Gene Expression Profiles Controlled by the Alternative Splicing Factor Nova2 in Endothelial Cells.
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ABSTRACT:
SUBMITTER: Belloni E
PROVIDER: S-EPMC6953062 | biostudies-literature | 2019 Nov
REPOSITORIES: biostudies-literature
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Project description:Angiogenesis is crucial for cancer progression. While several anti-angiogenic drugs are in use for cancer treatment, their clinical benefits are unsatisfactory. Thus, a deeper understanding of the mechanisms sustaining cancer vessel growth is fundamental to identify novel biomarkers and therapeutic targets. Alternative splicing (AS) is an essential modifier of human proteome diversity. Nevertheless, AS contribution to tumor vasculature development is poorly known. The Neuro-Oncological Ventral Antigen 2 (NOVA2) is a critical AS regulator of angiogenesis and vascular development. NOVA2 is upregulated in tumor endothelial cells (ECs) of different cancers, thus representing a potential driver of tumor blood vessel aberrancies. Here, we identified novel AS transcripts generated upon NOVA2 upregulation in ECs, suggesting a pervasive role of NOVA2 in vascular biology. In addition, we report that NOVA2 is also upregulated in ECs of gastric cancer (GC), and its expression correlates with poor overall survival of GC patients. Finally, we found that the AS of the Rap Guanine Nucleotide Exchange Factor 6 (RapGEF6), a newly identified NOVA2 target, is altered in GC patients and associated with NOVA2 expression, tumor angiogenesis, and poor patient outcome. Our findings provide a better understanding of GC biology and suggest that AS might be exploited to identify novel biomarkers and therapeutics for anti-angiogenic GC treatments.
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Project description:Vascular lumen formation is a fundamental step during angiogenesis; yet, the molecular mechanisms underlying this process are poorly understood. Recent studies have shown that neural and vascular systems share common anatomical, functional and molecular similarities. Here we show that the organization of endothelial lumen is controlled at the post-transcriptional level by the alternative splicing (AS) regulator Nova2, which was previously considered to be neural cell-specific. Nova2 is expressed during angiogenesis and its depletion disrupts vascular lumen formation in vivo. Similarly, Nova2 depletion in cultured endothelial cells (ECs) impairs the apical distribution and the downstream signalling of the Par polarity complex, resulting in altered EC polarity, a process required for vascular lumen formation. These defects are linked to AS changes of Nova2 target exons affecting the Par complex and its regulators. Collectively, our results reveal that Nova2 functions as an AS regulator in angiogenesis and is a novel member of the 'angioneurins' family.
| S-EPMC4633719 | biostudies-literature
Project description:BackgroundTumor-specific isoforms generated by alternative splicing (AS) are demonstrated to contribute to tumor progression and can represent potential biomarkers. NOVA2 is an AS factor that in physiological conditions regulates endothelial cells' (ECs) polarity and vessel lumen maturation, likely by mediating AS of apical-basal polarity regulators. However, NOVA2 expression in tumor ECs and its regulation have never been investigated.MethodsTo elucidate this, 40 colorectal cancer patients were enrolled and NOVA2 expression was investigated by immunohistochemistry in samples bearing both the normal mucosa and the tumor tissue.ResultsNOVA2 was found expressed in ECs of tumor vasculature and, importantly, it was upregulated in tumor ECs with respect to normal mucosa ECs in all cases (P<0.001). The same samples analyzed by immunohistochemistry for the expression HIF1α, a marker of hypoxia, showed a positive and significant association with NOVA2 levels (P=0.045). Of note, NOVA2 was upregulated by hypoxia also in an in vitro ECs model.ConclusionOur results provide, for the first time, evidence of NOVA2 expression and upregulation in tumor ECs and highlight hypoxia as a potential regulatory factor. These findings open a completely new perspective to study tumor vasculature and to uncover NOVA2 as a potential source of biomarkers and therapeutic targets based on AS isoforms.
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Project description:BackgroundAdaptation to low oxygen by changing gene expression is vitally important for cell survival and tissue development. The sprouting of new blood vessels, initiated from endothelial cells, restores the oxygen supply of ischemic tissues. In contrast to the transcriptional response induced by hypoxia, which is mainly mediated by members of the HIF family, there are only few studies investigating alternative splicing events. Therefore, we performed an exon array for the genome-wide analysis of hypoxia-related changes of alternative splicing in endothelial cells.Methodology/principal findingsHuman umbilical vein endothelial cells (HUVECs) were incubated under hypoxic conditions (1% O(2)) for 48 h. Genome-wide transcript and exon expression levels were assessed using the Affymetrix GeneChip Human Exon 1.0 ST Array. We found altered expression of 294 genes after hypoxia treatment. Upregulated genes are highly enriched in glucose metabolism and angiogenesis related processes, whereas downregulated genes are mainly connected to cell cycle and DNA repair. Thus, gene expression patterns recapitulate known adaptations to low oxygen supply. Alternative splicing events, until now not related to hypoxia, are shown for nine genes: six which are implicated in angiogenesis-mediated cytoskeleton remodeling (cask, itsn1, larp6, sptan1, tpm1 and robo1); one, which is involved in the synthesis of membrane-anchors (pign) and two universal regulators of gene expression (cugbp1 and max).Conclusions/significanceFor the first time, this study investigates changes in splicing in the physiological response to hypoxia on a genome-wide scale. Nine alternative splicing events, until now not related to hypoxia, are reported, considerably expanding the information on splicing changes due to low oxygen supply. Therefore, this study provides further knowledge on hypoxia induced gene expression changes and presents new starting points to study the hypoxia adaptation of endothelial cells.
