• Journal of Microbiology and Biotechnology
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• 제24권12호
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• pp.1644-1653
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• 2014

Wuyiencin is produced by Streptomyces ahygroscopicus var. wuyiensis CK-15 and is widely used as an antifungal agent in agriculture. Analysis of wuyiencin biosynthetic gene clusters reveals wysR, a member of the LAL-family of transcriptional regulatory genes. WysR consists of an N-terminal PAS domain and a LuxR family C-terminal helix-turn-helix motif. However, the roles of wysR in wuyiencin biosynthesis are largely unknown. In this study, we showed that inactivation of wysR resulted in the complete loss of wuyiencin production, which could be restored by complementation with a single copy of wysR. Furthermore, we successfully increased wuyiencin production to a significantly higher level by overexpression of wysR in S. wuyiensis CK-15. Quantitative real-time RT-PCR analysis showed that WysR regulates wuyiencin biosynthesis by modulating other putative regulatory genes. Thus, WysR was identified as an activator of wuyiencin biosynthesis, and overexpression of wysR gene proved to be an effective strategy for improving wuyiencin production.

• BMB Reports
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• 제49권8호
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• pp.401-402
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• 2016

S6K1 is a key regulator of cell growth, cell size, and metabolism. Although the role of cytosolic S6K1 in cellular processes is well established, the function of S6K1 in the nucleus remains poorly understood. Our recent study has revealed that S6K1 is translocated into the nucleus upon adipogenic stimulus where it directly binds to and phosphorylates H2B at serine 36. Such phosphorylation promotes EZH2 recruitment and subsequent histone H3K27 trimethylation on the promoter of its target genes including Wnt6, Wnt10a, and Wnt10b, leading to repression of their expression. S6K1-mediated suppression of Wnt genes facilitates adipogenic differentiation through the expression of adipogenic transcription factors PPARγ and Cebpa. White adipose tissues from S6K1-deficient mice consistently exhibit marked reduction in H2BS36 phosphorylation (H2BS36p) and H3K27 trimethylation (H3K27me3), leading to enhanced expression of Wnt genes. In addition, expression levels of H2BS36p and H3K27me3 are highly elevated in white adipose tissues from mice fed on high-fat diet or from obese humans. These findings describe a novel role of S6K1 as a transcriptional regulator controlling an epigenetic network initiated by phosphorylation of H2B and trimethylation of H3, thus shutting off Wnt gene expression in early adipogenesis.

• Asian Pacific Journal of Cancer Prevention
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• 제15권8호
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• pp.3817-3823
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• 2014

Background: The MDM2 oncogene, a negative regulator of p53, has a functional polymorphism in the promoter region (SNP309) that is associated with multiple kinds of cancers including non-melanoma skin cancer. SNP309 has been shown to associate with accelerated tumor formation by increasing the affinity of the transcriptional activator Sp1. It remains unknown whether there are other factors involved in the regulation of MDM2 transcription through a trans-regulatory mechanism. Methods: In this study, SNP309 was verified to be associated with overexpression of MDM2 in tumor cells. Bioinformatics predicts that the T to G substitution at SNP309 generates a stronger E2F1 binding site, which was confirmed by ChIP and luciferase assays. Results: E2F1 knockdown downregulates the expression of MDM2, which confirms that E2F1 is a functional upstream regulator. Furthermore, tumor cells with the GG genotype exhibited a higher proliferation rate than TT, correlating with cyclin D1 expression. E2F1 depletion significantly inhibits the proliferation capacity and downregulates cyclin D1 expression, especially in GG genotype skin fibroblasts. Notably, E2F1 siRNA effects could be rescued by cyclin D1 overexpression. Conclusion: Taken together, a novel modulator E2F1 was identified as regulating MDM2 expression dependent on SNP309 and further mediates cyclin D1 expression and tumor cell proliferation. E2F1 might act as an important factor for SNP309 serving as a rate-limiting event in carcinogenesis.

• Journal of Microbiology and Biotechnology
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• 제30권6호
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• pp.811-821
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• 2020

Mycobacterium tuberculosis produces mycolic acids which are relevant for persistence, recalcitrance to antibiotics and defiance to host immunity. c-di-GMP is a second messenger involved in transition from planktonic cells to biofilms, whose levels are controlled by diguanylate cyclases (DGC) and phosphodiesterases (PDE). The transcriptional regulator dosR, is involved in response to low oxygen, a condition likely happening to a subset of cells within biofilms. Here, we found that in M. bovis BCG, expression of both BCG1416c and BCG1419c genes, which code for a DGC and a PDE, respectively, decreased in both stationary phase and during biofilm production. The kasA, kasB, and fas genes, which are involved in mycolic acid biosynthesis, were induced in biofilm cultures, as was dosR, therefore suggesting an inverse correlation in their expression compared with that of genes involved in c-di-GMP metabolism. The relative abundance within trehalose dimycolate (TDM) of α-mycolates decreased during biofilm maturation, with methoxy mycolates increasing over time, and keto species remaining practically stable. Moreover, addition of synthetic c-di-GMP to mid-log phase BCG cultures reduced methoxy mycolates, increased keto species and practically did not affect α-mycolates, showing a differential effect of c-di-GMP on keto- and methoxy-mycolic acid metabolism.

