• BMB Reports
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• 제46권4호
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• pp.219-224
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• 2013

The cell-fate specification of the anchor cell (AC) and a ventral uterine precursor cell (VU) in Caenorhabditis elegans is initiated by a stochastic interaction between LIN-12/Notch receptor and LAG-2/Delta ligand in two neighboring Z1.ppp and Z4.aaa cells. Both cells express lin-12 and lag-2 before specification, and a small difference in LIN-12 activity leads to the exclusive expressions of lin-12 in VU and lag-2 in the AC, through a feedback mechanism of unknown nature. Here we show that the expression pattern of lag-1/CSL, a transcriptional repressor itself that turns into an activator upon binding of the intracellular domain of Notch, overlaps with that of lin-12. Site-directed mutagenesis of LAG-1 binding sites in lag-1 maintains its expression in the AC, and eliminates it in the VU. Thus, AC/VU cell-fate specification appears to involve direct regulation of lag-1 expression by the LAG-1 protein, activating its transcription in VU cells, but repressing it in the AC.

• The Plant Pathology Journal
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• 제27권1호
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• pp.20-25
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• 2011

Gibberella zeae (anamorph: Fusarium graminearum), a homothallic (self-ferile) ascomycete with ubiquitous geographic distribution, causes serious diseases in several cereal crops. Ascospores (sexual spores) produced by this fungal pathogen have been suggested as the main source of primary inoculum in disease development. Here, we report the function of a gene designated GzRUM1, which is essential for ascospore formation in G. zeae. The deduced product of GzRUM1 showed significant similarities to the human retinoblastoma (tumor suppressor) binding protein 2 and a transcriptional repressor, Rum1 in the corn smut fungus (Ustilago maydis). The transcript of GzRUM1 was detected during the both vegetative and sexual stages, but was more highly accumulated during the latter stage. In addition, no GzRUM1 transcript was detected in a G. zeae strain lacking a mating-type gene (MAT1-2), a master regulator for sexual development in G. zeae. Targeted deletion of GzRUM1 caused no dramatic changes in several traits except ascospore formation. The ${\Delta}$GzRUM1 strain produced perithecia (sexual fruit bodies) but not asci nor ascospores within them. This specific defect leading to an arrest in ascospore development suggests that GzRUM1, as Rum1 in U. maydis, functions as a transcriptional regulator during sexual reproduction in G. zeae.

• 대한화장품학회지
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• 제40권4호
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• pp.403-412
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• 2014

효능이 우수한 신규 미백 소재 개발을 위하여, 민간 및 전통 미백 처방에 사용되어 온 오이(Cucumis sativus L.)에서 활성 물질 분획 추적 연구를 통하여 cucurbitacin B를 분리 정제하고, cucurbitacin B의 멜라닌 합성에 미치는 효과를 B16F1 멜라노마 세포를 이용해 확인하였다. Cucurbitacin B는 세포 독성을 보이지 않는 농도에서 실험한 결과, 멜라닌 생합성을 농도 의존적으로 감소시켰다. Cucurbitacin B는 mushroom tyrosinase의 활성은 직접적으로 저해하지 않았지만, 세포에 처리했을 때 세포 내의 tyrosinase의 활성을 감소시킴을 확인하였다. 또한, cucurbitacin B의 이러한 멜라닌 합성 저해의 기전 연구를 위하여, 멜라닌 합성에 중요한 단백질인 tyrosinase와 microphthalmia-associated transcription factor (MITF)의 단백질 발현을 조사한 결과, cucurbitacin B가 tyrosinase와 MITF의 단백질의 발현을 농도 의존적으로 감소시키는 결과를 확인하였다. 또한, cucurbitacin B는 자외선에 의한 피부암 발생 억제인자(tumor repressor) 및 $Wnt/{\beta}$-catenin 신호전달 과정에 대한 억제 기능이 밝혀진 WW domain-containing oxidoreductase (WWOX)의 단백질 발현을 증가시킴을 추가적으로 확인하였다. 따라서, 이상의 연구 결과를 통해, 오이에서 분리 정제된 cucurbitacin B는 멜라닌 세포(melanocytes)에서 멜라닌 합성을 저해하는 효능이 있음을 확인하였으며, 향후 피부 미백 소재로 활용될 수 있을 것으로 판단된다.

