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Background: Primary ciliary dyskinesia (PCD) is considered a rare cause of chronic rhinosinusitis with nasal polyposis (CRSwNP), which is reported in 6% of children with PCD. The forms of PCD associated with the variants of the GAS8 gene identified so far seem to be linked to recurrent respiratory infections (sinusitis, otitis, and bronchiectasis) without situs inversus. Case presentation: We report a case of an 11-year-old girl with recurrent otitis media, productive cough, and chronic rhinosinusitis with nasal polyposis with homozygosity for a novel nonsense mutation in the GAS8. Conclusion: Children with CRSwNP should be treated in a multidisciplinary manner (ENT, pulmonologist, allergist, pathologist, pediatrician, and geneticist) because nasal polyposis often hides etiologies that must be recognized.
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PURPOSE: Long noncoding RNAs (lncRNAs) play an essential role in the epigenetic regulation of various key genes involved in vital cellular functions. A somatic dinucleotide mutation in the lncRNA GAS8-AS1 was reported in Chinese papillary thyroid cancer. However, GAS8-AS1 dinucleotide alteration and its impact have never been explored in differentiated thyroid cancers and other populations. METHODS: We extracted genomic DNA from 265 DTCs and 97 normal healthy subjects, PCR amplified and Sanger sequenced to examine the GAS8-AS1 dinucleotide alteration. Calculated genotype/allele frequency to test Hardy-Weinberg Equilibrium (HWE) and performed a genetic model of inheritance to determine its association with DTC risk. Correlated the GAS8-AS1 dinucleotide variant distribution with clinical characteristics to find the association. Predicted GAS8-AS1 RNA secondary structure for wild type and variant using RemuRNA and RNAfold to assess the conformational changes. RESULTS: GAS8-AS1 dinucleotide alteration (n.713A > G, rs55742939; n.714T > C, rs61118444) identified in DTCs is a germline variant not somatic. The GAS8-AS1 genotype and allele frequency significantly deviated for HWE in DTCs (χ2 = 37.954; p = 0.0001) though not associated with its risk. Dinucleotide variant distribution was remarkably associated with early-stage disease (p = 0.002), lymph node (p = 0.01), and distant metastasis (p = 0.01) in DTCs. The GAS8-AS1 bearing dinucleotide variant markedly showed conformational change compared to that of its wild type. CONCLUSIONS: These findings indicate that GAS8-AS1 is genetically deregulated and implicated in several stages of DTC tumorigenesis suggesting it could be a promising prognostic biomarker in DTCs.
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Metástasis Linfática , ARN Largo no Codificante , Neoplasias de la Tiroides , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Genotipo , Metástasis Linfática/genética , Estadificación de Neoplasias , ARN Largo no Codificante/genética , Cáncer Papilar Tiroideo/genética , Cáncer Papilar Tiroideo/patología , Neoplasias de la Tiroides/genética , Neoplasias de la Tiroides/patologíaRESUMEN
Sperm flagella share an evolutionary conserved microtubule-based structure with motile cilia expressed at the surface of several cell types, such as the airways epithelial cells. As a result, male infertility can be observed as an isolated condition or a syndromic trait, illustrated by Primary Cilia Dyskinesia (PCD). We report two unrelated patients showing multiple morphological abnormalities of the sperm flagella (MMAF) and carrying distinct homozygous truncating variants in the PCD-associated gene CCDC65. We characterized one of the identified variants (c.1208del; p.Asn403Ilefs*9), which induces the near absence of CCDC65 protein in patient sperm. In Chlamydomonas, CCDC65 ortholog (DRC2, FAP250) is a component of the Nexin-Dynein Regulatory complex (N-DRC), which interconnects microtubule doublets and coordinates dynein arms activity. In sperm cells from the patient, we also show the loss of GAS8, another component of the N-DRC, supporting a structural/functional link between the two proteins. Our work indicates that, similarly to ciliary axoneme, CCDC65 is required for sperm flagellum structure. Importantly, our work provides first evidence that mutations in the PCD-associated gene CCDC65 also cause asthenozoospermia.
