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1.
Cell Prolif ; 52(4): e12614, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30983072

RESUMO

OBJECTIVES: To reveal the role of circular RNA (circRNA) DOCK1 (circDOCK1) as a potential biomarker and therapeutic target and its competing endogenous RNA mechanism in bladder carcinoma (BC). METHODS: The next-generation sequencing (NGS) technology was introduced to screen the circRNA expression profiles of BC using microarray. qPCR and Western blots assay were employed to measure the gene expression in different groups. Cell counting kit-8, EdU and transwell assays were applied to detect the cell viability, proliferation and migration potential, respectively. Luciferase reporter assay was used to test the binds between hsa-miR-132-3p/Sox5. Xenografted tumour growth of nude mice was performed to test the role of circDOCK1 in vivo. RESULTS: CircDOCK1 was upregulated in BC tissues and cell lines. Repression of circDOCK1 reduced cell viability, inhibited cell proliferation and curbed the cell migration potential of BC cell. CircDOCK1 played its role via regulation of circDOCK1/hsa-miR-132-3p/Sox5 pathway in BC cells. Suppression circDOCK1 inhibited the tumour growth in vivo. CONCLUSION: In this study, we revealed that circDOCK1 affected the progression of BC via modulation of circDOCK1/hsa-miR-132-3p/Sox5 pathway both in vitro and in vivo and providing a potential biomarker and therapeutic targets for BC.


Assuntos
MicroRNAs/genética , Fatores de Transcrição SOXD/genética , Transdução de Sinais/genética , Neoplasias da Bexiga Urinária/genética , Proteínas rac de Ligação ao GTP/genética , Animais , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Sobrevivência Celular/genética , Progressão da Doença , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , RNA/genética , Regulação para Cima/genética , Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/patologia
2.
J Assist Reprod Genet ; 36(4): 759-768, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30863997

RESUMO

PURPOSE: Male infertility is a multifactorial syndrome encompassing a wide variety of disorders. A previous Chinese genome-wide single-nucleotide polymorphism (SNP) association studies have identified four SNPs (rs12097821 in PRMT6 gene, rs2477686 in PEX10 gene, rs6080550 in SIRPA-SIRPG, and rs10842262 in SOX5 gene) as being significantly associated with risk factors for nonobstructive azoospermia (NOA). However, the results were not fully repeated in later studies, which calls for further investigations. METHODS: We here performed a case-control study in a central Chinese population to explore the association between the four SNPs and male infertility, which included 631 infertile men (NOA and oligozoospermia) and 720 healthy fertile men. The genotyping was performed using the polymerase chain reaction-restriction fragment length polymorphism and confirmed by sequencing. RESULTS: The results showed that rs12097821 and rs10842262 were strongly associated with the risk of NOA but not total male infertility or oligozoospermia, while rs2477686 and rs6080550 were not associated with the risk of total male infertility, NOA, or oligozoospermia. To improve the statistical strength, a meta-analysis was conducted. The results suggested that rs2477686, rs6080550, and rs10842262 were significantly associated with male infertility, especially with NOA, while rs12097821 was only found to be associated with total male infertility. CONCLUSIONS: Collectively, the rs2477686, rs6080550, and rs10842262 may indeed be the genetic risk factors for NOA, which requires further investigation using larger independent sets of samples in different ethnic populations.


Assuntos
Antígenos de Diferenciação/genética , Azoospermia/genética , Peroxinas/genética , Receptores Citoplasmáticos e Nucleares/genética , Receptores Imunológicos/genética , Fatores de Transcrição SOXD/genética , Adulto , Azoospermia/patologia , Estudos de Associação Genética , Predisposição Genética para Doença , Humanos , Infertilidade Masculina , Masculino , Polimorfismo de Nucleotídeo Único/genética
3.
Biochem Biophys Res Commun ; 509(2): 603-610, 2019 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-30606481

RESUMO

MicroRNA-135a-5p has been reported to play a potential role in the generation of new neurons. However, the underlying targets of miR-135a-5p in regulating neuronal differentiation have been poorly understood. Our study recently has uncovered that Sox6 and CD44 genes were significantly downregulated during neuronal differentiation of P19 cells, a multipotent cell type. We then found that Sox6 directly bound to the promoter of CD44. Importantly, we identified Sox6 as a direct target of miR-135a-5p. Additionally, we demonstrated that miR-135a-5p is crucial for the neuronal differentiation of P19 cells. More significantly, we found that Sox6 overexpression could overturn miR-135a-5p-mediated neuronal differentiation and dendrite development. In conclusion, these findings indicated that miR-135a-5p/Sox6/CD44 axis provides an important molecular target mechanism for neurodifferentiation.