• 생명과학회지
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• 제24권3호
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• pp.323-328
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• 2014

MicroRNA는 21~25개의 뉴클레오티드로 이루어진 단일 염기가닥의 RNA로 지방분화를 포함한 세포내 여러 생리학적 기전에 영향을 미친다. MicroRNA-17 (miR-17)은 지방전구세포의 지방분화를 촉진하고, 세포 내 지방을 축적시킨다. 그렇지만, 지방분화시 miR-17 전사조절 기전은 알려진 것이 없다. 본 연구에서는 $PPAR{\gamma}$ 전사인자가 miR-17의 전사를 조절하는지 여부를 조사하였다. 먼저 지방전구세포의 분화를 유도한 다음 $PPAR{\gamma}$와 miR-17의 발현을 조사한 결과 두 유전자 모두 분화 이후 발현이 증가하였다. 또한 miR-17 프로모터 부위에는 $PPAR{\gamma}$ 반응하는 부위(PPRE)가 세 군데 발견되었다. Chromatin immunoprecipitation과 luciferase assay을 실시한 결과, miR-17 프로모터의 PPRE3 (-677/-655) 부위가 $PPAR{\gamma}$와 직접 결합함을 관찰하였다. 이러한 연구 결과는 3T3-L1 세포주에서 $PPAR{\gamma}$가 miR-17의 전사를 활성화 시키고 있음을 나타낸다. 향후 $PPAR{\gamma}$에 의한 표적 microRNA을 대량 발굴한다면 비만과 관련한 $PPAR{\gamma}$의 기능을 보다 구체적으로 파악하는데 중요한 정보를 제공할 수 있을 것으로 생각된다.

• 대한소아치과학회지
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• 제29권3호
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• pp.337-344
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• 2002

조골 세포의 분화에 중요한 역할을 하는 전사 인자인 Runx2는 그 역할은 많이 알려져 있지만, 이를 조절하는 신호 전달체계에 대해서는 많이 알려지지 않았다. 이에 본 연구에서는 조골 세포의 분화 및 증식에 영향을 미친다고 알려진 PKC 및 MAPK pathway가 Runx2에 미치는 영향을 알아보고자 하였다. PKC활성화에 따른 Runx2의 전사 활성 및 발현 양상을 관찰하기 위해 6XOSE2-C2C12 cell에 PKC 활성제를 처리하여 luciferase assay와 Northern blot analysis를 시행하였다. MAPK 활성화에 따른 Runx2의 전사 활성을 관찰하기 위해 MAPK 활성제를 6XOSE2-C2C12 cell에 처리하여 luciferase assay를 시행하였다. 두 신호 전달 체계의 활성화에 따른 골 표지 유전자의 전사 양상을 관찰하기 위해 osteocalcin과 osteopontin을 transient transfection한 C2C12 cell에 각 신호 전달 체계의 활성제를 처리하여 luciferase assay를 시행하였다. 또한 각 신호 전달 체계가 상호 작용하는지 알아보기 위하여 MAPK 억제제를 전처리하여 MAPK pathway를 차단한 1 시간 뒤 PKC 활성제를 처리하고 luciferase assay를 시행하여 Runx2의 전사 활성을 관찰하였다. 이상의 실험으로 다음과 같은 결론을 얻었다. - PKC pathway의 활성화는 Runx2의 전사 활성 및 발현을 증가시키고 이로 인해 그의 영향을 받는 골 표지 유전자 (osteopontin, osteocalcin)의 전사도 증가한다. - MAPK pathway의 활성화는 Runx2 및 골 표지 유전자 (osteopontin, osteocalcin)의 전사활성을 증가시킨다. - PKC pathway는 MAPK pathway를 경유하여 Runx2의 전사 활성을 조절한다.