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Infertilidad Masculina , Cola del Espermatozoide , Humanos , Masculino , Cola del Espermatozoide/metabolismo , Axonema/genética , Semillas/metabolismo , Proteínas Asociadas a Microtúbulos/genética , Mutación/genética , Dineínas/genética , Infertilidad Masculina/genética , Glicoproteínas/genéticaRESUMEN
Motile cilia and flagella are closely related organelles structured around a highly conserved axoneme whose formation and maintenance involve proteins from hundreds of genes. Defects in many of these genes have been described to induce primary ciliary dyskinesia (PCD) mainly characterized by chronic respiratory infections, situs inversus and/or infertility. In men, cilia/flagella-related infertility is usually caused by asthenozoospermia due to multiple morphological abnormalities of the sperm flagella (MMAF). Here, we investigated a cohort of 196 infertile men displaying a typical MMAF phenotype without any other PCD symptoms. Analysis of WES data identified a single case carrying a deleterious homozygous GAS8 variant altering a splice donor consensus site. This gene, also known as DRC4, encodes a subunit of the Nexin-Dynein Regulatory Complex (N-DRC), and has been already associated to male infertility and mild PCD. Confirming the deleterious effect of the candidate variant, GAS8 staining by immunofluorescence did not evidence any signal from the patient's spermatozoa whereas a strong signal was present along the whole flagella length in control cells. Concordant with its role in the N-DRC, transmission electron microscopy evidenced peripheral microtubule doublets misalignments. We confirm here the importance of GAS8 in the N-DRC and observed that its absence induces a typical MMAF phenotype not necessarily accompanied by other PCD symptoms.
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Axonema , Infertilidad Masculina , Masculino , Humanos , Axonema/genética , Mutación , Semen , Cola del Espermatozoide , Infertilidad Masculina/genética , Espermatozoides , Flagelos , Proteínas Asociadas a Microtúbulos/genética , Dineínas/genéticaRESUMEN
PURPOSE: Ciliopathies are broadly classified as non-motile or motile (primary ciliary dyskinesia). Early-onset retinal dystrophy is common in non-motile ciliopathy, but retinal dystrophy is not considered a feature of primary ciliary dyskinesia. The subject of this report is woman referred as a case of Stargardt disease who in fact had retinal dystrophy apparently related to GAS8-related primary ciliary dyskinesia. METHODS: Retrospective case report. RESULTS: A 43-year-old Emirati woman was referred for further evaluation of Stargardt disease. Her only ophthalmic complaints were related to dry eye disease. Past ocular history was significant for refractive surgery in her early 30's. Past medical history was significant for primary ciliary dyskinesia, which included recurrent bronchiectasis and sino-pulmonary infections since childhood. Clinical examination confirmed retinopathy resembling Stargardt disease. Electroretinography revealed cone-rod dysfunction. Whole exome sequencing with attention to ABCA4 was unrevealing for retinal dystrophy genes but did uncover a homozygous GAS8 deletion, molecularly confirming the diagnosis of primary ciliary dyskinesia. Literature review revealed a report of a 34-year-old North African male with GAS8-related primary ciliary dyskinesia who also had been diagnosed with Stargardt disease in the absence of pathogenic ABCA4 variants. DISCUSSION: Longer follow-up of individuals with primary ciliary dyskinesia may reveal findings more typically associated with non-motile ciliopathy such as retinal dystrophy. GAS8-related retinal dystrophy can resemble Stargardt disease.
