• Molecules and Cells
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• 제40권2호
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• pp.151-161
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• 2017

Proper synaptic function in neural circuits requires precise pairings between correct pre- and post-synaptic partners. Errors in this process may underlie development of neuropsychiatric disorders, such as autism spectrum disorder (ASD). Development of ASD can be influenced by genetic factors, including copy number variations (CNVs). In this study, we focused on a CNV occurring at the 16p11.2 locus in the human genome and investigated potential defects in synaptic connectivity caused by reduced activities of genes located in this region at Drosophila larval neuromuscular junctions, a well-established model synapse with stereotypic synaptic structures. A mutation of rolled, a Drosophila homolog of human mitogen-activated protein kinase 3 (MAPK3) at the 16p11.2 locus, caused ectopic innervation of axonal branches and their abnormal defasciculation. The specificity of these phenotypes was confirmed by expression of wild-type rolled in the mutant background. Albeit to a lesser extent, we also observed ectopic innervation patterns in mutants defective in Cdk2, Gq, and Gp93, all of which were expected to interact with Rolled MAPK3. A further genetic analysis in double heterozygous combinations revealed a synergistic interaction between rolled and Gp93. In addition, results from RT-qPCR analyses indicated consistently reduced rolled mRNA levels in Cdk2, Gq, and Gp93 mutants. Taken together, these data suggest a central role of MAPK3 in regulating the precise targeting of presynaptic axons to proper postsynaptic targets, a critical step that may be altered significantly in ASD.

• Asian Pacific Journal of Cancer Prevention
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• 제14권4호
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• pp.2361-2366
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• 2013

Diverse studies have shown that miR-155 is overexpressed in different tumor types. However, the precise molecular mechanism of the ectopic expression of miR-155 in breast cancer is still poorly understood. To further explore the role of miR-155 in breast tumorigenesis, we here assessed the influence of miR-155 antisense oligonucleotide (miR-155 ASO) on MDA-MB-157 cell viability and apoptosis in vitro. Furthermore, the effects of inhibitory effects of miR-155 on the growth of xenograft tumors in vivo were determined with performance of immunohistochemistry to detect expression of caspase-3, a pivotal apoptosis regulatory factor, in xenografts. Transfection efficiency detected by laser confocal microscope was higher than 80%. The level of miR-155 expression was significantly decreased (P<0.05) in the cells transfected with miR-155 ASO, compared with that in cells transfected with a negative control. After being transfected with miR-155 ASO, the viability of MDA-MB-157 cells was reduced greatly (P<0.05) and the number of apoptotic cells was increased significantly. Additionally, miR-155 ASO inhibited the growth of transplanted tumor in vivo and significantly increased the expression of caspase-3. Taken together, our study revealed that miR-155 ASO can induce cell apoptosis and inhibit cell proliferation in vitro. Moreover, miR-155 ASO could significantly repress tumor growth in vivo, presumably by inducing apoptosis via caspase-3 up-regulation. These findings provide experimental evidence for using miR-155 as a therapeutic target of breast carcinoma.

• Asian Pacific Journal of Cancer Prevention
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• 제13권11호
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• pp.5799-5804
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• 2012

Family with sequence similarity 189, member B (FAM189B), alias COTE1, a putative oncogene selected by microarray, for the first time was here found to be significantly up-regulated in hepatocellular carcinoma (HCC) specimens and HCC cell lines. mRNA expression of COTE1 in HCC samples and cell lines was detected by reverse transcription-polymerase chain reaction (RT-PCR) and real-time PCR, while protein expression of COTE1 in HCC tissues was assessed by immunohistochemistry. In addition, invasion of HCC cells was observed after overexpressing or silencing COTE1. In the total of 48 paired HCC specimens, compared with the adjacent non-cancer tissues, the expression of COTE1 was up-regulated in 31 (p<0.01). In HCC cell lines, COTE1 expression was significantly higher than in normal human adult liver (p<0.01). Overexpression of COTE1 enhanced HCC-derived LM6 and MHCC-L cellular invasion in vitro. In contrast, COTE1 knockdown via RNAi markedly suppressed these phenotypes, as documented in LM3 and MHCC-H HCC cells. Mechanistic analyses indicated that COTE1 could physically associate with WW domain oxidoreductase (WWOX), a tumor suppressor. COTE1 may be closely correlated with invasion of hepatocellular carcinoma (HCC) cells and thus may serve as an effective target for gene therapy.

