• BMB Reports
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• 제53권9호
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• pp.484-489
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• 2020

Epilepsy is a neurological disorder characterized by unpredictable seizures, which are bursts of electrical activity that temporarily affect the brain. Cereblon (CRBN), a DCAFs (DDB1 and CUL4-associated factors), is a well-established protein associated with human mental retardation. Being a substrate receptor of the cullin-RING E3 ubiquitin ligase (CRL) 4 complex, CRBN mediates ubiquitination of several substrates and conducts multiple biological processes. In the central nervous system, the large-conductance Ca²⁺-activated K⁺ (BK_Ca) channel, which is the substrate of CRBN, is an important regulator of epilepsy. Despite the functional role and importance of CRBN in the brain, direct injection of pentylenetetrazole (PTZ) to induce seizures in CRBN knock-out mice has not been challenged. In this study, we investigated the effect of PTZ in CRBN knock-out mice. Here, we demonstrate that, compared with WT mice, CRBN knock-out mice do not show the intensification of seizures by PTZ induction. Moreover, electroencephalography recordings were also performed in the brains of both WT and CRBN knockout mice to identify the absence of significant differences in the pattern of seizure activities. Consistently, immunoblot analysis for validating the protein level of the CRL4 complex containing CRBN (CRL4^Crbn) in the mouse brain was carried out. Taken together, we found that the deficiency of CRBN does not affect PTZ-induced seizure.

• Asian Pacific Journal of Cancer Prevention
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• 제13권11호
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• pp.5865-5869
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• 2012

RHOXF1 has been shown to be expressed in embryonic stem cells, adult germline stem cells and some cancer lines. It has been proposed as a candidate gene to encode transcription factors regulating downstream genes in the human testis with antiapoptotic effects. Its expression in cancer cell lines has implied a similar role in the process of tumorigenesis. The human breast cancer cell lines MDA-MB-231 and MCF-7 were cultured in DMEM medium and transfected with a pGFP-V-RS plasmid bearing an RHOXF1 specific shRNA. Quantitative real-time RT-PCR was performed for RHOXF1, CASP8, BCL2 and HPRT genes. Decreased RHOXF1 expression was confirmed in cells after transfection. shRNA knock down of RHOXF1 resulted in significantly decreased BCL2 expression in both cell lines but no change in CASP8 expression. shRNA targeting RHOXF1 was shown to specifically mediate RHOXF1 gene silencing, so RHOXF1 can mediate transcriptional activation of the BCL2 in cancers and may render tumor cells resistant to apoptotic cell death induced by anticancer therapy. shRNA mediated knock down of RHOXF1 can be effective in induction of apoptotic pathway in cancer cells via BCL2 downregulation, so it can have potential therapeutic utility for human breast cancer.

• Asian Pacific Journal of Cancer Prevention
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• 제14권11호
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• pp.6625-6629
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• 2013

Breast cancer accounts for one third of new cancer cases among women. The need for biomarkers for early detection is the stimulus to researchers to evaluate altered expression of genes in tumours. Cancer-testis (CT) genes are a group with limited expression in normal tissues except testis but up-regulation in a wide variety of cancers. We here evaluated expression of two CT genes named FBXO39 and TDRD4 in 32 invasive ductal carcinoma samples, 10 fibroadenomas and 6 normal breast tissue samples, in addition to two breast cancer cell lines, MCF-7 and MDA-MB-231, by the means of quantitative real time RT-PCR. FBXO39 showed significant up-regulation in invasive ductal carcinoma samples in comparison with normal samples. It also was expressed in both cell lines and after RHOXF1 gene knock down it was down-regulated in MCF-7 but up-regulated in the MDA-MB-231 cell line. TDRD4 was not expressed in the MCF-7 cell line and any of the tissue samples except testis. However, it was expressed in MDA-MB-231 and was up-regulated after RHOXF1 gene knock down. Our results show that FBXO39 but not TDRD4 can be used for cancer detection and if proved to be immunogenic, might be a putative candidate for breast cancer immunotherapy.