• International Journal of Industrial Entomology and Biomaterials
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• 제12권2호
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• pp.57-61
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• 2006

Suppressor of cytokine signaling (SOCS) is known to be as a negative feedback regulator in Janus kinase signal transducer and activator of transcription signaling. Highly conserved SOCS box domain was cloned from a Korean malaria vector, Anopheles sinensis. Sequence analysis indicates that it has identity to Anopheles gambiae (96%), Aedes aegypti (94%), Drosophila melanogaster (78%), Mus musculus (72%) and Homo sapiens (72%), respectively. Tissue specificity RT-PCR demonstrated that the expression level of AsSOCS transcript was high at abdomen, midgut, and ovary, whereas developmental expression patterns showed that the level of AsSOCS was high at egg, early pupae, and adult female. On the other hand, RT-PCR analysis after bacterial challenge showed that SOCS mRNA was strongly induced in larvae. In addition, it was also induced by various immune elicitors such as lipoteicoic acid, CpG-DNA, and laminarin. It seems that AsSOCS, repressor of JAK-STAT pathway, is highly conserved in mosquito, and may play an important role in mosquito innate immune response.

• Journal of Microbiology and Biotechnology
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• 제14권6호
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• pp.1249-1255
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• 2004

This study has developed Corynebacterium ammoniagenes (c. ammoniagenes) purine gene transcriptional reporters (purF-lacZ and purE-lacZ) that function in Escherichia coli (E. coli) DH5a. After transformation of a C. ammoniagenes gDNA library into E. coli cells harboring either purF-lacZ or purE-lacZ, C. ammoniagenes clones were obtained that repress purF-lacZ and purE-lacZ gene expression. The potential purE and purF regulatory genes are homologous to the genes encoding transcription regulators, the regulatory subunit of RNA polymerase, and genes for purine nucleotide biosynthesis of various bacteria. The C. ammoniagenes purE-lacZ and purF-lacZ reporters were repressed by adenine and guanine within E. coli, indicating similarity in the regulatory mechanism of purine biosynthesis in C. ammoniagenes and E. coli. Gene regulation of pur-lacZ by adenine and guanine was partly abolished in cells expressing potential purine regulatory genes, indicating functionality of the purine gene regulators in repression of purE-lacZ and purF-lacZ. The purE-lacZ and purF-lacZ reporters can be used for the screening of genes involved in the regulation of the de novo synthesis of the purine nucleotides.

• 한국자기공명학회논문지
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• 제3권2호
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• pp.100-108
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• 1999

Cop protein is the transcription repressor protein in rolling circle replication plasmid. With antisense RNA, Cop protein controls the copy number of plasmid. Cop family proteins have been found in various plasmids. Among Cop family proteins, Cop studied in this paper consists of 55 amino acids (Mw. 6,400), and was known to have trimer structure. Since no structural facts are elucidated, we have carried out preliminary experiments aimed at the elucidation of its three dimensional structure. The secondary structure of Cop is studied by CD and NMR. To solve the aggregation of Cop at high concentration, we tested various detergents and salts. The addition of detergents and salts could not solve the aggregation problem. However, we found that concentration is important in solving the aggregation problem. We knew that 0.18mM in 50mM potassium phosphate without any other ingredients is maximum concentration not to aggregate. Wa also investigated the pH dependence of Cop protein, and knew that Cop protein is more stable in acid state. At various temperatures, 15N-1H HSQC spectra were measured in order to find the optimal experimental condition. To enhance the peak resolution, 3D NOESY-HSQC spectrum is acquired. Since there are NOE peaks in the NH-NH region, we knew that Cop protein has $\alpha$-helical content, which was also confirmed by CD.

• Molecules and Cells
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• 제41권9호
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• pp.842-852
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• 2018

Notch signaling is an evolutionarily conserved pathway and involves in the regulation of various cellular and developmental processes. Ligand binding releases the intracellular domain of Notch receptor (NICD), which interacts with DNA-bound CSL [CBF1/Su(H)/Lag-1] to activate transcription of target genes. In the absence of NICD binding, CSL down-regulates target gene expression through the recruitment of various corepressor proteins including SMRT/NCoR (silencing mediator of retinoid and thyroid receptors/nuclear receptor corepressor), SHARP (SMRT/HDAC1-associated repressor protein), and KyoT2. Structural and functional studies revealed the molecular basis of these interactions, in which NICD coactivator and corepressor proteins competitively bind to ${\beta}-trefoil$ domain (BTD) of CSL using a conserved ${\varphi}W{\varphi}P$ motif (${\varphi}$ denotes any hydrophobic residues). To date, there are conflicting ideas regarding the molecular mechanism of SMRT-mediated repression of CSL as to whether CSL-SMRT interaction is direct or indirect (via the bridge factor SHARP). To solve this issue, we mapped the CSL-binding region of SMRT and employed a 'one- plus two-hybrid system' to obtain CSL interaction-defective mutants for this region. We identified the CSL-interaction module of SMRT (CIMS; amino acid 1816-1846) as the molecular determinant of its direct interaction with CSL. Notably, CIMS contains a canonical ${\varphi}W{\varphi}P$ sequence (APIWRP, amino acids 1832-1837) and directly interacts with CSL-BTD in a mode similar to other BTD-binding corepressors. Finally, we showed that CSL-interaction motif, rather than SHARP-interaction motif, of SMRT is involved in transcriptional repression of NICD in a cell-based assay. These results strongly suggest that SMRT participates in CSL-mediated repression via direct binding to CSL.

