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1.
Front Cell Dev Biol ; 11: 1147625, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37936982

RESUMO

Introduction: Proline/arginine-rich end leucine-rich repeat protein (PRELP), is a small secreted proteoglycan expressed by pericytes and vascular smooth muscle cells surrounding the brain vasculature of adult mouse. Methods: We utilised a Prelp knockout (Prelp -/-) mouse model to interrogate vasculature integrity in the brain alongside performing in vitro assays to characterise PRELP application to endothelial cells lines. Our findings were supplemented with RNA expression profiling to elucidate the mechanism of how PRELP maintains neurovasculature function. Results: Prelp -/- mice presented with neuroinflammation and reducedneurovasculature integrity, resulting in IgG and dextran leakage in the cerebellum and cortex. Histological analysis of Prelp -/- mice revealed reducedcell-cell integrity of the blood brain barrier, capillary attachment of pericytes andastrocyte end-feet. RNA-sequencing analysis found that cell-cell adhesion andinflammation are affected in Prelp -/- mice and gene ontology analysis as well as gene set enrichment analysis demonstrated that inflammation related processes and adhesion related processes such as epithelial-mesenchymal transition and apical junctions were significantly affected, suggesting PRELP is a regulator of cell-cell adhesion. Immunofluorescence analysis showed that adhesion junction protein expression levels of cadherin, claudin-5, and ZO-1, was suppressed in Prelp -/- mice neurovasculature. Additionally, in vitro studies revealed that PRELP application to endothelial cells enhances cell-cell integrity, induces mesenchymal-endothelial transition and inhibits TGF-ß mediated damage to cell-cell adhesion. Discussion: Our study indicates that PRELP is a novel endogenous secreted regulator of neurovasculature integrity and that PRELP application may be a potential treatment for diseases associated with neurovascular damage.

2.
NAR Cancer ; 5(3): zcad046, 2023 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-37636315

RESUMO

Constant communication between mitochondria and nucleus ensures cellular homeostasis and adaptation to mitochondrial stress. Anterograde regulatory pathways involving a large number of nuclear-encoded proteins control mitochondrial biogenesis and functions. Such functions are deregulated in cancer cells, resulting in proliferative advantages, aggressive disease and therapeutic resistance. Transcriptional networks controlling the nuclear-encoded mitochondrial genes are known, however alternative splicing (AS) regulation has not been implicated in this communication. Here, we show that IQGAP1, a scaffold protein regulating AS of distinct gene subsets in gastric cancer cells, participates in AS regulation that strongly affects mitochondrial respiration. Combined proteomic and RNA-seq analyses of IQGAP1KO and parental cells show that IQGAP1KO alters an AS event of the mitochondrial respiratory chain complex I (CI) subunit NDUFS4 and downregulates a subset of CI subunits. In IQGAP1KO cells, CI intermediates accumulate, resembling assembly deficiencies observed in patients with Leigh syndrome bearing NDUFS4 mutations. Mitochondrial CI activity is significantly lower in KO compared to parental cells, while exogenous expression of IQGAP1 reverses mitochondrial defects of IQGAP1KO cells. Our work sheds light to a novel facet of IQGAP1 in mitochondrial quality control that involves fine-tuning of CI activity through AS regulation in gastric cancer cells relying highly on mitochondrial respiration.

3.
Cancers (Basel) ; 14(19)2022 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-36230849

RESUMO

Retinoblastoma (RB) is the most common intraocular pediatric cancer. Nearly all cases of RB are associated with mutations compromising the function of the RB1 tumor suppressor gene. We previously demonstrated that PRELP is widely downregulated in various cancers and our in vivo and in vitro analysis revealed PRELP as a novel tumor suppressor and regulator of EMT. In addition, PRELP is located at chromosome 1q31.1, around a region hypothesized to be associated with the initiation of malignancy in RB. Therefore, in this study, we investigated the role of PRELP in RB through in vitro analysis and next-generation sequencing. Immunostaining revealed that PRELP is expressed in Müller glial cells in the retina. mRNA expression profiling of PRELP-/- mouse retina and PRELP-treated RB cells found that PRELP contributes to RB progression via regulation of the cancer microenvironment, in which loss of PRELP reduces cell-cell adhesion and facilitates EMT. Our observations suggest that PRELP may have potential as a new strategy for RB treatment.