• Molecules and Cells
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• 제38권1호
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• pp.65-74
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• 2015

Carbohydrate antigens expressed on pig cells are considered to be major barriers in pig-to-human xenotransplantation. Even after ${\alpha}1,3$-galactosyltransferase gene knock-out (GalT-KO) pigs are generated, potential non-Gal antigens are still existed. However, to the best of our knowledge there is no extensive study analyzing N-glycans expressed on the GalT-KO pig tissues or cells. Here, we identified and quantified totally 47 N-glycans from wild-type (WT) and GalT-KO pig fibroblasts using mass spectrometry. First, our results confirmed the absence of galactose-alpha-1,3-galactose (${\alpha}$-Gal) residue in the GalT-KO pig cells. Interestingly, we showed that the level of overall fucosylated N-glycans from GalT-KO pig fibroblasts is much higher than from WT pig fibroblasts. Moreover, the relative quantity of the N-glycolylneuraminic acid (NeuGc) antigen is slightly higher in the GalT-KO pigs. Thus, this study will contribute to a better understanding of cellular glycan alterations on GalT-KO pigs for successful xenotransplantation.

• Reproductive and Developmental Biology
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• 제31권1호
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• pp.21-28
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• 2007

During early embryo development, Oct-4 is an important transcription factor for the early differentiation the present study was first examined methylation status in distal enhancer and promoter region of Oct-4 during mouse pre-implantation embryo development. In oocyte and sperm, high methylation was observed in both distal and proximal of promoter in Oct-4. Following fertilization relatively high methylation level remained until 8-cell stage embryos, but decreased at the morula and blastocyst stage. Specific gene knock down of Oct-4 by siRNA injection into zygote induced higher methylation rates of both distal and proximal region of promoter of Oct-4. These results suggest a functional link between the DNA methylation status of distal and promoter resign in the Oct-4 gene and the gene sequence-specific transcriptional silencing by exogenous siRNA injection during mouse preimplantation embryos.

• 생물정신의학
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• 제9권1호
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• pp.8-14
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• 2002

Patients with attention deficit hyperactivity disorder(ADHD) have symptoms of inattention, hyperactivity, impulsivity. Symptoms of ADHD are responsive to medications such as methylphenidate, dextroamphetamine, pemoline, and bupropion. The functional change of the dopamine transporter is related to the therapeutic effect of these drugs. This can be one reason for the dopamine transporter to be emphasized in the research field of ADHD. ADHD has a genetic tendency. Since dopamine transporter gene(DAT1) knock out mice were reported to be several times more active than normal mice in a novel situation, lights has been shed on DAT1 as a candidate gene for ADHD. Though there have been several studies which reported an association between DAT1 and ADHD, the association between DAT1 and ADHD is not conclusive. Since Vandenbergh reported the DAT1 polymorphism with variable number of tandem repeats(VNTR), and the racial differences in allelic frequencies of the DAT1, wide ethnic variation in the distribution of the DAT1 polymorphism had been confirmed. Wide ethnic variation in the distribution of the DAT1 suggested that there might be ethnic difference in the association between DAT1 and ADHD. Before applying previous findings to Koreans, verification might be needed for Korean patients with ADHD.

• Bulletin of the Korean Chemical Society
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• 제30권2호
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• pp.323-326
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• 2009

SR proteins are essential splicing regulators and also modulate alternative splicing events, which function both as redundant and substrate-specific manner. The Drosophila B52/SRp55, a member of the SR protein family, is essential for the fly development in vivo, as deletion of B52 gene results in lethality of animals at the second instar larval stage. Identification of the splicing target genes of B52 thus should be crucial for the understanding of the specific developmental role of B52 in vivo. In this study, we performed whole-genome DNA microarray experiments with a B52- knock-out animal. Analysis of the microarray data not only provided the B52-dependent gene expression signature, but also revealed a larval-stage specific, alternative splicing target gene of B52. Our result thus provides a starting point to understand the essential function of B52 at the organismal level.

