor (SFT) is a mesenchymal neoplasm displaying variable morphologic and clinical features. To identify pathogenetically important genetic rearrangements, 44 SFTs were analyzed using a variety of techniques. Chromosome banding and fluorescence in situ hybridization (FISH) showed recurrent breakpoints in 12q13, clustering near the NAB2 and STAT6 genes, and single nucleotide polymorphism array analysis disclosed frequent deletions affecting STAT6. Quantitative real-time PCR revealed high expression levels of the 5'-end of NAB2 and the 3'-end of STAT6, which at deep sequencing of enriched DNA corresponded to NAB2/STAT6 fusions. Subsequent reverse-transcriptase PCR (RT-PCR) analysis identified a NAB2/STAT6 fusion in 37/41 cases, confirming that this fusion gene underlies the pathogenesis of SFT. The hypothesis that the NAB2/STAT6 fusions will result in altered properties of the transcriptional co-repressor NAB2--a key regulator of the early growth response 1 (EGR1) transcription factor - was corroborated by global gene expression analysis; SFTs showed deregulated expression of EGR1 target genes, as well as of other, developmentally important genes. We also identified several nonrandom secondary changes, notably loss of material from 13q and 14q. As neither chromosome banding nor FISH analysis identify more than a minor fraction of the fusion-positive cases, and because multiple primer combinations are required to identify all possible fusion transcripts by RT-PCR, alternative diagnostic markers might instead be found among deregulated genes identified at global gene expression analysis. Indeed, using immunohistochemistry on tissue microarrays, the top up-regulated gene, GRIA2, was found to be differentially expressed also at the protein level.