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Trastornos de la Motilidad Ciliar , Ciliopatías , Distrofias Retinianas , Transportadoras de Casetes de Unión a ATP/genética , Trastornos de la Motilidad Ciliar/diagnóstico , Trastornos de la Motilidad Ciliar/genética , Femenino , Humanos , Masculino , Mutación , Fenotipo , Distrofias Retinianas/genética , Estudios Retrospectivos , Enfermedad de StargardtRESUMEN
Long non-coding (lnc)RNAs have been recognized as important regulators in gastric cancer. lncRNA GAS8-AS1 is considered a tumor suppressor in multiple types of cancer, such as papillary thyroid carcinoma, ovarian cancer and colorectal cancer. However, the specific role of GAS8-AS1 in gastric cancer remains to be fully elucidated. The aim of the present study was to investigate the role of GAS8-AS1 in gastric cancer and its potential underlying mechanisms of action. The expression levels of GAS8-AS1, microRNA (miR)-21-3p, PTEN and pyruvate dehydrogenase (E1) alpha subunit gene (PDHA1) in gastric cancer and non-cancerous tissues, as well as in gastric cancer cell lines, were detected using reverse transcription-quantitative PCR. Cell proliferation was detected by using a Cell Counting Kit-8 assay. Cell migration and invasion were detected using a Transwell assay. Results of the present study demonstrated that the expression levels of GAS8-AS1 in gastric cancer tissues were significantly decreased, whereas its expression did not differ among cancer tissues at different clinical stages. Low expression levels of GAS8-AS1 predicted poor 5-year survival rates for 70 patients with gastric adenocarcinoma from the Affiliated Hospital of Xuzhou Medical University (Xuzhou, China) during patient follow-up. In addition, the expression levels of miR-21-3p were markedly increased in cancer tissues, and miR-21-3p expression was negatively associated with the expression of GAS8-AS1. The direct interaction between GAS8-AS1 and miR-21-3p was predicted using the starBase database and was confirmed by using an RNA pull-down assay. In gastric cell lines, the overexpression of GAS8-AS1 reduced the expression levels of mature miR-21-3p but did not affect the expression of miR-21-3p precursor, while the overexpression of miR-21-3p did not, in turn, affect the expression of GAS8-AS1. In addition, the overexpression of GAS8-AS1 inhibited cancer cell proliferation, while the overexpression of miR-21-3p promoted cancer cell proliferation and attenuated the effects of GAS8-AS1. Overexpression of miR-21-3p promoted cancer cell migration and invasion, whereas overexpression of GAS8-AS1 did not affect cell migration or invasion. In summary, results of the present study have demonstrated that GAS8-AS1 acts as a tumor suppressor in gastric cancer, and it may inhibit cancer cell proliferation by downregulating miR-21-3p.
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BACKGROUND: LncRNA GAS8-AS1 has been reported to participate in several types of cancer, while its role in glioblastoma (GBM) is unknown. In the present study, we aimed to investigate the function of GAS8-AS1 in GBM and the underlying mechanisms. METHODS: The expression levels of GAS8-AS1 and NEAT1 in GBM patients and the healthy controls were measured by performing RT-qPCR. Diagnostic values of plasma GAS8-AS1 and NEAT1 for GBM were analyzed by performing ROC curve analysis with GBM patients as true positive cases and the healthy controls as true negative cases. Linear regression analysis was performed to study the correlation between the expression levels of GAS8-AS1 and NEAT1. The expression levels of GAS8-AS1 and NEAT1 in GBM cells were also determined by RT-qPCR. CCK-8 and transwell invasion assays were performed to detect the proliferation and invasion of GBM cells. Western blot assay was performed to detect the expression levels of ß-catenin, Axin2, c-myc, cyclin D1, and GAPDH in GBM cells. RESULTS: GAS8-AS1 was downregulated, while lncRNA NEAT1 was upregulated in the plasma of GBM patients. Altered expression levels of GAS8-AS1 and NEAT1 distinguished GBM patients from the healthy controls. The expression of GAS8-AS1 and NEAT1 was inversely correlated only in GBM patients. Overexpression of GAS8-AS1 reduced the expression levels of NEAT1 in GBM cells, while knock-down of GAS8-AS1 increased the expression levels of NEAT1. However, overexpression of NEAT1 showed no significant effects on the expression of GAS8-AS1. Knock-down of GAS8-AS1 promoted GBM cell proliferation and invasion and enhanced the activation of the Wnt/ß-catenin pathway. However, the effects of knock-down of GAS8-AS1 were alleviated by the knock-down of NEAT1. CONCLUSION: Overexpression of GAS8-AS1 inhibits GBM cell proliferation and invasion by downregulating NEAT1.