• Reproductive and Developmental Biology
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• 제29권1호
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• pp.19-24
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• 2005

저밀도 리포단백질 수용체 관련 단백질 5(LRP5)는 간과 췌장을 포함하여 많은 조직에서 발현하며 아포리포단백질 E와 결합한다. 이와 같은 LRP5 유전자의 체내 기능을 규명하기 위하여 LRP5 유전자가 결손된 생쥐를 개발하였다. 먼지 LRP5 genomic DNA는 TT2 ES 세포로부터 분리하였으며 LRP5 유전자의 엑손 18에 neo 유전자를 삽입한 vector를 구축하고 TT2 ES 세포에 도입하였다. 178개의 G418 내성을 보인 세포 중 상동유전자 재조합에 의하여 targeting vector가 LRP5 유전자 위치에 삽입된 clone은 3개였다. 키메라 생쥐는 상실배기 수정을 ES 세포와 응집시켜 생산하였으며 생산된 키메라 생쥐는 C57BL/6 생쥐와 교미를 유도하여 heterozygous를 얻었다. 또한 이들 heterozygous간의 교배에 의하여 LRP5 유전자 결손 생쥐를 생산하였다. 이러한 생쥐는 LRP5 유전자의 체내 기능연구에 있어서 모델로 이용될 것으로 생각된다.

• Journal of Plant Biotechnology
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• 제37권4호
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• pp.483-493
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• 2010

Knock-out of a gene by insertional mutagenesis is a direct way to address its function through the mutant phenotype. Among ca. 15,000 gene-trapped Ds insertion lines of rice, we identified one line from selected sensitive lines in highly salt stress. We conducted gene tagging by TAIL-PCR, and DNA gel blot analysis from salt sensitive mutant. A gene encoding an oligopeptide transporter (OPT family) homologue was disrupted by the insertion of a Ds transposon into the OsOPT10 gene that was located shot arm of chromosome 8. The OsOPT10 gene (NP_001062118.) has 6 exons and encodes a protein (752 aa) containing the OPT family domain. RT-PCR analysis showed that the expression of OsOPT10 gene was rapidly and strongly induced by stresses such as high-salinity (250 mM), osmotic, drought, $100\;{\mu}M$ ABA. The subcellular localization assay indicated that OsOPT10 was localized specifically in the plasma membrane. Overexpression of OsOPT10 in Arabidopsis thaliana and rice conferred tolerance of transgenic plants to salt stress. Further we found expression levels of some stress related genes were inhibited in OsOPT10 transgenic plants. These results suggested that OsOPT10 might play crucial but differential roles in plant responses to various abiotic stresses.

• Animal Bioscience
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• 제35권6호
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• pp.791-803
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• 2022

Genome/gene-editing (GE) techniques, characterized by a low technological barrier, high efficiency, and broad application among organisms, are now being employed not only in medical science but also in agriculture/veterinary science. Different engineered CRISPR/Cas9s have been identified to expand the application of this technology. In pig production, GE is a precise new breeding technology (NBT), and promising outcomes in improving economic traits, such as growth, lean or healthy meat production, animal welfare, and disease resistance, have already been documented and reviewed. These promising achievements in porcine gene editing, including the Myostatin gene knockout (KO) in indigenous breeds to improve lean meat production, the uncoupling protein 1 (UCP1) gene knock-in to enhance piglet thermogenesis and survival under cold stress, the generation of GGTA1 and CMP-N-glycolylneuraminic acid hydroxylase (CMAH) gene double KO (dKO) pigs to produce healthy red meat, and the KO or deletion of exon 7 of the CD163 gene to confer resistance to porcine reproductive and respiratory syndrome virus infection, are described in the present article. Other related approaches for such purposes are also discussed. The current trend of global regulations or legislation for GE organisms is that they are exempted from classification as genetically modified organisms (GMOs) if no exogenes are integrated into the genome, according to product-based and not process-based methods. Moreover, an updated case study in the EU showed that current GMO legislation is not fit for purpose in term of NBTs, which contribute to the objectives of the EU's Green Deal and biodiversity strategies and even meet the United Nations' sustainable development goals for a more resilient and sustainable agri-food system. The GE pigs generated via NBT will be exempted from classification as GMOs, and their global valorization and commercialization can be foreseen.