• Title/Summary/Keyword: SiRNA

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Identification and Characterization of Protein Arginine Methyltransferase 1 in Acanthamoeba castellanii

  • Moon, Eun-Kyung;Kong, Hyun-Hee;Hong, Yeonchul;Lee, Hae-Ahm;Quan, Fu-Shi
    • Parasites, Hosts and Diseases
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    • v.55 no.2
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    • pp.109-114
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    • 2017
  • Protein arginine methyltransferase (PRMT) is an important epigenetic regulator in eukaryotic cells. During encystation, an essential process for Acanthamoeba survival, the expression of a lot of genes involved in the encystation process has to be regulated in order to be induced or inhibited. However, the regulation mechanism of these genes is yet unknown. In this study, the full-length 1,059 bp cDNA sequence of Acanthamoeba castellanii PRMT1 (AcPRMT1) was cloned for the first time. The AcPRMT1 protein comprised of 352 amino acids with a SAM-dependent methyltransferase PRMT-type domain. The expression level of AcPRMT1 was highly increased during encystation of A. castellanii. The EGFP-AcPRMT1 fusion protein was distributed over the cytoplasm, but it was mainly localized in the nucleus of Acanthamoeba. Knock down of AcPRMT1 by synthetic siRNA with a complementary sequence failed to form mature cysts. These findings suggested that AcPRMT1 plays a critical role in the regulation of encystation of A. castellanii. The target gene of AcPRMT1 regulation and the detailed mechanisms need to be investigated by further studies.

Production of 3-Hydroxypropionic Acid from Acrylic Acid by Newly Isolated Rhodococcus erythropolis LG12

  • Lee, Sang-Hyun;Park, Si-Jae;Park, Oh-Jin;Cho, Jun-Hyeong;Rhee, Joo-Won
    • Journal of Microbiology and Biotechnology
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    • v.19 no.5
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    • pp.474-481
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    • 2009
  • A novel microorganism, designated as LG12, was isolated from soil based on its ability to use acrylic acid as the sole carbon source. An electron microscopic analysis of its morphological characteristics and phylogenetic classification by 16S rRNA homology showed that the LG12 strain belongs to Rhodococcus erythropolis. R. erythropolis LG12 was able to metabolize a high concentration of acrylic acid (up to 40 g/l). In addition, R. erythropolis LG12 exhibited the highest acrylic acid-degrading activity among the tested microorganisms, including R. rhodochrous, R. equi, R. rubber, Candida rugosa, and Bacillus cereus. The effect of the culture conditions of R. erythropo/is LG12 on the production of 3-hydroxypropionic acid (3HP) from acrylic acid was also examined. To enhance the production of 3HP, acrylic acid-assimilating activity was induced by adding 1 mM acrylic acid to the culture medium when the cell density reached an $OD_{600}$ of 5. Further cultivation of R. erythropo/is LG 12 with 40 g/l of acrylic acid resulted in the production of 17.5 g/l of 3HP with a molar conversion yield of 44% and productivity of 0.22 g/l/h at $30^{\circ}C$ after 72 h.

Histone H3 is Digested by Granzyme A During Compromised Cell Death in the Raji Cells

  • Lee, Phil Young;Park, Byoung Chul;Chi, Seung Wook;Bae, Kwang-Hee;Kim, Sunhong;Cho, Sayeon;Kim, Jeong-Hoon;Park, Sung Goo
    • Journal of Microbiology and Biotechnology
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    • v.25 no.9
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    • pp.1578-1582
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    • 2015
  • Granzyme A (GzmA) was identified as a cytotoxic T lymphocyte protease protein expressed in the nucleus. A number of nuclear proteins are well known as GzmA substrates, and GzmA is related with caspase-independent apoptosis. Histones H1, H2B, and H3 were identified as GzmA substrates through in vitro experiment with purified nucleosome. Here, we demonstrated that histone H3 was cleaved by GzmA in vivo during staurosporine-induced cell death. Moreover, histone H3 cleavage was blocked by the GzmA inhibitor nafamostat mesylate and by GzmA knockdown using siRNA. Taken together, we verified that histone H3 is a real substrate for GzmA in vivo in the Raji cells treated by staurosporin.