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Glioblastoma , ARN Largo no Codificante , Línea Celular Tumoral , Proliferación Celular/genética , Proteínas del Citoesqueleto , Regulación Neoplásica de la Expresión Génica , Glioblastoma/genética , Humanos , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , ARN Largo no Codificante/genéticaRESUMEN
Long non-coding RNAs (lncRNAs) represent a group of ncRNAs with more than 200 nucleotides. These RNAs can specifically regulate gene expression at both the transcriptional and the post-transcriptional levels, and increasing evidence indicates that they play vital roles in a variety of disease-related cellular processes. The lncRNA GAS8 antisense RNA 1 (GAS8-AS1, also known as C16orf3) is located in the second intron of GAS8 and has been reported to be both abnormally expressed in several diseases and closely correlated with many clinical characteristics. GAS8-AS1 has been shown to affect many biological functions, including cell proliferation, migration, invasiveness, and autophagy using several signaling pathways. In this review, we have summarized current studies on GAS8-AS1 roles in disease and discuss its potential clinical utility. GAS8-AS1 may be a promising biomarker for both diagnoses and prognoses, and a novel target for many disease therapies.
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Enfermedad/genética , ARN Largo no Codificante/genética , Animales , Biomarcadores , Diagnóstico , Humanos , PronósticoRESUMEN
Long non-coding RNAs (lncRNAs) play an essential regulatory role in multiple cancers. However, the role of lncRNAs in papillary thyroid carcinoma (PTC) is still unknown. Here, GAS8-AS1, a novel lncRNA that is significantly downregulated in PTC, was selected for further investigation. The roles of GAS8-AS1 in PTC cells were verified by gain- and loss-of-function experiments. The functional mechanism of GAS8-AS1 on the microRNA (miR)-187-3p/ATG5 axis and miR-1343-3p/ATG7 axis in PTC cells was evaluated using bioinformatics analysis, luciferase reporter assay, Cell Counting Kit-8 (CCK-8) assay, immunohistochemistry analysis, transmission electron microscopy, and immunofluorescence. We found that GAS8-AS1 was downregulated in PTC tissues and cell lines. In patients with PTC, low GAS8-AS1 expression was associated with higher tumor-node-metastasis (TNM) stage and lymph node metastasis (LNM). Functionally, GAS8-AS1 significantly promoted autophagy and inhibited PTC cell proliferation in vitro and promoted tumorigenesis in vivo. Mechanistically, GAS8-AS1 acted as a sponge of miR-187-3p and miR-1343-3p and upregulated ATG5 and ATG7 expression, respectively. The transcription factor ATF2 regulated GAS8-AS1 by binding to the GAS8-AS1 promoter. In conclusion, upregulation of ATF2 activated GAS8-AS1-promoted autophagy of PTC cells by sponging oncogenic miR-187-3p and miR-1343-3p and upregulating the expression of ATG5 and ATG7, respectively, making GAS8-AS1 a potential prognostic biomarker and therapeutic target for PTC.
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BACKGROUND: Early detection and diagnosis of ovarian cancer (OC) is complicated due to the concealment of the ovarian anatomical position and the lack of clinical manifestations and specific indicators of early OC. Therefore, it is urgent to study the pathogenesis of OC, especially the molecular mechanism. RESULTS: LncRNA GAS8-AS1 was decreased in OC tissues and cell lines, and high expression of GAS8-AS1 indicated a higher 5-year survival rate of OC patients. Overexpression of GAS8-AS1 suppressed growth of OC cells, while deletion of GAS8-AS1 promoted the progression of OC cells. Further data indicated GAS8-AS1 activated autophagy in OC cells. Functional experiments showed that 3-MA removed the inhibitory effect of GAS8-AS1 in OC cells. On the contrary, Rapamycin reversed the promoting effect of GAS8-AS1 in OC cells. Furthermore, GAS8-AS1 bound with Beclin1 and promoted its expression, and silencing of Beclin1 reversed the inhibitory role of GAS8-AS1 in OC progression. In vivo tumorigenesis assay showed GAS8-AS1 suppressed OC progression and activated Beclin1 mediated autophagy. CONCLUSION: Our study suggested GAS8-AS1 inhibited OC progression by activating autophagy via binding with Beclin1, and GAS8-AS1 might be a potential therapeutic target for OC clinical treatment.
