The kinase Activity of N-Acetylglucosamine Kinase is not Essential for the Role of the Enzyme in the Development of Neuronal Dendrites

HyunSook Lee,Il Soo Moon 한국생명과학회 2008 한국생명과학회 심포지움 Vol.49 No.-

Identification of Potential Substrates of N-acetylglucosamine Kinase by a Proteomic Approach

HyunSook Lee(이현숙),Il Soo Moon(문일수) 한국생명과학회 2013 생명과학회지 Vol.23 No.4

단백질 번역 후 O-GlcNAc 수식은 단백질 조절의 새로운 기전으로 대두되고 있다. 전통적인 당수식과 달리 O-GlcNAc 수식은 단 한번의 O-GlcNAc 전달로 이루어지며, 핵 및 세포질단백질 모두에 수식될 수 있다. O-GlcNAc은 이 분자를 끝으로 하는 최종수식으로 생각되어 왔으나, 최근의 논문(J Proteome Res. 201110:2725-2733)은 AP180 단백질에 O-GlcNAc-P가 존재함을 보고하였다. 이 논문은 O-GlcNAc-P가 일반적인 단백질 수식인지에 대한 중요한 질문을 던진다. 이에 답하고자 저자들은 HEK293T 세포에 O-GlcNAc 인산화효소 NAGK를 DsRed2에 연결한 DsRed2-NAGKWT 혹은 효소활성이 없는 돌연변이 NAGK를 표현하는 DsRed2-NAGKD107A를 표현시키고, 단백질 추출물을 얻어 2D-PAGE로 분리한 후 인산화 정도를 측정하여, NAGKWT에 의하여 인산화가 증가되는 15개의 단백질 스폿을 선별하였다. 이 가운데 7개 스팟을 동정한 결과 2개의 스폿은 O-GlcNAc 수식 단백질인 HSP90β, 다른 2개의 스폿도 O-GlcNAc 수식 단백질인 ENO1로 동정되었으며, 나머지(dUTP nucleotidohydrolase mitochondrial isoform 2, glutathione S-transferase P, grp94)는 O-GlcNAc수식 여부를 아직 모르는 단백질이였다. NAGK에 의하여 O-GlcNAc 단백질의 인산화가 증가된다는 사실은 O-GlcNAc이 인산화되어 O-GlcNAc-P로 수식됨을 시사하며, 따라서 본 연구의 결과는 O-GlcNAc이 최종 수식이 아님을 지지한다. Post-translational O-GlcNAc modification (O-GlcNAcylation) of serine or threonine is a new protein modulation mechanism. In contrast to the classical glycosylation, O-GlcNAcylation occurs in a onestep transfer of O-GlcNAc on both nuclear and cytoplasmic proteins. In contrast to the general consensus that O-GlcNAc is a final modification, a recent paper (J Proteome Res. 2011 10:2725?2733) showed the presence of O-GlcNAc-P on a synaptic assembly protein AP180. This finding raises a fundamental question about its prevalence. To address this question, we used proteomics to identify those proteins that were phospho-signal enriched by GlcNAc kinase (NAGK). Comparison of pDsRed2-NAGKWT-transfected HEK293T cell extract with pDsRed2-NAGKD107A-transfected control culture revealed 15 phospho-signal increased spots. Excluding those spots that had no detectable amount of protein expression yielded 7 spots, which were selected for ID determination. Among these, two duplicate spots (two HSP90β and two ENO1 spots) were shown to be O-GlcNAcylated, two (dUTP nucleotidohydrolase mitochondrial isoform 2, glutathione S-transferase P) were not known to be involved in OGlcNAcylation, and one (heat shock protein gp96 precursor or grp94) was a glycoprotein. The increase in the phospho-levels of O-GlcNAc by NAGK strongly indicates that these proteins are phosphorylated on O-GlcNAc. Our present data support the idea that O-GlcNAc is not a terminal modification.

From Epistemic Verbs to Concessives

Hyunsook Lee(이현숙) 담화·인지언어학회 2010 담화와 인지 Vol.17 No.3

In English, there are many markers to express concessive relationships. Among them, this paper aims at verbal origin concessives considering, seeing, granting, and supposing. They are all derived from verbs and acquire grammatical status as concessives. Users of language appear to opt for a different linguistic realization of the same concept (Hoffman 2005). Verbal origin concessives are not frequently used as much as although, though, in spite of, and a few others. However, because those items of content source have grammaticality and are used by speakers as one of the methods to denote the sense of concession, they should be shed light on in terms of grammaticalization. The present investigation, therefore, will look into the actual process in which English concessive markers derived from verbal origin are grammaticalized. First, items in question will be examined about grammaticality and the effect of analogy, and then diachronic semantic changes will be discussed through a perspective shift, subjectification and metonymy.

Cydonia oblonga Miller fruit extract exerts anti-obesity effect in 3T3-L1 adipocyte by upregulating the AMPK signaling pathway

Lee, Hyunsook 한국생활과학회 2023 한국생활과학회 학술대회 Vol.2023 No.5

Vasculogenic property of endothelial colony forming cells combined with mesenchymal progenitor cells in high glucose condition.