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Project description:Farnesyl diphosphate synthase (FDPS), an essential enzyme involved in the mevalonate pathway, is implicated in various diseases, including multiple types of cancer. As an RNA-binding protein (RBP), FDPS is also involved in transcriptional and post-transcriptional regulation. However, to the best of our knowledge, transcriptome-wide targets of FDPS still remain unknown. In the present study, FDPS expression patterns in pan-cancer were analyzed. In addition, it was investigated how FDPS overexpression (FDPS-OE) regulates the transcriptome in HeLa cells. FDPS-OE increased the proliferation rate in HeLa cells by MTT assay. Using transcriptome-wide high throughput sequencing and bioinformatics analysis, it was found that FDPS upregulated the expression levels of genes enriched in cell proliferation and extracellular matrix organization, including the laminin subunit γ2, interferon-induced proteins with tetratricopeptide repeats 2 and matrix metallopeptidase 19 genes. According to alternative splicing (AS) analysis, FDPS modulated the splicing patterns of the bone morphogenic protein 1, semaphorin 4D, annexin A2 and sirtuin 2 genes, which are enriched in the cell cycle and DNA repair, and are related to cell proliferation. To corroborate the FDPS-regulated transcriptome findings, FDPS was overexpressed in human osteosarcoma cells. Differentially expressed genes and regulated AS genes in the cells were both validated by reverse transcription-quantitative PCR. The results suggested that, as an emerging RBP, FDPS may serve an important role in transcriptome profiles by altering gene expression and regulating AS. FDPS also affected the cell proliferation rate. These findings broaden the understanding of the molecular functions of FDPS, and the potential of FDPS as a target in therapy should be investigated.
| S-EPMC10018273 | biostudies-literature
Project description:The biological players involved in angiogenesis are only partially defined. Here, we report that endothelial cells (ECs) express a novel isoform of the cell-surface adhesion molecule L1CAM, termed L1-?TM. The splicing factor NOVA2, which binds directly to L1CAM pre-mRNA, is necessary and sufficient for the skipping of L1CAM transmembrane domain in ECs, leading to the release of soluble L1-?TM. The latter exerts high angiogenic function through both autocrine and paracrine activities. Mechanistically, L1-?TM-induced angiogenesis requires fibroblast growth factor receptor-1 signaling, implying a crosstalk between the two molecules. NOVA2 and L1-?TM are overexpressed in the vasculature of ovarian cancer, where L1-?TM levels correlate with tumor vascularization, supporting the involvement of NOVA2-mediated L1-?TM production in tumor angiogenesis. Finally, high NOVA2 expression is associated with poor outcome in ovarian cancer patients. Our results point to L1-?TM as a novel, EC-derived angiogenic factor which may represent a target for innovative antiangiogenic therapies.
| S-EPMC6398979 | biostudies-literature
Project description:Angiogenesis is crucial for cancer progression and anti-angiogenic drugs are in use for its treatment. However, their clinical benefits are unsatisfactory. Thus, a deeper understanding of the mechanisms sustaining cancer vessel growth is crucial to identify better biomarkers and novel therapeutic targets. Alternative Splicing (AS) is an essential modifier of human proteome diversity. Nevertheless, its contribution to tumor vasculature development is poorly known. NOVA2 is an important AS regulator of angiogenesis and vascular development and it is up-regulated in tumor endothelial cells (ECs) of different cancers, thus representing a potential driver of tumor blood vessels aberrancies. Here, we identified novel AS transcripts generated upon NOVA2 upregulation in ECs, suggesting a pervasive role of NOVA2 in vascular biology. In addition, we report that NOVA2 is also upregulated in ECs of gastric cancer (GC), with its expression correlating with poor overall survival of the GC patients. We also found that the AS of the newly identified NOVA2 target RapGEF6 is altered in GC patients and associated with NOVA2 expression, tumor angiogenesis and poor patient outcome. Our findings provide a better understanding of GC biology and suggest that AS might represent a potential source of biomarkers and therapeutics for novel anti-angiogenic GC treatments.
2023-08-04 | GSE224823 | GEO
Project description:Alternative splicing (AS) is crucial for cell-type-specific gene transcription and plays a critical role in neuronal differentiation and synaptic plasticity. De novo frameshift variants in NOVA2, encoding a neuron-specific key splicing factor, have been recently associated with a new neurodevelopmental disorder (NDD) with hypotonia, neurological features, and brain abnormalities. We investigated eight unrelated individuals by exome sequencing (ES) and identified seven novel pathogenic NOVA2 variants, including two with a novel localization at the KH1 and KH3 domains. In addition to a severe NDD phenotype, novel clinical features included psychomotor regression, attention deficit-hyperactivity disorder (ADHD), dyspraxia, and urogenital and endocrinological manifestations. To test the effect of the variants on splicing regulation, we transfected HeLa cells with wildtype and mutant NOVA2 complementary DNA (cDNA). The novel variants NM_002516.4:c.754_756delCTGinsTT p.(Leu252Phefs*144) and c.1329dup p.(Lys444Glnfs*82) all negatively affected AS events. The distal p.(Lys444Glnfs*82) variant, causing a partial removal of the KH3 domain, had a milder functional effect leading to an intermediate phenotype. Our findings expand the molecular and phenotypic spectrum of NOVA2-related NDD, supporting the pathogenic role of AS disruption by truncating variants and suggesting that this is a heterogeneous condition with variable clinical course.
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