Assuntos
Células-Tronco de Carcinoma Embrionário/patologia , Receptores de Hialuronatos/genética , MicroRNAs/genética , Neurogênese , Fatores de Transcrição SOXD/genética , Animais , Linhagem Celular Tumoral , Células-Tronco de Carcinoma Embrionário/citologia , Células-Tronco de Carcinoma Embrionário/metabolismo , Regulação Neoplásica da Expressão Gênica , Células HEK293 , Humanos , Receptores de Hialuronatos/metabolismo , Camundongos , Fatores de Transcrição SOXD/metabolismo , Transdução de Sinais
4.
Cell Prolif ; 52(3): e12573, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30667104

RESUMO

OBJECTIVES: Long non-coding RNAs (LncRNAs) play important roles in epigenetic regulatory function during the development processes. In this study, we found that through alternative splicing, LncRNA C130071C03Riken variants Riken-201 (Riken-201) and Riken-203 (Riken-203) are both expressed highly in brain, and increase gradually during neural differentiation. However, the function of Rik-201 and Rik-203 is unknown. MATERIALS AND METHODS: Embryonic stem cells (ESCs); RNA sequencing; gene expression of mRNAs, LncRNAs and miRNAs; over-expression and RNA interference of genes; flow cytometry; real-time quantity PCR; and Western blot were used in the studies. RNA pull-down assay and PCR were employed to detect any miRNA that attached to Rik-201 and Rik-203. The binding of miRNA with mRNA of Sox6 was presented by the luciferase assay. RESULTS: Repression of Rik-201 and Rik-203 inhibited neural differentiation from mouse embryonic stem cells. Moreover, Rik-201 and Rik-203 functioned as the competing endogenous RNA (ceRNA) to repress the function of miR-96 and miR-467a-3p, respectively, and modulate the expression of Sox6 to further regulate neural differentiation. Knockout of the Rik-203 and Rik-201 induced high ratio of brain developmental retardation. Further we found that C/EBPß might potentially activated the transcription of Rik-201 and Rik-203. CONCLUSIONS: These findings identify the functional role of Rik-201 and Rik-203 in facilitating neural differentiation and further brain development, and elucidate the underlying miRNAs-Sox6-associated molecular mechanisms.


Assuntos
MicroRNAs/genética , MicroRNAs/metabolismo , Neurogênese/genética , RNA Longo não Codificante/genética , Fatores de Transcrição SOXD/genética , Processamento Alternativo , Animais , Sequência de Bases , Encéfalo/anormalidades , Encéfalo/embriologia , Encéfalo/metabolismo , Diferenciação Celular/genética , Epigênese Genética , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/metabolismo , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Gravidez , RNA Longo não Codificante/antagonistas & inibidores , RNA Interferente Pequeno/genética
5.
Pathol Res Pract ; 215(2): 335-342, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30580904

RESUMO

BACKGROUND: The aim of the study was to measure the expression of microRNA (miR)-181b in patients with lung cancer, investigate its biological function and elucidate the underlying mechanisms associated with the development of lung cancer. METHODS: miR-181b expression in tissues was measured via RT-qPCR. After A549 cells were transfected with miR-181b mimic or si-Sox6, the proliferation, migration and cell cycle distribution of A549 were evaluated using cell counting kit-8 assay, transwell assay and flow cytometry. The levels of cell cycle-related proteins and Sox6 were analyzed by western blotting. Gene targets of miR-181b were predicted via bioinformatics analysis and verified using a dual-luciferase reporter gene assay. RESULTS: Expression of miR-181b was significantly downregulated in lung cancer tissues (P < 0.05), and was inversely correlated with the degree of cell differentiation and clinical stages of lung cancer (both P < 0.05). Additionally, the expression of miR-181b was significantly lower in adenocarcinoma compared with squamous cell carcinoma in the lungs (P < 0.05). Overexpression of miR-181b significantly decreased the protein level of Sox6 and significantly suppressed the cell proliferation and metastasis (both P < 0.05); this effect was also observed in A549 cells transfected with si-Sox6. The luciferase activity of a Sox6 3'-untranslated region-based reporter construct was significantly lower when transfected with miR-181b (P < 0.05), which suggests that Sox6 is a direct target of miR-181b. CONCLUSION: The results of the present study suggest that miR-181b may function as a tumor inhibitor in the development of lung cancer via targeting Sox6 to decrease the proliferation and metastasis of lung cancer cells.


Assuntos
Regulação Neoplásica da Expressão Gênica/genética , Neoplasias Pulmonares/patologia , MicroRNAs/genética , Fatores de Transcrição SOXD/biossíntese , Células A549 , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Proliferação de Células/genética , Humanos , Neoplasias Pulmonares/genética , MicroRNAs/metabolismo , Invasividade Neoplásica/genética , Fatores de Transcrição SOXD/genética , Carcinoma de Pequenas Células do Pulmão/genética , Carcinoma de Pequenas Células do Pulmão/patologia
6.
Gene ; 678: 343-348, 2018 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-30098430