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The Growth arrest specific 8 (GAS8) and its anti-sense transcript (GAS8-AS1) are located in a genomic region that is frequently mutated in breast cancer. These transcripts have established tumor suppressor effects in some human malignancies. In the current investigation, we aimed at identification of the role of GAS8 and GAS8-AS1 in breast cancer. We measured gene expression of GAS8 and GAS8-AS1 in paired tumoral and non-tumoral tissues obtained from 88 breast cancer patients by means of real time PCR. No significant differences were identified in expressions of GAS8 and GAS8-AS1 in tumoral samples compared with non-tumoral tissues (Fold changes = 1.53 and 1.71; P values = .28 and 0.14 respectively). Transcript levels of GAS8-AS1 was significantly correlated with estrogen receptor (ER) status (P = .01). Expression of GAS8 gene was associated with history of oral contraceptive use (P = .04). The similar expressions of GAS8 and GAS8-AS1 genes in tumoral and non-tumoral tissues of breast in spite of previous reports regarding their fundamental tumor suppressor roles in other tissues show that these genes are not involved in the pathogenesis of breast cancer. So, these genes have distinct roles in diverse tissues.
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Neoplasias de la Mama/genética , Proteínas del Citoesqueleto/genética , Proteínas de Neoplasias/genética , ARN Largo no Codificante/genética , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias de la Mama/patología , Línea Celular Tumoral , Anticonceptivos Orales/efectos adversos , Femenino , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Persona de Mediana EdadRESUMEN
BACKGROUND: Osteosarcoma (OS) is the most common type of primary bone tumor that mainly affects adolescents and young adults. The present study explored the role of lncRNA GAS8-AS1 in OS. METHODS: A total of 48 OS patients were selected from the 82 OS patients admitted by Luoyang Orthopedic Hospital of Henan Province between May 2010 and May 2013. Transient cell transfections, Transwell cell migration and invasion assay, RT-qPCR, and patient follow-up were carried out during the research. RESULTS: The results showed that GAS8-AS1 was downregulated, while UCA1 was upregulate in cancer tissues in comparison to adjacent non-cancer tissues of OS patients. GAS8-AS1 was not affected by clinical stage. Follow-up study showed that downregulated GAS8-AS1 in cancer tissues was closely correlated with poor survival. GAS8-AS1 and UCA1 were inversely correlated in cancer tissues. Overexpression of UCA1 failed to affect the expression of GAS8-AS1, while overexpression of GAS8-AS1 led to downregulated expression of UCA1 in OS cells, while the molecular mediators between these two lncRNAs are unknown. Overexpression of GAS8-AS1 did not affect OS cell proliferation but significantly inhibited cancer cell migration and invasion. Overexpression of UCA1 promoted the migration and invasion of OS cells and attenuated the effects of overexpressing GAS8-AS1. CONCLUSIONS: Therefore, GAS8-AS1 may inhibit OS cell migration and invasion by downregulating oncogenic UCA1.
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Neoplasias Óseas/metabolismo , Movimiento Celular/fisiología , Regulación hacia Abajo/fisiología , Osteosarcoma/metabolismo , ARN Largo no Codificante/biosíntesis , Adolescente , Adulto , Biomarcadores de Tumor/antagonistas & inhibidores , Biomarcadores de Tumor/biosíntesis , Neoplasias Óseas/patología , Línea Celular Tumoral , Niño , Femenino , Estudios de Seguimiento , Humanos , Masculino , Invasividad Neoplásica/patología , Osteosarcoma/patología , ARN Largo no Codificante/antagonistas & inhibidores , Adulto JovenRESUMEN
AIM: To evaluate the expression of the growth arrest-specific 8 (GAS8) and its antisense (GAS8-AS1) in gastric cancer. BACKGROUND: GAS8 exists in a genomic region that is recurrently deleted in breast and prostate cancer. This gene contains a long non-coding RNA, namely GAS8-AS1 whose roles in the regulation of GAS8 has been reported in hepatocytes. GAS8-AS1 has also been regarded as a putative tumor suppressor gene in papillary thyroid cancer and hepatocellular carcinoma. METHODS: In the present study, we evaluated expression levels of GAS8 and GAS8-AS1 in 30 gastric cancer tissues and their corresponding adjacent non-cancerous tissues (ANCTs). RESULTS: GAS8 was significantly down-regulated in tumor tissues compared to ANCTs (Expression ratio=0.29, p<0.001). Although the expression of GAS8-AS1 was higher in tumor tissues compared to ANCTs (Expression ratio=2.15), it did not reach the level of significance (p=0.12). GAS8 expression was associated with the site of the primary tumor (p=0.01). GAS8-AS1 expression was significantly higher in tumors with lymphatic/ vascular invasion compared with those without lymphatic/ vascular invasion (p=0.03). Significant pairwise correlations were detected between expression levels of GAS8 and GAS8-AS1 in tumor tissues and ANCTs. Based on the results of the ROC curve, the diagnostic power of transcript levels of GAS8 in gastric tissues was estimated to be 76%. CONCLUSION: The current study underscores the roles of GAS8 and GAS8-AS1 in gastric carcinogenesis and warrants future functional studies to unravel the underlying mechanism of such contribution.