• Tuberculosis and Respiratory Diseases
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• 제78권2호
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• pp.99-105
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• 2015

Background: Aryl hydrocarbon receptor (AhR), a ligand-dependent transcription factor, binds to a wide variety of synthetic and naturally occurring compounds. AhR is involved in the regulation of inflammatory response during acute and chronic respiratory diseases. We investigated whether nuclear receptor coactivator 7 (NCOA7) could regulate transcriptional levels of AhR target genes and inflammatory cytokines in 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-treated human bronchial epithelial cells. This study was based on our previous study that NCOA7 was differentially expressed between normal and chronic obstructive pulmonary disease lung tissues. Methods: BEAS-2B and A549 cells grown under serum-free conditions were treated with or without TCDD (0.15 nM and 6.5 nM) for 24 hours after transfection of pCMV-NCOA7 isoform 4. Expression levels of cytochrome P4501A1 (CYP1A1), IL-6, and IL-8 were measured by quantitative real-time polymerase chain reaction. Results: The transcriptional activities of CYP1A1 and inflammatory cytokines were strongly induced by TCDD treatment in both BEAS-2B and A549 cell lines. The NCOA7 isoform 4 oppositely regulated the transcriptional activities of CYP1A1 and inflammatory cytokines between BEAS-2B and A549 cell lines. Conclusion: Our results suggest that NCOA7 could act as a regulator in the TCDD-AhR signaling pathway with dual roles in normal and abnormal physiological conditions.

• Journal of Microbiology and Biotechnology
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• 제17권8호
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• pp.1344-1352
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• 2007

An opportunistic human pathogen, Pseudomonas aeruginosa, contains the major catalase KatA, which is required to cope with oxidative and osmotic stresses. As an attempt to uncover the $H_2O_2$-dependent regulatory mechanism delineating katA gene expression, four prototrophic $H_2O_2$-sensitive mutants were isolated from about 1,500 TnphoA mutant clones of P. aeruginosa strain PA14. Arbitrary PCR and direct cloning of the transposon insertion sites revealed that one insertion is located within the katA coding region and two are within the coding region of oxyR, which is responsible for transcriptional activation of several antioxidant enzyme genes in response to oxidative challenges. The fourth insertion was within PA3815 (IscR), which encodes a homolog of the Escherichia coli iron-sulfur assembly regulator, IscR. The levels of catalase and SOD activities were significantly reduced in the iscR mutant, but not in the oxyR mutant, during the normal planktonic culture conditions. These results suggest that both IscR and OxyR are required for the optimal resistance to $H_2O_2$, which involves the expression of multiple antioxidant enzymes including KatA.

• Journal of Microbiology
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• 제35권4호
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• pp.277-283
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• 1997

Promoters inducible by paraquat, a superocide-generating agent, were isolated from Escherichia coli using a promoter-probing plasmid pRS415 with promoterless lacA gene. Twenty one promoters induced by paraquat were selected and further characterized. From sequence analysis, thirteen of the promoters were mapped to their specific loci on the Escherichia coli chromosome. Several promoters were mapped to the upstream of known genes such as usgl, katG, and mglB, whose relationships with superoxide response have not been previously reported. Other promoters were mapped to the upstream region of unknown open reading frames. Downstream of HC 96 promoter are uncharacterized ORFs whose sequences are homologous to ABC-transporter subunits. Downstream of HC84 promoter is an ORF encoding a transcriptional regulator-like protein, which contains a LysR family-specific HTH (helix-turn-helix) DNA bindign motif. We investigated whether these promoters belong to the soxRS regulon. All promoters except HC96 were found to belong to the soxRS regulon. The HC96 promoter was significantly induced by paraquat in the soxRS deletion mutant strain. The basal transcription level of three promoters (HE43, HC71, HD94) significantly increased at the stationary phase, implying that they are regulated by RpoS. However, paraquat inducibility of all promoters disappeared in the stationary phase, suggesting that SoxRS regulatory system is active only in rapidly growing cells.

• Molecules and Cells
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• 제24권3호
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• pp.445-451
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• 2007

Translation initiation factor 4E (eIF4E) is a key regulator of protein synthesis. Abnormal regulation of eIF4E is closely linked to oncogenic transformation. Several regulatory mechanisms affecting eIF4E are discussed, including transcriptional regulation, phosphorylation and binding of an inhibitor protein. However it is not clear how the level of eIF4E protein is regulated under basal conditions. Here we demonstrate that Diap1 (Drosophila Inhibitor of Apoptosis Protein), a cell death inhibitor, binds directly to eIF4E and poly-ubiquitinates it via its E3 ligase activity, promoting its proteasome-dependent degradation. Expression of Diap1 caused a reduction of Cyclin D1 protein level and inhibited the growth stimulation induced by overexpression of eIF4E. Taken together, our results suggest that the level of eIF4E protein is regulated by Diap1, and that IAPs may play a role in cap-dependent translation by regulating the level of eIF4E protein.