RESUMO

It has been demonstrated that microRNAs (miRNAs) play important roles in the regulation of melanogenesis and hair color in mammals. Short tandem target mimic (STTM) has been used to effectively block small RNA functions in plants and animals. We previously showed that miR508-3p plays a functional role in regulating melanogenesis in alpaca melanocytes by directly targeting microphthalmia (MITF). To verify the effect of miR-508-3p function on melanogenesis in alpaca melanocytes, miR-508-3p was blocked using STTM technology in the present research. miR508-3p was predicted to target the gene encoding SRY-box6 (SOX6) by bioinformatics. The luciferase reporter assay indicated that miR508-3p regulates SRY-box6 (SOX6) expression by targeting its 3'UTR. Here, STTM-miR508-3p overexpression in alpaca melanocytes blocked the expression of miR-508-3p and up-regulated SOX6 expression at both the mRNA and protein levels, resulting in increasing the expression of key melanogenic genes, including cAMP responsive element (CRE) binding protein (CREB), MITF, tyrosinase (TYR) and tyrosinase-related protein 1 and 2 (TYRP1 and TYRP2). STTM-miR508-3p overexpression in melanocytes also resulted in increased melanin production, including total alkali soluble melanogenesis (ASM), eumelanogenesis (EM) and pheomelanogenesis (PM). Additionally, we identified melanin granules in alpaca melanocytes transfected with STTM-miR508-3p under Fontana-Masson staining. These results suggest that STTM-miR508-3p could up-regulate melanogenesis by effectively blocking miR508-3p.


Assuntos
Camelídeos Americanos/genética , Inativação Gênica , Melaninas/biossíntese , MicroRNAs/genética , Regiões 3' não Traduzidas , Animais , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Melanócitos/metabolismo , Fatores de Transcrição SOXD/genética
7.
Dev Biol ; 442(2): 262-275, 2018 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-30071218

RESUMO

In anamniotes, somite compartimentalization in the lateral somitic domain leads simultaneously to myotome and dermomyotome formation. In the myotome, Xenopus Sox5 is co-expressed with Myod1 in the course of myogenic differentiation. Here, we studied the function of Sox5 using a Myod1-induced myogenic transcription assay in pluripotent cells of animal caps. We found that Sox5 enhances myogenic transcription of muscle markers Des, Actc1, Ckm and MyhE3. The use of chimeric transactivating or transrepressive Sox5 proteins indicates that Sox5 acts as a transrepressor and indirectly stimulates myogenic transcription except for the slow muscle-specific genes Myh7L, Myh7S, Myl2 and Tnnc1. We showed that this role is shared by Sox6, which is structurally similar to Sox5, both belonging to the SoxD subfamily of transcription factors. Moreover, Sox5 can antagonize the inhibitory function of Meox2 on myogenic differentiation. Meox2 which is a dermomyotome marker, represses myogenic transcription in Myod-induced myogenic transcription assay and in Nodal5-induced mesoderm from animal cap assay. The inhibitory function of Meox2 and the pro-myogenic function of Sox5 were confirmed during Xenopus normal development by the use of translation-blocking oligomorpholinos and dexamethasone inducible chimeric Sox5 and Meox2 proteins. We have therefore identified a new function for SoxD proteins in muscle cells, which can indirectly enhance myogenic transcription through transrepression, in addition to the previously identified function as a direct repressor of slow muscle-specific genes.


Assuntos
Fatores de Transcrição SOXD/metabolismo , Proteínas de Xenopus/metabolismo , Animais , Diferenciação Celular/fisiologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Mesoderma/metabolismo , Células Musculares/metabolismo , Desenvolvimento Muscular/genética , Músculos/metabolismo , Proteína MyoD/genética , Proteína MyoD/metabolismo , Fatores de Transcrição SOXD/genética , Somitos/metabolismo , Ativação Transcricional/fisiologia , Proteínas de Xenopus/genética , Xenopus laevis
8.
Mol Cells ; 41(6): 575-581, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29890823

RESUMO

Postmenopausal osteoporosis (PMOP) is a common systemic skeletal disease characterized by reduced bone mass and microarchitecture deterioration. Although differentially expressed SOX5 has been found in bone marrow from ovariectomized mice, its role in osteogenic differentiation in human mesenchymal stem cells (hMSCs) from bone marrow in PMOP remains unknown. In this study, we investigated the biological function of SOX5 and explore its molecular mechanism in hMSCs from patients with PMOP. Our findings showed that the mRNA and protein expression levels of SOX5 were upregulated in hMSCs isolated from bone marrow samples of PMOP patients. We also found that SOX5 overexpression decreased the alkaline phosphatase (ALP) activity and the gene expression of osteoblast markers including Collagen I, Runx2 and Osterix, which were increased by SOX5 knockdown using RNA interference. Furthermore, TNF-α notably upregulated the SOX5 mRNA expression level, and SOX5 knockdown reversed the effect of TNF-α on osteogenic differentiation of hMSCs. In addition, SOX5 overexpression increased Kruppel-like factor 4 (KLF4) gene expression, which was decreased by SOX5 silencing. KLF4 knockdown abrogated the suppressive effect of SOX5 overexpression on osteogenic differentiation of hMSCs. Taken together, our results indicated that TNF-α-induced SOX5 upregulation inhibited osteogenic differentiation of hMSCs through KLF4 signal pathway, suggesting that SOX5 might be a novel therapeutic target for PMOP treatment.