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LncRNA GAS8-AS1 inhibits thyroid carcinoma, while its role in colorectal cancer (CRC) is unknown. In the present study we found that plasma GAS8-AS1 was upregulated in early stage CRC patients, and downregulation of GAS8-AS1 effectively distinguished CRC patients from healthy controls. LncRNA AFAP1-AS1 was upregulated in CRC patients and was inversely correlated with GAS8-AS1 only in CRC patients but not in healthy controls. GAS8-AS1 overexpression mediated the downregulation of AFAP1-AS1 in colon cancer cells, while AFAP1-AS1 overexpression did not significantly affect GAS8-AS1 expression. Expression level of GAS8-AS1 decreased, while expression level of AFAP1-AS1 increased with the increase of primary tumor diameters. GAS8-AS1 overexpression led to inhibited, while AFAP1-AS1 overexpression led to promoted proliferation of CRC cells, and AFAP1-AS1 overexpression reduced the inhibitory effects of GAS8-AS1 overexpression on cancer cell proliferation. Therefore, GAS8-AS may inhibit CRC cell proliferation by downregulating AFAP1-AS1.
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Neoplasias Colorrectales/patología , Regulación hacia Abajo , Proteínas de Neoplasias/genética , ARN Largo no Codificante/genética , Adulto , Anciano , Línea Celular Tumoral , Proliferación Celular , Neoplasias Colorrectales/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Carga TumoralRESUMEN
Expressions of the Growth arrest specific 8 (GAS8) and its naturally occurring anti-sense RNA (GAS8-AS1) have been assessed in tumoral tissues of different origins. However, their association with immune-related disorders has been poorly understood. In the current study, we evaluated expression levels of these genes in 50 relapsing-remitting multiple sclerosis (RRMS) patients compared with age- and sex-matched controls. Expressions of both genes were significantly higher in total MS patients compared with controls (P = 0.001 and P < 0.0001 respectively). The difference in GAS8 expression was also significant in total female patients and females aged less than 50 when compared with the corresponding control subjects (P = 0.002 and 0.006 respectively). GAS8-AS1 was higher in male patients in both age-based subgroups compared with the corresponding healthy subjects (P < 0.0001). Expressions of both genes were inversely correlated with age of male study participants but no other subgroups. GAS8-AS1 transcript levels had 99.6% accuracy in diagnosis of disease status in male subjects. The current study shows significance of GAS8 and GAS8-AS1 in the pathogenesis of MS and the putative role of GAS8-AS1 as a diagnostic biomarker in a subset of patients.
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Proteínas del Citoesqueleto/genética , Esclerosis Múltiple/diagnóstico , Esclerosis Múltiple/genética , Proteínas de Neoplasias/genética , ARN Largo no Codificante/genética , Adulto , Edad de Inicio , Estudios de Casos y Controles , Femenino , Humanos , Masculino , Persona de Mediana Edad , Esclerosis Múltiple/metabolismo , Curva ROCRESUMEN
The aim of the present study was to investigate the potential role of GAS8 antisense RNA 1 (GAS8-AS1) in papillary thyroid carcinoma (PTC). PcDNA3.1-GAS8-AS1 and si-GAS8-AS1, miR-135b-5p mimic and si-CCND2 were transfected into PTC cells. Cell proliferation was evaluated by Cell Counting Kit-8 (CCK-8). QRT-PCR was used to determine expressions of GAS8-AS1, miR-135b-5p, and CCND2, and Western blot were detected protein level of CCND2. The miRNA target gene prediction site TargetScan was used to predict potential targets of GAS8-AS1 and miR-135b-5p. Cell cycle progression was analyzed by flow cytometry. We found that GAS8-AS1 was down-regulated in PTC cell lines and inhibited proliferation and cycle of PTC cell. GAS8-AS1 directly targets miR-135b-5p, and GAS8-AS1 could regulate a downstream target of miR-135b-5p, Cyclin G2 (CCNG2), in an miR-135b-5p-mediated manner. In addition, we also proved that overexpressed GAS8-AS1 inhibited tumor formation in vivo GAS8-AS1 suppresses PTC cell growth through the miR-135b-5p/CCND2 axis.