Silencing of COX-2 by RNAi Modulates Epithelial-Mesenchymal Transition in Breast Cancer Cells Partially Dependent on the PGE2 Cascade

  • Cao, Juan;Yang, Xiao;Li, Wen-Tong;Zhao, Chun-Ling;Lv, Shi-Jun
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.22
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    • pp.9967-9972
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    • 2014
  • In order to prove whether downregulation of COX-2 (Cyclooxygenase-2) could modulate the epithelial-mesenchymal transition (EMT) of breast cancer, celecoxib and siRNA were respectively used to inhibit COX-2 function and expression in MDA-MB-231 cells. The EMT reversal effect in the RNAi treated group was better than that of the celecoxib group while there were no obvious differences in the medium $PGE_2$ levels between the two groups. The results show that COX-2 pathways may contribute considerably to EMT of breast cancer cells, partially dependent on the PGE2 cascade. Akt2, ZEB2 and Snail were measured to clarify the underlying mechanisms of COX-2 on EMT; COX-2 may modulate EMT of breast cancer by regulating these factors. This finding may be helpful to elucidate the mechanisms of selective COX-2 inhibitor action in EMT modulation in breast cancer.

ZNF217 is Overexpressed and Enhances Cell Migration and Invasion in Colorectal Carcinoma

  • Zhang, Zi-Chao;Zheng, Li-Qiang;Pan, Li-Jie;Guo, Jin-Xing;Yang, Guo-Shan
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.6
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    • pp.2459-2463
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    • 2015
  • Background: To investigate the expression and clinical significance of zinc finger protein 217 (ZNF217) in human colorectal carcinoma (CRC). Materials and Methods: The expression of ZNF217 in 60 CRC tissues and matched tumor adjacent tissues, collected between January 2013 and June 2014, was assessed immunohistochemically. The relationship between the expression of ZNF217 and clinicopathlogical features was analyzed by Pearson chi-square test. In addition, siRNA was used to down-regulate the expression of ZNF217 in CRC cells. The effects of ZNF217 for cell migration and invasion were measured by wound healing assay and transwell assay, respectively. Results: The expression level of ZNF217 was significantly higher in CRC tissues than in tumor adjacent tissues (p<0.05), positively correlating with tumor size, lymphatic metastasis and advanced TNM stage (p<0.05). Down-regulation of ZNF217 in CRC cells could significantly suppress cell migration and invasion. Conclusions: ZNF217 is overexpressed in colorectal carcinoma tissues and is associated with tumor malignant clinicopathological features. ZNF217 may promote CRC progression by inducing cell migration and invasion.

Inhibition of SMP30 Gene Expression Influences the Biological Characteristics of Human Hep G2 Cells

  • Zhang, Sheng-Chang;Liang, Ming-Kang;Huang, Guang-Lin;Jiang, Kui;Zhou, Su-Fang;Zhao, Shuang
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.3
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    • pp.1193-1196
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    • 2014
  • Senescence marker protein 30 (SMP30), a hepatocellular carcinoma (HCe) associated antigen had been identified by our research group. To study its mechanisms of regulation and associations with the occurrence and development of HCe, we inhibited expression by RNAi technique, and observed effects on the biological characteristics of Hep G2 cells. In cell viability assays, cell growth in the experimental group (with siRNA transfection) was elevated. In Transwell invasion assays, compared with blank and control groups, numbers of invading cells in the experimental group were significantly increased, whereas in apoptosis assays, the percentage apoptosis demonstrated no differences, but after UV irradiation, that in the experimental group was higher than the other two groups. In a word, SMP30 can inhibit the proliferation and invasion of human hepatoma cells and thus can be regarded as a cancer suppressive factor.

Roles of GST-π and polβ Genes in Chemoresistance of Esophageal Carcinoma Cells

  • Tang, Yue;Xuan, Xiao-Yan;Li, Min;Dong, Zi-Ming
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.12
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    • pp.7375-7379
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    • 2013
  • The main aim of this study was to investigate the roles of GST-${\pi}$ and $pol{\beta}$ genes in the chemoresistance of esophageal carcinoma cells. Eukaryotic expression vectors containing each gene were constructed and transfected into EC9706 cells, and the biological effects of the two genes assessed based on a resistance index. We additionally investigated the in vitro and in vivo anti-resistance effects of GST-${\pi}$ and $pol{\beta}$ genes using recombinant lentiviruses carrying siRNAs against the two genes. Our results showed that upregulation of GST-${\pi}$ and $pol{\beta}$ genes suppresses chemosensitivity of esophageal carcinoma cells to cisplatin, while downregulation of these two genes with RNAi technology reverses this chemoresistance. Multi-site injection of recombinant lentivirus targeting the GST-${\pi}$ gene into transplanted cDDP tumors effectively reversed their chemoresistant phenotype. However, the same treatment against the $pol{\beta}$ gene did not lead to significant efficacy against chemoresistance.