Assuntos
Fatores de Transcrição Kruppel-Like/metabolismo , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Fatores de Transcrição SOXD/biossíntese , Fator de Necrose Tumoral alfa/farmacologia , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Estudos de Casos e Controles , Diferenciação Celular/fisiologia , Feminino , Expressão Gênica , Humanos , Fatores de Transcrição Kruppel-Like/genética , Células-Tronco Mesenquimais/efeitos dos fármacos , Camundongos , Pessoa de Meia-Idade , Osteoporose Pós-Menopausa/tratamento farmacológico , Osteoporose Pós-Menopausa/genética , Osteoporose Pós-Menopausa/metabolismo , Osteoporose Pós-Menopausa/patologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Fatores de Transcrição SOXD/genética , Fatores de Transcrição SOXD/metabolismo , Transdução de Sinais , Regulação para Cima/efeitos dos fármacos
9.
PLoS Genet ; 14(4): e1007260, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29621239

RESUMO

Mechanisms generating diverse cell types from multipotent progenitors are fundamental for normal development. Pigment cells are derived from multipotent neural crest cells and their diversity in teleosts provides an excellent model for studying mechanisms controlling fate specification of distinct cell types. Zebrafish have three types of pigment cells (melanocytes, iridophores and xanthophores) while medaka have four (three shared with zebrafish, plus leucophores), raising questions about how conserved mechanisms of fate specification of each pigment cell type are in these fish. We have previously shown that the Sry-related transcription factor Sox10 is crucial for fate specification of pigment cells in zebrafish, and that Sox5 promotes xanthophores and represses leucophores in a shared xanthophore/leucophore progenitor in medaka. Employing TILLING, TALEN and CRISPR/Cas9 technologies, we generated medaka and zebrafish sox5 and sox10 mutants and conducted comparative analyses of their compound mutant phenotypes. We show that specification of all pigment cells, except leucophores, is dependent on Sox10. Loss of Sox5 in Sox10-defective fish partially rescued the formation of all pigment cells in zebrafish, and melanocytes and iridophores in medaka, suggesting that Sox5 represses Sox10-dependent formation of these pigment cells, similar to their interaction in mammalian melanocyte specification. In contrast, in medaka, loss of Sox10 acts cooperatively with Sox5, enhancing both xanthophore reduction and leucophore increase in sox5 mutants. Misexpression of Sox5 in the xanthophore/leucophore progenitors increased xanthophores and reduced leucophores in medaka. Thus, the mode of Sox5 function in xanthophore specification differs between medaka (promoting) and zebrafish (repressing), which is also the case in adult fish. Our findings reveal surprising diversity in even the mode of the interactions between Sox5 and Sox10 governing specification of pigment cell types in medaka and zebrafish, and suggest that this is related to the evolution of a fourth pigment cell type.


Assuntos
Linhagem da Célula , Melanócitos/metabolismo , Oryzias/genética , Pigmentação/genética , Fatores de Transcrição SOXD/genética , Fatores de Transcrição SOXE/genética , Proteínas de Peixe-Zebra/genética , Peixe-Zebra/genética , Alelos , Animais , Regulação da Expressão Gênica no Desenvolvimento , Melanócitos/citologia , Crista Neural/metabolismo , Fatores de Transcrição SOXD/metabolismo , Fatores de Transcrição SOXE/metabolismo , Proteínas de Peixe-Zebra/metabolismo
10.
Mol Med Rep ; 17(5): 6803-6811, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29512775

RESUMO

Transcription factor SOX6 (SOX6) has been reported to serve essential roles in numerous types of cancers. However, the expression and functions of SOX6 in osteosarcoma (OS) have not been analyzed. In the present study, the patterns of SOX6 expression in OS cell lines and tissues were investigated by reverse transcription­quantitative polymerase chain reaction and western blotting. The results of the present study revealed that SOX6 was notably downregulated in OS tissues and cell lines. Subsequently, gain­ and loss­of­function studies demonstrated that SOX6 inhibited OS cell migration and invasion. In addition, SOX6 may have suppressed epithelial­mesenchymal transition via twist­related protein 1 (TWIST1) modulation. Chromatin immunoprecipitation (ChIP), quantitative ChIP and dual luciferase activity assays were used to confirm the binding of SOX6 to the promoter region of TWIST1. Additionally, colony formation assays and Cell Counting Kit­8 assays demonstrated that SOX6 suppressed cell proliferation. The findings of the present study indicated that SOX6 serves as a tumor suppressor in OS and may be a potential therapeutic target for OS.