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Ciclina D2/genética , Proteínas del Citoesqueleto/genética , Regulación Neoplásica de la Expresión Génica , MicroARNs/genética , Proteínas de Neoplasias/genética , ARN Largo no Codificante/genética , Cáncer Papilar Tiroideo/genética , Neoplasias de la Tiroides/genética , Animales , Secuencia de Bases , Sitios de Unión , Ciclo Celular/genética , Línea Celular Tumoral , Proliferación Celular , Ciclina D2/metabolismo , Proteínas del Citoesqueleto/antagonistas & inhibidores , Proteínas del Citoesqueleto/metabolismo , Humanos , Masculino , Ratones , Ratones Desnudos , MicroARNs/metabolismo , Proteínas de Neoplasias/antagonistas & inhibidores , Proteínas de Neoplasias/metabolismo , ARN Largo no Codificante/metabolismo , Transducción de Señal , Cáncer Papilar Tiroideo/metabolismo , Cáncer Papilar Tiroideo/patología , Cáncer Papilar Tiroideo/terapia , Neoplasias de la Tiroides/metabolismo , Neoplasias de la Tiroides/patología , Neoplasias de la Tiroides/terapia , Carga Tumoral , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Long noncoding RNAs (lncRNAs) are vital players in cancers, including hepatocellular carcinoma (HCC). We previously identified an lncRNA, GAS8-AS1, that is located in intron 2 of GAS8 However, its involvement in HCC is still largely unknown. In this study, we report that both GAS8-AS1 and its host gene GAS8 act as HCC tumor suppressors. We found that expression of GAS8-AS1 or GAS8 is significantly decreased in HCC tissues and is associated with a poor prognosis among HCC patients. Interestingly, lncRNA GAS8-AS1 could promote GAS8 transcription. We detected a CpG island in the GAS8 promoter, but lncRNA GAS8-AS1 did not affect DNA methylation at this GAS8 promoter site. Moreover, we identified two GAS8-AS1-interacting proteins, mixed-lineage leukemia 1 (MLL1), a histone 3 Lys-4 (H3K4) methyltransferase, and its partner WD-40 repeat protein 5 (WDR5). RNA pulldown, ChIP, and RNA immunoprecipitation assays revealed that GAS8-AS1 is required for maintaining the GAS8 promoter in an open chromatin state by recruiting the MLL1/WDR5 complex and for enhancing RNA polymerase II activity and GAS8 transcription. Of note, GAS8-AS1-dependent GAS8 hyperactivation inhibited malignant transformation of hepatocytes. Our results provide important insights into how lncRNA GAS8-AS1 suppresses HCC development and suggest potential strategies for treating patients with liver cancer.