Depletion of PDCD4 Accelerates Stress Granule Assembly Through Sensitization of Stress Response Pathways

  • Kim, Jeeho;Chang, In Youb;Lee, Wooje;Ohn, Takbum
    • Journal of Integrative Natural Science
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    • v.12 no.4
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    • pp.127-132
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    • 2019
  • Programmed cell death 4 (PDCD4) is a novel tumor suppressor that function in the nucleus and the cytoplasm and appears to be involved in the regulation of transcription and translation. Stress granules (SGs) are cytoplasmic foci at which untranslated mRNAs accumulate when cells exposed to environmental stresses. Since PDCD4 has implicated in translation repression through direct interaction with eukaryotic translation initiation factor 4A (eIF4A), we here investigated if PDCD4 has a functional role in the process of SG assembly under oxidative stresses. Using immunofluorescence microscopy, we found that PDCD4 is localized to SGs under oxidative stresses. Next, we tested if knockdown of PDCD4 has an effect on the assembly of SG using PDCD4-specific siRNA. Interestingly, SG assembly was accelerated and this effect was caused by sensitization of phosphorylation of eIF2α and dephosphorylation of eIF4E binding protein (4E-BP). These results suggest that PDCD4 has an effect on SG dynamics and possibly involved in cap-dependent translation repression under stress conditions.

Comparison of Immuno-Suppressive Activities of Pinitol Isolated from Soybean (콩으로부터 추출한 Pinitol의 면역억제 활성 비교)

  • Park, Chul-Hong;Heo, Jin-Chul;Nam, Dong-Yun;Lee, Si-Rim;Nam, So-Hyun;Son, Min-Sik;Hwang, Young-Hyun;Lee, Sang-Han
    • Food Science and Preservation
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    • v.16 no.3
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    • pp.449-453
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    • 2009
  • The experiment was conducted to validate anti-inflammatory effects of pinitol from bean. It was evaluated for some molecule targets by wound healing assay and RT-PCR. The results of wound healing assay was shown dose-dependent inhibition of cell migration in cancer cells and inhibited RNA expression of ICAM-1, CD 44, MMP-17, MMP-14 and ARF2. Immune suppression activity in a mouse provoked by DNFB observed that inflammatory reaction with pinitol were reduced ear swelling and inflammatory cells infiltration in mouse atopic models. The result confirmed that pinitol have the effect of dose-dependent immune suppression activity.

Effect of Bee Venom Death Receptor Dependent Apoptosis and JAK2/STAT3 Pathway in the Ovarian Cancer (난소암에서 봉독이 세포자멸사와 JAK2/STAT3 Pathway의 억제에 미치는 영향)

  • Ahn, Byeong-Joon;Song, Ho-Sueb
    • Journal of Acupuncture Research
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    • v.29 no.1
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    • pp.47-59
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    • 2012
  • 목적 : 이 연구는 봉독이 사람의 난소암 세포인 SKOV3와 PA-1에서 death receptor의 발현을 높여 세포자멸사를 촉진함으로써 암세포의 성장을 억제하는지 밝히고자 하였다. 방법 : 난소암의 세포자멸사의 관찰에는 DAPI, TUNEL staining assay를 시행하였으며, 세포자멸사 조절 단백질의 변동 관찰에는 western blot analysis를 시행하였고, 난소암 세포에서 death receptor의 변화를 관찰하기 위해 RT-PCR analysis를 시행하였다. 결과 : 1. DAPI, TUNEL staining assay 결과, 봉독은 투여량에 따라 세포자멸사의 유도를 통해 SKOV3와 PA-1 난소암세포의 증식을 억제하였고, 세포자멸사와 동반하여 DR4와 DR6의 발현이 두 암세포 모두에서 증가하였고, DR3의 출현은 PA-1 세포에서 증가하였다. 2. Death Receptor의 발현 증가에 따라 caspase-3, 8, 9 and Bax를 포함하는 세포자멸사 촉진 단백질의 발현이 동반하여 상승하였고 JAK2, STAT3의 인산화와 Bcl-2의 발현은 억제되었다. 3. siRNA 처리 시 봉독에 의한 DR3, DR4, DR6 발현증가와 STAT3의 활성억제가 역전되었다. 결론 : 이러한 결과는 봉독이 난소암 세포에서 DR3, DR4, DR6의 증가와 JAK2/STAT3 pathway의 억제를 통하여 세포자멸사를 유발한다는 것을 시사하며, 난소암의 예방과 치료에 효과적으로 활용될 수 있을 것으로 기대된다.