Assuntos
Neoplasias Ósseas/metabolismo , Movimento Celular , Transição Epitelial-Mesenquimal , Regulação Neoplásica da Expressão Gênica , Proteínas Nucleares/biossíntese , Osteossarcoma/metabolismo , Fatores de Transcrição SOXD/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Proteína 1 Relacionada a Twist/biossíntese , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Humanos , Invasividade Neoplásica , Proteínas Nucleares/genética , Osteossarcoma/genética , Osteossarcoma/patologia , Fatores de Transcrição SOXD/genética , Proteínas Supressoras de Tumor/genética , Proteína 1 Relacionada a Twist/genética
11.
Gene ; 655: 65-70, 2018 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-29477873

RESUMO

Lamb-Shaffer syndrome (OMIM: 616803) is a neurodevelopmental disorder characterized by developmental delay, mild to moderate intellectual disability, speech delay, and mild characteristic facial appearance caused by SOX5 haploinsufficiency on chromosome 12p12.1. There are clinical variabilities among the patients with genomic alterations, such as intragenic deletions, a point mutation, and a chromosomal translocation of t(11;12)(p13;p12.1), in SOX5. We report herein a 5-year-old Japanese male with a de novo balanced reciprocal translocation t(12;20)(p12.1;p12.3) presenting a mild intellectual disability, speech delay, characteristic facial appearance, and autistic features. We determined the translocation breakpoints of the patient to be in intron 4 of SOX5 and the intergenic region in 20p12.3 via FISH and nucleotide sequence analyses. Thus, the present patient has SOX5 haploinsufficiency affecting 2 long forms of SOX5 and is the second reported case of Lamb-Shaffer syndrome caused by a de novo balanced reciprocal translocation. This report confirmed that haploinsufficiency of the 2 long forms of SOX5 presents common clinical features, including mild intellectual disability and autistic features, which could be useful for the clinical diagnosis of Lamb-Shaffer syndrome.


Assuntos
Anormalidades Múltiplas/genética , Anormalidades Múltiplas/patologia , Cromossomos Humanos Par 12 , Cromossomos Humanos Par 20 , Haploinsuficiência , Fatores de Transcrição SOXD/genética , Translocação Genética , Transtorno Autístico/genética , Transtorno Autístico/patologia , Pré-Escolar , Cromossomos Humanos Par 12/genética , Cromossomos Humanos Par 20/genética , Análise Mutacional de DNA , Deficiências do Desenvolvimento/genética , Deficiências do Desenvolvimento/patologia , Humanos , Deficiência Intelectual/genética , Deficiência Intelectual/patologia , Masculino
12.
Eur J Pharmacol ; 823: 65-71, 2018 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-29355560

RESUMO

Prostate cancer is one of the most severe malignancies in men, and many genes and non-coding RNAs, included microRNAs (miRs), have been demonstrated to regulate prostate cancer progression. In the present study, we investigated the role of miR-671 in prostate cancer cell proliferation. We found that miR-671 was significantly upregulated in human prostate cancer tissues and cells. miR-671 overexpression promoted prostate cancer cell proliferation, while its downregulation inhibited prostate cancer cell proliferation, as determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays, colony formation assays, soft agar growth assays, and bromodeoxyuridine (BrdU) incorporation assays. miR-671 directly targets the 3' untranslated region (UTR) of the tumor suppressor SOX6 (encoding SRY (sex determining region Y)-box 6) to inhibit its expression. Double knockdown of miR-671 and SOX6 promoted PC3 cell proliferation, suggesting that miR-671 promotes prostate cancer cell proliferation by inhibiting SOX6.


Assuntos
MicroRNAs/genética , Neoplasias da Próstata/patologia , Fatores de Transcrição SOXD/genética , Regiões 3' não Traduzidas/genética , Sequência de Bases , Linhagem Celular Tumoral , Proliferação de Células/genética , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino
13.
Cell Mol Life Sci ; 75(1): 67-79, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-28864883

RESUMO

Transcriptional regulation of proteins involved in neuronal polarity is a key process that underlies the ability of neurons to transfer information in the central nervous system. The Collapsin Response Mediator Protein (CRMP) family is best known for its role in neurite outgrowth regulation conducting to neuronal polarity and axonal guidance, including CRMP5 that drives dendrite differentiation. Although CRMP5 is able to control dendritic development, the regulation of its expression remains poorly understood. Here we identify a Sox5 consensus binding sequence in the putative promoter sequence upstream of the CRMP5 gene. By luciferase assays we show that Sox5 increases CRMP5 promoter activity, but not if the putative Sox5 binding site is mutated. We demonstrate that Sox5 can physically bind to the CRMP5 promoter DNA in gel mobility shift and chromatin immunoprecipitation assays. Using a combination of real-time RT-PCR and quantitative immunocytochemistry, we provide further evidence for a Sox5-dependent upregulation of CRMP5 transcription and protein expression in N1E115 cells: a commonly used cell line model for neuronal differentiation. Furthermore, we report that increasing Sox5 levels in this neuronal cell line inhibits neurite outgrowth. This inhibition requires CRMP5 because CRMP5 knockdown prevents the Sox5-dependent effect. We confirm the physiological relevance of the Sox5-CRMP5 pathway in the regulation of neurite outgrowth using mouse primary hippocampal neurons. These findings identify Sox5 as a critical modulator of neurite outgrowth through the selective activation of CRMP5 expression.