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Carcinoma Hepatocelular/genética , Proteínas del Citoesqueleto/genética , Neoplasias Hepáticas/genética , Proteínas de Neoplasias/genética , ARN Largo no Codificante/genética , Animales , Carcinogénesis , Carcinoma Hepatocelular/metabolismo , Línea Celular Tumoral , Transformación Celular Neoplásica , Islas de CpG , Proteínas del Citoesqueleto/metabolismo , Metilación de ADN , Epigénesis Genética , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias Hepáticas/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Proteínas de Neoplasias/metabolismo , Regiones Promotoras Genéticas , ARN Largo no Codificante/metabolismoRESUMEN
PURPOSE: The long non-coding RNA GAS8 antisense RNA 1 (lncRNA GAS8-AS1) is a tumor suppressor in papillary thyroid cancer (PTC), but the mechanisms underlying how GAS8-AS1 regulates PTC biology remain unclear. Here, we evaluated the molecular function of GAS8-AS1 in regulating autophagy in PTC cell lines. METHODS: GAS8-AS1 was overexpressed and knocked down in PTC cell lines by transfecting with expression plasmids or short interfering RNAs (siRNAs). Cell proliferation was evaluated using the Cell Counting Kit-8 (CCK-8). qRT-PCR and western blot were used to determine changes in expression of autophagy-related genes. Autophagy was evaluated by immunofluorescence and transmission electron microscopy. RESULTS: Relative GAS8-AS1 expression was lower in the PTC cell lines, TPC1 and BCPAP, compared to a normal thyroid cell line. Overexpression of GAS8-AS1 inhibited proliferation, significantly increased the ratio of LC3-II/LC3-I, and reduced p62 expression, whereas GAS8-AS1 knockdown demonstrated opposite effects. In GAS8-AS1 overexpressing cell lines, LC3 immunofluorescence staining demonstrated increased punctate aggregates of LC3 staining, and transmission electron microscopy revealed increased numbers of autophagosomes. Autophagy-related gene 5 (ATG5) was markedly upregulated by GAS8-AS1 overexpression and downregulated by GAS8-AS1 knockdown. Finally, silencing of ATG5 attenuated autophagy activation and rescued the inhibition of cell proliferation caused by GAS8-AS1. CONCLUSIONS: In PTC cell lines, GAS8-AS1 inhibited proliferation, activated autophagy, and increased ATG5 expression. Downregulation of ATG5 reversed GAS8-AS1-mediated activation of autophagy leading to cell death, revealing a novel mechanism of the GAS8-AS1-ATG5 axis in PTC cell lines. This provided a new experimental basis to explore the effects of lncRNA on autophagy in the treatment of thyroid cancer.
Asunto(s)
Proteína 5 Relacionada con la Autofagia/metabolismo , Autofagia/fisiología , Carcinoma Papilar/metabolismo , Proliferación Celular/fisiología , Proteínas del Citoesqueleto/metabolismo , Proteínas de Neoplasias/metabolismo , ARN Largo no Codificante/metabolismo , Neoplasias de la Tiroides/metabolismo , Carcinoma Papilar/patología , Línea Celular Tumoral , Humanos , Neoplasias de la Tiroides/patologíaRESUMEN
Objective To investigate the expression level and clinical significance of long non-coding RNA(LncRNA) growth arrest specific gene-antisense 1(GAS8-AS1) in papillary thyroid microcarcinoma(PTMC) patients. Methods We investigated the expression profile of GAS8-AS1 in tissue samples of patients with PTMC as well as nodular goiter(NG) by quantitative real-time polymerase chain reaction(RT-qPCR). Results GAS8-AS1 in cancer tissue was down-regulated in PTMC patients compared with adjacent thyroid tissue and NG samples(P<0.05). Lower level of GAS8-AS1 was also correlated with central cervical lymph node metastasis(CLNM, P<0.05). The area under the ROC curve for GAS8-AS1 was up to 0.717 3 in CLNM prediction(P<0.05). Conclusion GAS8-AS1 may act as a potential biomarker for PTC diagnosis and CLNM prediction.
RESUMEN
Primary ciliary dyskinesia (PCD) is an autosomal recessive disease characterized by chronic respiratory infections of the upper and lower airways, hypofertility, and, in approximately half of the cases, situs inversus. This complex phenotype results from defects in motile cilia and sperm flagella. Among the numerous genes involved in PCD, very few-including CCDC39 and CCDC40-carry mutations that lead to a disorganization of ciliary axonemes with microtubule misalignment. Focusing on this particular phenotype, we identified bi-allelic loss-of-function mutations in GAS8, a gene that encodes a subunit of the nexin-dynein regulatory complex (N-DRC) orthologous to DRC4 of the flagellated alga Chlamydomonas reinhardtii. Unlike the majority of PCD patients, individuals with GAS8 mutations have motile cilia, which, as documented by high-speed videomicroscopy, display a subtle beating pattern defect characterized by slightly reduced bending amplitude. Immunofluorescence studies performed on patients' respiratory cilia revealed that GAS8 is not required for the proper expression of CCDC39 and CCDC40. Rather, mutations in GAS8 affect the subcellular localization of another N-DRC subunit called DRC3. Overall, this study, which identifies GAS8 as a PCD gene, unveils the key importance of the corresponding protein in N-DRC integrity and in the proper alignment of axonemal microtubules in humans.