4.
Oncogene ; 40(36): 5518-5532, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34294847

RESUMO

In response to oncogenic signals, Alternative Splicing (AS) regulators such as SR and hnRNP proteins show altered expression levels, subnuclear distribution and/or post-translational modification status, but the link between signals and these changes remains unknown. Here, we report that a cytosolic scaffold protein, IQGAP1, performs this task in response to heat-induced signals. We show that in gastric cancer cells, a nuclear pool of IQGAP1 acts as a tethering module for a group of spliceosome components, including hnRNPM, a splicing factor critical for the response of the spliceosome to heat-shock. IQGAP1 controls hnRNPM's sumoylation, subnuclear localisation and the relevant response of the AS machinery to heat-induced stress. Genome-wide analyses reveal that IQGAP1 and hnRNPM co-regulate the AS of a cell cycle-related RNA regulon in gastric cancer cells, thus favouring the accelerated proliferation phenotype of gastric cancer cells. Overall, we reveal a missing link between stress signals and AS regulation.


Assuntos
Neoplasias Gástricas , Processamento Alternativo , Estudo de Associação Genômica Ampla , Humanos , Estômago , Proteínas Ativadoras de ras GTPase
5.
Cancers (Basel) ; 12(11)2020 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-33202923

RESUMO

Osteomodulin (OMD) and proline/arginine-rich end leucine repeat protein (PRELP) are secreted extracellular matrix proteins belonging to the small leucine-rich proteoglycans family. We found that OMD and PRELP were specifically expressed in umbrella cells in bladder epithelia, and their expression levels were dramatically downregulated in all bladder cancers from very early stages and various epithelial cancers. Our in vitro studies including gene expression profiling using bladder cancer cell lines revealed that OMD or PRELP application suppressed the cancer progression by inhibiting TGF-ß and EGF pathways, which reversed epithelial-mesenchymal transition (EMT), activated cell-cell adhesion, and inhibited various oncogenic pathways. Furthermore, the overexpression of OMD in bladder cancer cells strongly inhibited the anchorage-independent growth and tumorigenicity in mouse xenograft studies. On the other hand, we found that in the bladder epithelia, the knockout mice of OMD and/or PRELP gene caused partial EMT and a loss of tight junctions of the umbrella cells and resulted in formation of a bladder carcinoma in situ-like structure by spontaneous breakdowns of the umbrella cell layer. Furthermore, the ontological analysis of the expression profiling of an OMD knockout mouse bladder demonstrated very high similarity with those obtained from human bladder cancers. Our data indicate that OMD and PRELP are endogenous inhibitors of cancer initiation and progression by controlling EMT. OMD and/or PRELP may have potential for the treatment of bladder cancer.

6.
Invest Ophthalmol Vis Sci ; 56(1): 164-76, 2014 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-25414197

RESUMO

PURPOSE: Mutations in the Prominin-1 (Prom1) gene are known to cause retinitis pigmentosa and Stargardt disease, both of which are associated with progressive photoreceptor cell death. There are no effective therapies for either disorder. The aim of this study was to investigate the mechanism of the retinal degeneration in Prom1-deficient mouse models. METHODS: We constructed Prom1 knockout mice with two distinct genetic backgrounds of C57BL/6 and C57BL/6xCBA/NSlc, and investigated the photoreceptor degeneration by means of histology and functional tests.. In addition, we examined the effect of light on the Prom1(-/-) retina by rearing the mice in the normal light/dark cycle and completely dark conditions. Finally, we investigated if the retinoic-acid derivative Fenretinide slowed the pace of retinal degeneration in these mouse models. RESULTS: The Prom1(-/-)-knockout mice with both backgrounds developed photoreceptor degeneration after eye opening, but the CB57/BL6-background mice developed photoreceptor cell degeneration much faster than the C57BL/6xCBA/NSlc mice, demonstrating genetic background dependency.. Interestingly, our histologic and functional examination showed that the photoreceptor cell degeneration of Prom1-knockout mice was light-dependent, and was almost completely inhibited when the mutant mice were kept in the dark. The Prom1-knockout retina showed strong downregulation of expression of the visual cycle components, Rdh12 and Abca4. Furthermore, administration of Fenretinide, which lowers the level of the toxic lipofuscin, slowed the degeneration of photoreceptor cells. CONCLUSIONS: These findings improve our understanding of the mechanism of cell death in Prominin-1-related disease and provide evidence that fenretinide may be worth studying in human disease.