Assuntos
Amidoidrolases/genética , Regulação da Expressão Gênica , Crescimento Neuronal/genética , Fatores de Transcrição SOXD/genética , Amidoidrolases/metabolismo , Animais , Sítios de Ligação/genética , Encéfalo/embriologia , Encéfalo/metabolismo , Linhagem Celular Tumoral , Humanos , Camundongos , Mutação , Neuritos/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Regiões Promotoras Genéticas/genética , Ligação Proteica , Fatores de Transcrição SOXD/metabolismo
14.
J Cell Biochem ; 119(3): 2512-2519, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-28941328

RESUMO

ß-thalassemia is a common autosomal recessive disorder characterized by a deficiency in the synthesis of ß-chains. Evidences show that increased HbF levels improve the symptoms in patients with ß-thalassemia or sickle cell anemia. In this study, ZFN technology was applied to induce a mutation in the binding domain region of SOX6 to reactivate γ-globin expression. The sequences coding for ZFP arrays were designed and sub cloned in TDH plus as a transfer vector. The ZFN expression was confirmed using Western blot analysis. In the next step, using the site-directed mutagenesis strategy through the overlap PCR, a missense mutation (D64V) was induced in the catalytic domain of the integrase gene in the packaging plasmid and verified using DNA sequencing. Then, the integrase minus lentivirus containing ZFN cassette was packaged. Transduction of K562 cells with this virus was performed. Mutation detection assay was performed. The indel percentage of the cells transducted with lenti virus containing ZFN was 31%. After 5 days of erythroid differentiation with 15 µg/mL cisplatin, the levels of γ-globin mRNA were sixfold in the cells treated with ZFN compared to untreated cells. In the meantime, the measurement of HbF expression levels was carried out using hemoglobin electrophoresis and showed the same results. Integrase minus lentivirus can provide a useful tool for efficient transient gene expression and helps avoid disadvantages of gene targeting using the native virus. The ZFN strategy applied here to induce indel on SOX6 gene in adult erythroid progenitors may provide a method to activate fetal hemoglobin expression in individuals with ß-thalassemia.


Assuntos
Edição de Genes/métodos , Terapia Genética/métodos , Fatores de Transcrição SOXD/genética , Talassemia beta/genética , gama-Globinas/genética , Humanos , Células K562 , Mutação , Transdução Genética , Nucleases de Dedos de Zinco , gama-Globinas/biossíntese
15.
Gastroenterology ; 154(3): 624-636, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29031500

RESUMO

BACKGROUND & AIMS: The enteric nervous system (ENS) regulates gastrointestinal function via different subtypes of neurons, organized into fine-tuned neural circuits. It is not clear how cell diversity is created within the embryonic ENS; information required for development of cell-based therapies and models of enteric neuropathies. We aimed to identify proteins that regulate ENS differentiation and network formation. METHODS: We generated and compared RNA expression profiles of the entire ENS, ENS progenitor cells, and non-ENS gut cells of mice, collected at embryonic days 11.5 and 15.5, when different subtypes of neurons are formed. Gastrointestinal tissues from R26ReYFP reporter mice crossed to Sox10-CreERT2 or Wnt1-Cre mice were dissected and the 6 populations of cells were isolated by flow cytometry. We used histochemistry to map differentially expressed proteins in mouse and human gut tissues at different stages of development, in different regions. We examined enteric neuronal diversity and gastric function in Wnt1-Cre x Sox6fl/fl mice, which do not express the Sox6 gene in the ENS. RESULTS: We identified 147 transcription and signaling factors that varied in spatial and temporal expression during development of the mouse ENS. Of the factors also analyzed in human ENS, most were conserved. We uncovered 16 signaling pathways (such as fibroblast growth factor and Eph/ephrin pathways). Transcription factors were grouped according to their specific expression in enteric progenitor cells (such as MEF2C), enteric neurons (such as SOX4), or neuron subpopulations (such as SATB1 and SOX6). Lack of SOX6 in the ENS reduced the numbers of gastric dopamine neurons and delayed gastric emptying. CONCLUSIONS: Using transcriptome and histochemical analyses of the developing mouse and human ENS, we mapped expression patterns of transcription and signaling factors. Further studies of these candidate determinants might elucidate the mechanisms by which enteric stem cells differentiate into neuronal subtypes and form distinct connectivity patterns during ENS development. We found expression of SOX6 to be required for development of gastric dopamine neurons.