• 생명과학회지
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• 제26권5호
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• pp.614-619
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• 2016

Par-4는 다양한 세포사멸 자극에 세포 사멸을 조절하고, 종양 억제기능을 가지고 있다. 그러나, Par-4에 의한 암세포의 이동 및 침윤에 대한 연구는 수행되지 않았다. 본 연구에서 Par-4단백질의 과발현이 인간 신장암 Caki세포에서 MMP-2의 활성화를 억제하지만 MMP-9 활성에는 영향을 주지 않았다. Par-4에 의한 MMP-2의 활성 억제는 leucine zipper domain이 결실된 Par-4 에서는 확인되지 않았다. Par-4 siRNA를 이용한 knock-down 실험에서 PMA 처리 시 세포이동 및 침윤 증가함을 확인하였다. Par-4의 과발현과 knock-dwon에서 MMP-2 mRNA 발현의 변화를 확인 할 수 없었다. 이 점은 Par-4 매개의 MMP-2 활성 억제는 전사 후 조절을 통하여 야기됨을 추측 할 수 있다.

• BMB Reports
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• 제45권11호
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• pp.629-634
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• 2012

The human glycoprotein, stanniocalcin 2 (STC2) plays multiple roles in several tumor types, however, its function and clinical significance in hepatocellular carcinoma (HCC) remain unclear. In this study, we detected STC2 expression by quantitative real-time PCR and found STC2 was upregulated in HCC tissues, correlated with tumor size and multiplicity of HCC. Ectopic expression of STC2 markedly promoted HCC cell proliferation and colony formation, while silencing of endogenous STC2 resulted in a reduced cell growth by cell cycle delay in G0/G1 phase. Western blot analysis demonstrated that STC2 could regulate the expression of cyclin D1 and activate extracellular signal-regulated kinase 1/2 (ERK1/2) in a dominant-positive manner. Transwell chamber assay also indicated altered patterns of STC2 expression had an important effect on cell migration. Our findings suggest that STC2 functions as a potential oncoprotein in the development and progression of HCC as well as a promising molecular target for HCC therapy.

• Asian Pacific Journal of Cancer Prevention
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• 제14권2호
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• pp.835-840
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• 2013

Accumulating evidence has shown that microRNAs are involved in cancer development and progression. However, it remains unknown about the potential role of miR-19a in the pathogenesis of gastric cancer. Here, we report that suppressor of cytokine signaling 1 (SOCS1) is a novel target of miR-19a in gastric cancer cells and that miR-19a expression is inversely correlated with SOCS1 expression in gastric cancer cells and a subset of gastric cancer tissues. Ectopic expression of miR-19a dramatically promoted proliferation and tumorigenicity of gastric cancer cells both in vitro and in vivo. Moreover, we showed that silencing of SOCS1 promoted cell growth and colony formation resembling that of miR-19a overexpression, whereas re-introduction of SOCS1 (without the 3'-UTR) attenuated the pro-tumorigenic functions. Taken together, our findings suggest that the SOCS1 gene is a direct target of miR-19a, which functions as an oncogenic miRNA in gastric cancer by repressing the expression of tumor suppressor SOCS1.

• Molecules and Cells
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• 제38권1호
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• pp.75-80
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• 2015

Osteoclasts are unique cells responsible for the resorption of bone matrix. MicroRNAs (miRNAs) are involved in the regulation of a wide range of physiological processes. Here, we examined the role of miR-26a in RANKL-induced osteoclastogenesis. The expression of miR-26a was upregulated by RANKL at the late stage of osteoclastogenesis. Ectopic expression of an miR-26a mimic in osteoclast precursor cells attenuated osteoclast formation, actin-ring formation, and bone resorption by suppressing the expression of connective tissue growth factor/CCN family 2 (CTGF/CCN2), which can promote osteoclast formation via upregulation of dendritic cell-specific transmembrane protein (DC-STAMP). On the other hand, overexpression of miR-26a inhibitor enhanced RANKL-induced osteoclast formation and function as well as CTGF expression. In addition, the inhibitory effect of miR-26a on osteoclast formation and function was prevented by treatment with recombinant CTGF. Collectively, our results suggest that miR-26a modulates osteoclast formation and function through the regulation of CTGF.