• Journal of Microbiology
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• 제38권4호
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• pp.239-244
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• 2000

The cats gene encodes the major catalase in Sreptomyces coelicolor, whose production increases upon H$_2$O$_2$treatment. Besides the previously identified primary promoter (catApl), a minor promoter (catAp2) was newly assigned by S1 nuclease mapping. The catAp2 transcript was observed transiently upon entry into the stationary phase in liquid culture and upon differentiation on solid plates, whereas the level of catApl transcription did not chance significantly during this growth transition. ThecatApl promoter was transcribed by the major vegetative RNA polymerase holoenzyme containing $\sigma$$\^$HrdB/, whereas the catAp2 was transcribed in vitro by the holoenzyme containing $\sigma$$\^$R/ that is activated under oxidative conditions. The cia-element regulating the H$_2$O$_2$-inducibility of catApl was identified within the 23 bp inverted repeat sequence located between -65 and -43 of the catApl promoter. We roamed this sequence HRE (H$_2$O$_2$-responsive Element). The distal half of the inverted repeat was more crucial for H$_2$O$_2$-dependent induction of the catApl transcript than the proximal half. HRE most likely serves as a binding site for the H$_2$O$_2$-responsive repressor CatR.

• Biomolecules & Therapeutics
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• 제29권2호
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• pp.234-247
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• 2021

We used a heterozygous gene deletion library of fission yeasts comprising all essential and non-essential genes for a microarray screening of target genes of the antifungal terbinafine, which inhibits ergosterol synthesis via the Erg1 enzyme. We identified 14 heterozygous strains corresponding to 10 non-essential [7 ribosomal-protein (RP) coding genes, spt7, spt20, and elp2] and 4 essential genes (tif302, rpl2501, rpl31, and erg1). Expectedly, their erg1 mRNA and protein levels had decreased compared to the control strain SP286. When we studied the action mechanism of the non-essential target genes using cognate haploid deletion strains, knockout of SAGA-subunit genes caused a down-regulation in erg1 transcription compared to the control strain ED668. However, knockout of RP genes conferred no susceptibility to ergosterol-targeting antifungals. Surprisingly, the RP genes participated in the erg1 transcription as components of repressor complexes as observed in a comparison analysis of the experimental ratio of erg1 mRNA. To understand the action mechanism of the interaction between the drug and the novel essential target genes, we performed isobologram assays with terbinafine and econazole (or cycloheximide). Terbinafine susceptibility of the tif302 heterozygous strain was attributed to both decreased erg1 mRNA levels and inhibition of translation. Moreover, Tif302 was required for efficacy of both terbinafine and cycloheximide. Based on a molecular modeling analysis, terbinafine could directly bind to Tif302 in yeasts, suggesting Tif302 as a potential off-target of terbinafine. In conclusion, this genome-wide screening system can be harnessed for the identification and characterization of target genes under any condition of interest.

• BMB Reports
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• 제40권5호
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• pp.757-764
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• 2007

Marbling of cattle meat is dependent on the coordinated expression of multiple genes. Cattle dramatically increase their intramuscular fat content in the longissimus dorsi muscle between 12 and 27 months of age. We used the annealing control primer (ACP)-differential display RT-PCR method to identify differentially expressed genes (DEGs) that may participate in the development of intramuscular fat between early (12 months old) and late fattening stages (27 months old). Using 20 arbitrary ACP primers, we identified and sequenced 14 DEGs. BLAST searches revealed that expression of the MDH, PI4-K, ferritin, ICER, NID-2, WDNMI, telethonin, filamin, and desmin (DES) genes increased while that of GAPD, COP VII, ACTA1, CamK II, and nebulin decreased during the late fattening stage. The results of functional categorization using the Gene Ontology database for 14 known genes indicated that MDH, GAPD, and COP VII are involved in metabolic pathways such as glycolysis and the TCA cycle, whereas telethonin, filamin, nebulin, desmin, and ACTA1 contribute to the muscle contractile apparatus, and PI4-K, CamK II, and ICER have roles in signal transduction pathways regulated by growth factor or hormones. The final three genes, NID-2, WDNMI, and ferritin, are involved in iron transport and extracellular protein inhibition. The expression patterns were confirmed for seven genes (MDH, PI4-K, ferritin, ICER, nebulin, WDNMI, and telethonin) using real-time PCR. We found that the novel transcription repressor ICER gene was highly expressed in the late fattening stage and during bovine preadipocyte differentiation. This information may be helpful in selecting candidate genes that participate in intramuscular fat development in cattle.