Assuntos
Neurônios Dopaminérgicos/metabolismo , Sistema Nervoso Entérico/metabolismo , Transdução de Sinais , Estômago/inervação , Fatores de Transcrição/metabolismo , Transcrição Genética , Animais , Comunicação Autócrina , Sistema Nervoso Entérico/embriologia , Esvaziamento Gástrico , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Genótipo , Idade Gestacional , Humanos , Camundongos Knockout , Comunicação Parácrina , Fenótipo , Fatores de Transcrição SOXD/genética , Fatores de Transcrição SOXD/metabolismo , Fatores de Transcrição SOXE/genética , Fatores de Transcrição SOXE/metabolismo , Especificidade da Espécie , Fatores de Transcrição/genética
16.
Development ; 144(15): 2837-2851, 2017 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-28694260

RESUMO

Distinct cortical interneuron (CIN) subtypes have unique circuit functions; dysfunction in specific subtypes is implicated in neuropsychiatric disorders. Somatostatin- and parvalbumin-expressing (SST+ and PV+) interneurons are the two major subtypes generated by medial ganglionic eminence (MGE) progenitors. Spatial and temporal mechanisms governing their cell-fate specification and differential integration into cortical layers are largely unknown. We provide evidence that Coup-TF1 and Coup-TF2 (Nr2f1 and Nr2f2) transcription factor expression in an arc-shaped progenitor domain within the MGE promotes time-dependent survival of this neuroepithelium and the time-dependent specification of layer V SST+ CINs. Coup-TF1 and Coup-TF2 autonomously repress PV+ fate in MGE progenitors, in part through directly driving Sox6 expression. These results have identified, in mouse, a transcriptional pathway that controls SST-PV fate.


Assuntos
Fator II de Transcrição COUP/metabolismo , Fator I de Transcrição COUP/metabolismo , Interneurônios/metabolismo , Neocórtex/citologia , Animais , Fator I de Transcrição COUP/genética , Fator II de Transcrição COUP/genética , Células Cultivadas , Imunoprecipitação da Cromatina , Feminino , Regulação da Expressão Gênica no Desenvolvimento/genética , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Imuno-Histoquímica , Hibridização In Situ , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Parvalbuminas/genética , Parvalbuminas/metabolismo , Fatores de Transcrição SOXD/genética , Fatores de Transcrição SOXD/metabolismo , Somatostatina/genética , Somatostatina/metabolismo
17.
Cell Cycle ; 16(13): 1295-1301, 2017 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-28632999

RESUMO

Long non-coding RNAs (LncRNAs) have been recently regarded as systemic regulators in multiple biologic processes including tumorigenesis. In this study, we observed the expression of lncRNA lnc-sox5 was significantly increased in colorectal cancer (CRC). Despite the CRC cell growth, cell cycle and cell apoptosis was not affected by lnc-sox5 knock-down, lnc-sox5 knock-down suppressed CRC cell migration and invasion. In addition, xenograft animal model suggested that lnc-sox5 knock-down significantly suppressed the CRC tumorigenesis. Our results also showed that the expression of indoleamine 2,3-dioxygenase 1 (IDO1) was significantly reduced by lnc-sox5 knock-down and therefore modulated the infiltration and cytotoxicity of CD3+CD8+T cells. Taken together, these results suggested that lnc-sox5 unbalances tumor microenvironment to regulate colorectal cancer progression.


Assuntos
Neoplasias Colorretais/patologia , RNA Longo não Codificante/metabolismo , Fatores de Transcrição SOXD/genética , Animais , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Nus , Interferência de RNA , RNA Longo não Codificante/antagonistas & inibidores , RNA Longo não Codificante/genética , RNA Interferente Pequeno/metabolismo , Transplante Heterólogo , Microambiente Tumoral , Vimentina/metabolismo
18.
Animal ; 11(12): 2268-2274, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28490391

RESUMO

MicroRNAs (miRNAs) are a class of small non-coding RNAs that negatively regulate gene expression of target messenger RNAs (mRNAs) and miRNAs have been proven to play vital roles in skeletal muscle development. The miRNA-499-5p has been reported to be negatively related with the expression of Sox6, a critical transcription factor for the maintenance of fast-twitch skeletal muscle. In this study, we amplified a length of 2012-bp mRNA that contains a 1512-bp porcine Sox6 (pSox6) 3'UTR from skeletal muscle of a Duroc×Landrace×Yorkshire pig. By luciferase reporter assay we verified that pSox6 is a target of miR-499-5p. In extensor digitorum longus and Soleus muscles of pigs, the expression levels of miR-499-5p and pSox6 mRNA were also inversely correlated. Besides, overexpression of miR-499-5p in porcine satellite cells promoted the expression of MyHC I and MyHC IIa mRNA, along with a reduction of pSox6 mRNA. Taken together, these results indicate that miR-499-5p may facilitate the oxidative myofibers formation by downregulating pSox6 expression.