• Molecules and Cells
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• 제28권5호
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• pp.479-487
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• 2009

Glutathione reductase (GR) is an enzyme that recycles a key cellular antioxidant molecule glutathione (GSH) from its oxidized form (GSSG) thus maintaining cellular redox homeostasis. A recombinant plasmid to overexpress a GR of Brassica rapa subsp. pekinensis (BrGR) in E. coli BL21 (DE3) was constructed using an expression vector pKM260. Expression of the introduced gene was confirmed by semi-quantitative RT-PCR, immunoblotting and enzyme assays. Purification of the BrGR protein was performed by IMAC method and indicated that the BrGR was a dimmer. The BrGR required NADPH as a cofactor and specific activity was approximately 458 U. The BrGR-expressing E. coli cells showed increased GR activity and tolerance to $H_2O_2$, menadione, and heavy metal ($CdCl_2$, $ZnCl_2$ and $AlCl_2$)-mediated growth inhibition. The ectopic expression of BrGR provoked the co-regulation of a variety of antioxidant enzymes including catalase, superoxide dismutase, glutathione peroxidase, and glucose-6-phosphate dehydrogenase. Consequently, the transformed cells showed decreased hydroperoxide levels when exposed to stressful conditions. A proteomic analysis demonstrated the higher level of induction of proteins involved in glycolysis, detoxification/oxidative stress response, protein folding, transport/binding proteins, cell envelope/porins, and protein translation and modification when exposed to $H_2O_2$ stress. Taken together, these results indicate that the plant GR protein is functional in a cooperative way in the E. coli system to protect cells against oxidative stress.

• BMB Reports
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• 제53권12호
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• pp.628-633
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• 2020

WNT11 is a member of the non-canonical Wnt family and plays a crucial role in tumor progression. However, the regulatory mechanisms underlying WNT11 expression are unclear. Tumor necrosis factor-alpha (TNFα) is a major inflammatory cytokine produced in the tumor microenvironment and contributes to processes associated with tumor progression, such as tumor invasion and metastasis. By using site-directed mutagenesis and introducing a serial deletion in the 5'-regulatory region of WNT11, we observed that TNFα activates the early growth response 1 (EGR1)-binding sequence (EBS) in the proximal region of WNT11 and that the transcription factor EGR1 is necessary for the TNFα-induced transcription of WNT11. EGR1 bound directly to the EBSs within the proximal 5'-regulatory region of WNT11 and ectopic expression of EGR1 stimulated WNT11 promoter activity, whereas the knockdown of EGR1 expression by RNA interference reduced TNFα-induced WNT11 expression in T47D breast cancer cells. We also observed that mitogen-activated protein kinases (MAPK), extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 kinase mediated TNFα-induced transcription of WNT11 via EGR1. Our results suggest that EGR1 directly targets WNT11 in response to TNFα stimulation in breast cancer cells.

• Molecules and Cells
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• 제46권12호
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• pp.746-756
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• 2023

A recent study revealed that the loss of Deup1 expression does not affect either centriole amplification or multicilia formation. Therefore, the deuterosome per se is not a platform for amplification of centrioles. In this study, we examine whether gain-of-function of Deup1 affects the development of multiciliated ependymal cells. Our time-lapse study reveals that deuterosomes with an average diameter of 300 nm have two different fates during ependymal differentiation. In the first instance, deuterosomes are scattered and gradually disappear as cells become multiciliated. In the second instance, deuterosomes self-organize into a larger aggregate, called a deuterosome cluster (DC). Unlike scattered deuterosomes, DCs possess centriole components primarily within their large structure. A characteristic of DC-containing cells is that they tend to become primary ciliated rather than multiciliated. Our in utero electroporation study shows that DCs in ependymal tissue are mostly observed at early postnatal stages, but are scarce at late postnatal stages, suggesting the presence of DC antagonists within the differentiating cells. Importantly, from our bead flow assay, ectopic expression of Deup1 significantly impairs cerebrospinal fluid flow. Furthermore, we show that expression of mouse Deup1 in Xenopus embryos has an inhibitory effect on differentiation of multiciliated cells in the epidermis. Taken together, we conclude that the DC formation of Deup1 in multiciliated cells inhibits production of multiple centrioles.