Assuntos
Antígenos CD/genética , DNA/genética , Glicoproteínas/genética , Mutação , Peptídeos/genética , Células Fotorreceptoras de Vertebrados/ultraestrutura , Degeneração Retiniana/genética , Antígeno AC133 , Animais , Antígenos CD/metabolismo , Análise Mutacional de DNA , Modelos Animais de Doenças , Eletrorretinografia , Glicoproteínas/metabolismo , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Eletrônica , Peptídeos/metabolismo , Estimulação Luminosa , Células Fotorreceptoras de Vertebrados/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Degeneração Retiniana/metabolismo , Degeneração Retiniana/fisiopatologia
7.
Mol Ther ; 21(10): 1862-75, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23799534

RESUMO

Huntington's disease (HD) is a devastating neurodegenerative disorder caused by abnormal polyglutamine expansion in the huntingtin protein (Exp-Htt). Currently, there are no effective treatments for HD. We used bidirectional lentiviral transfer vectors to generate in vitro and in vivo models of HD and to test the therapeutic potential of vascular endothelial growth factor 165 (VEGF165). Lentiviral-mediated expression of Exp-Htt caused cell death and aggregate formation in human neuroblastoma SH-SY5Y and rat primary striatal cultures. Lentiviral-mediated VEGF165 expression was found to be neuroprotective in both of these models. Unilateral stereotaxic vector delivery of Exp-Htt vector in adult rat striatum led to progressive inclusion formation and striatal neuron loss at 10 weeks post-transduction. Coinjection of a lower dose VEGF165 significantly attenuated DARPP-32(+) neuronal loss, enhanced NeuN staining and reduced Exp-Htt aggregation. A tenfold higher dose VEGF165 led to overt neuronal toxicity marked by tissue damage, neovascularization, extensive astrogliosis, vascular leakage, chronic inflammation and distal neuronal loss. No overt behavioral phenotype was observed in these animals. Expression of VEGF165 at this higher dose in the brain of wild-type rats led to early mortality with global neuronal loss. This report raises important safety concerns about unregulated VEGF165 CNS applications.


Assuntos
Corpo Estriado/patologia , Terapia Genética , Doença de Huntington/patologia , Degeneração Neural/patologia , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Morte Celular , Linhagem Celular Tumoral , Células Cultivadas , Corpo Estriado/metabolismo , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica , Vetores Genéticos , Células HEK293 , Humanos , Proteína Huntingtina , Doença de Huntington/genética , Lentivirus/genética , Masculino , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neurônios/patologia , Fármacos Neuroprotetores , Ratos , Ratos Sprague-Dawley , Transdução Genética
8.
Dev Growth Differ ; 54(3): 327-40, 2012 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-22524604

RESUMO

The small leucine-rich repeat proteoglycan (SLRPs) family of proteins currently consists of five classes, based on their structural composition and chromosomal location. As biologically active components of the extracellular matrix (ECM), SLRPs were known to bind to various collagens, having a role in regulating fibril assembly, organization and degradation. More recently, as a function of their diverse proteins cores and glycosaminoglycan side chains, SLRPs have been shown to be able to bind various cell surface receptors, growth factors, cytokines and other ECM components resulting in the ability to influence various cellular functions. Their involvement in several signaling pathways such as Wnt, transforming growth factor-ß and epidermal growth factor receptor also highlights their role as matricellular proteins. SLRP family members are expressed during neural development and in adult neural tissues, including ocular tissues. This review focuses on describing SLRP family members involvement in neural development with a brief summary of their role in non-neural ocular tissues and in response to neural injury.


Assuntos
Decorina/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Neurogênese , Proteínas/metabolismo , Via de Sinalização Wnt , Animais , Desenvolvimento Embrionário , Matriz Extracelular/metabolismo , Matriz Extracelular/fisiologia , Proteínas da Matriz Extracelular/metabolismo , Humanos , Proteínas de Repetições Ricas em Leucina , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Ligação Proteica , Retina/citologia , Retina/embriologia , Retina/lesões , Retina/metabolismo , Nicho de Células-Tronco , Fator de Crescimento Transformador beta/metabolismo
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