Assuntos
Regulação da Expressão Gênica , MicroRNAs/genética , Fatores de Transcrição SOXD/genética , Suínos/genética , Animais , Células Cultivadas , Regulação para Baixo , Feminino , Genes Reporter , Células HEK293 , Humanos , Masculino , Desenvolvimento Muscular/genética , Músculo Esquelético/crescimento & desenvolvimento , RNA Mensageiro/genética , Células Satélites de Músculo Esquelético , Suínos/crescimento & desenvolvimento
19.
Tumour Biol ; 39(5): 1010428317705508, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28475012

RESUMO

SOX6 plays important roles in cell proliferation, differentiation, and cell fate determination. It has been confirmed that SOX6 is a tumor suppressor and downregulated in various cancers, including esophageal squamous cell carcinoma, hepatocellular carcinoma, and chronic myeloid leukemia. Netrin-1 is highly expressed in various human cancers and acts as an anti-apoptotic and proangiogenic factor to drive tumorigenesis. The role of SOX6 and netrin-1 in regulating the growth of ovarian tumor cells still remains unclear. Real-time polymerase chain reaction and western blot were used to determine the SOX6 messenger RNA and protein levels, respectively, in ovarian cancer cells and tumor tissues. Stable transfection of SOX6 was conducted to overexpress SOX6 in PA-1 and SW626 cells. Cell viability was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Invasion of ovarian cancer cells and migration of human umbilical vein endothelial cells were confirmed by Transwell assays. To overexpress netrin-1, ovarian cancer cells with SOX6 restoration was transduced with netrin-1 lentiviral particles. PA-1 xenografts in a nude mice model were used to conduct in vivo evaluation of the role of SOX6 and its relationship with netrin-1 in tumor growth and angiogenesis. In this study, we found significantly reduced SOX6 levels in PA-1, SW626, SK-OV-3, and CaoV-3 ovarian cancer cell lines and human tumor tissues in comparison with normal human ovarian epithelial cells or matched non-tumor tissues. SOX6 overexpression by stable transfection dramatically inhibited proliferation and invasion of PA-1 and SW626 cells. Also, conditioned medium from PA-1 and SW626 cells with SOX6 restoration exhibited reduced ability to induce human umbilical vein endothelial cells migration and tube formation compared with conditioned medium from the cells with transfection control. Furthermore, an inverse relationship between SOX6 and netrin-1 expression was observed in PA-1 and SW626 cells. Overexpression of netrin-1 in ovarian cancer cells with forced SOX6 expression remarkably abrogated the inhibitory effect of SOX6 on proliferation, invasion of the cells, and tumor xenograft growth and vascularity in vivo. Human umbilical vein endothelial cell migration and tube formation were enhanced in the conditioned medium from the ovarian cancer cells transduced with netrin-1 lentivirus particles. Our observations revealed that SOX6 is a tumor suppressor in ovarian cancer cells, and SOX6 exerts an inhibitory effect on the proliferation, invasion, and tumor cell-induced angiogenesis of ovarian cancer cells, whereas nerin-1 plays an opposite role and its expression is inversely correlated with SOX6. Moreover, our findings suggest a new role of SOX6 and netrin-1 for understanding the progression of ovarian cancer and have the potential for the development of new diagnosis and treatment strategies for ovarian cancer.


Assuntos
Neovascularização Patológica/genética , Fatores de Crescimento Neural/genética , Neoplasias Ovarianas/genética , Fatores de Transcrição SOXD/genética , Proteínas Supressoras de Tumor/genética , Animais , Linhagem Celular Tumoral , Proliferação de Células/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana , Humanos , Camundongos , Invasividade Neoplásica/genética , Neovascularização Patológica/patologia , Fatores de Crescimento Neural/biossíntese , Netrina-1 , Neoplasias Ovarianas/patologia , Fatores de Transcrição SOXD/biossíntese , Proteínas Supressoras de Tumor/biossíntese , Ensaios Antitumorais Modelo de Xenoenxerto
20.
Biochemistry (Mosc) ; 82(4): 438-445, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28371600

RESUMO

Long noncoding RNAs (lncRNAs) have been recently regarded as systemic regulators in multiple biological processes including tumorigenesis. In this study, we report an ultra-highly expressed lncRNA, lnc-Sox5, in tongue tumor tissues. The results imply that lnc-Sox5 may play vital role in tongue carcinoma progression. We observed that the growth of Tca8113 cells was suppressed by lnc-Sox5 downregulation. Additionally, lnc-Sox5 knockdown simultaneously increased Tca8113 cell apoptosis, but the cell cycle was arrested. RNA immunoprecipitation suggested that HuR directly bound to and stabilized lnc-Sox5 RNA. Consistently, HuR knockdown reduced the level of lnc-Sox5 in Tca8113 cells. However, overexpression of HuR induced more lnc-Sox5 in Tca8113 cells. Both lnc-Sox5 knockdown and HuR knockdown suppressed Tca8113 cell tumorigenesis in xenograft models. These results suggest that lnc-Sox5, which was stabilized by HuR, could regulate carcinogenesis of tongue cancer and may serve as a predicted target for tongue carcinoma therapies.


Assuntos
Carcinoma de Células Escamosas/patologia , Proteína Semelhante a ELAV 1 , RNA Longo não Codificante/genética , RNA Mensageiro/genética , Fatores de Transcrição SOXD/genética , Neoplasias da Língua/patologia , Animais , Carcinoma de Células Escamosas/genética , Linhagem Celular Tumoral , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias da Língua/genética
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