LEE Hyunsook,KANG Kyu-Tae 한국특수교육학회 2014 한국특수교육학회 학술대회 Vol.2014 No.-

Control of Epidermal cell proliferation by p63, a homologue of tumor suppressor p53

Lee, Hyunsook 이화여자대학교 세포신호전달연구센터 2002 고사리 세포신호전달 심포지움 Vol. No.4

Epidermal stem cells playa critical role in producing the multi-layered vertebrate skin. Products of the p63 gene not only mark the epidermal stem cells but are absolutely required for the formation of mammalian epidermis. We find that early zebrafish embryos express a dominant negative form of p63(ΔNp63), which accumulates in the nucleus just as epidermal growth begins. Using antisense morpholino oligonucleotides, we show that ΔNp63 is needed for epidermal growth and limb development, and is specifically required for the proliferation of epidermal cells by inhibiting p53 activity. In cultured mammalian cells, ΔNp63α is degraded upon UV exposure independent from p53 status, suggesting a novel role of ΔNp63α in UV-induced DNA damage response pathway. We postulate a model that in epithelial cells, balance between ΔNp63α and p53 is absolutely critical in the maintenance of epidermal stem cell population. When this balance is perturbed, malignant tumors may arise.

Microarray Analysis of Gene Expression by Rhei Rhizoma Water Extracts in a Hypoxia Model of Cultured Neurons

HyunSook Lee(이현숙),Jinyoung Song(송진영),Il Soo Moon(문일수) 한국생명과학회 2009 생명과학회지 Vol.19 No.1

대황(Rhei Rhizoma; RR, 大黃)은 Rheum officinale Baill.와 Rheum palmatum L. (Polygonaceae)의 땅속부분으로 남아시아의 민속의학에서 간 및 신장의 손상을 치료하는데 널리 이용되고 있다. 본 연구에서는 배양한 흰쥐 해마신경세포의 저산소증모델을 이용하여 대황의 물추출물이 유전자 표현에 미치는 영향을 microarray 방법을 이용하여 조사하였다. 배양 후 10일(DIV10)에 추출물을 배지에 2.5 μg/ml 농도로 첨가하고, DIV13에 저산소증(2% O₂/5% CO₂, 37℃, 3 h)을 유발한 후 24시간 후에 total RNA를 분리하여 microarray에 사용하였다. MA-plot에 의하면 표현이 변화된 대부분의 유전자는 ±2배 이내로 증감되었다. 이 가운데 Global M 값이 0.2(즉, 15%)보다 더 증가한 유전자는 472종, Global M 값이 -0.2(즉, -15%)보다 더 감소한 유전자는 725종이였다. 세포의 생존과 관련된 유전자가운데 세포자연사 억제유전자인 Tegt (2.4배), Nfkb1 (2.4배), Veg (1.8배), Ngfr (1.6배) 등이 크게 증가하였으며, 반면에 자연사 촉진유전자인 Bad (-64%), Cstb (-66%)는 감소하였다. 스트레스를 극복하는데 필요한 유전자인 Defb3 (2.7배), Cygb (2.2배), Ahsg (2.18배), Alox5 (2배) 등도 크게 증가하였다. 그리고 세포 성장을 촉진하는 유전자인 Erbb2 (1.84배), Mapk12 (1.8배)도 크게 증가하였다. 따라서 대황의 물추출물은 세포생존에 필요한 유전자를 증가시키고, 세포사를 유도하는 유전자는 감소시킴으로서 저산소증 스트레스에서 신경세포의 사망을 억제하는 것으로 해석된다. In this study, we investigated the effect of Rhei Rhizoma (RR; 大黃) water extract on gene expression in a hypoxia model of cultured rat hippocampal neurons. RR water extract (2.5 μg/ml) was added to the culture media on day 10 in vitro (DIV10), and a hypoxic shock (2% O₂/5% CO₂ , 37℃, 3 h) was given on DIV13. After maintaining the cultures in normoxia for 24 hr, total RNA was isolated and used for microarray analysis. The MA-plot indicated that most genes were up- or downregulated within 2-fold. There were more downregulated genes (725 ea) than upregulated ones (472 ea) when larger than Global M value 0.2 (i.e., >15% increase) or smaller than Global M value -0.2 (i.e., >15% decrease) were considered. Antiapoptosis genes such as Tegt (2.4-fold), Nfkb1 (2.4-fold) Veg (1.8-fold), Ngfr (1.6-fold) were upregulated, while pro-apoptosis genes such as Bad (-64%), Cstb (-66%) were downregulated. Genes for combating environmental stress (stress response genes) such as Defb3 (2.7-fold), Cygb (2.2-fold), Ahsg (2.18-fold), Alox5 (2-fold) were upregulated. Genes for cell proliferation (cell cycle-related genes) such as Erbb2 (1.84-fold), Mapk12 gene (1.8-fold) was upregulated. Therefore, RR water extracts upregulate many pro-survival genes while downregulating many pro-death genes. It is interpreted that these genes, in combination with other regulated genes, can promote neuronal survival in a stress such as hypoxia.

Advanced tube formation assay using human endothelial colony forming cells for in vitro evaluation of angiogenesis

Lee, Hyunsook,Kang, Kyu-Tae The Korean Society of Pharmacology 2018 The Korean Journal of Physiology & Pharmacology Vol.22 No.6

The tube formation assay is a widely used in vitro experiment model to evaluate angiogenic properties by measuring the formation of tubular structures from vascular endothelial cells (ECs). In vitro experimental results are crucial when considered the advisability of moving forward to in vivo studies. Thus, the additional attentions to the in vitro assay is necessary to improve the quality of the pre-clinical data, leading to better decision-making for successful drug discovery. In this study, we improved the tube formation assay system in three aspects. First, we used human endothelial colony forming cells (ECFCs), which are endothelial precursors that have a robust proliferative capacity and more defined angiogenic characteristics compared to mature ECs. Second, we utilized a real-time cell recorder to track the progression of tube formation for 48 hours. Third, to minimize analysis error due to the limited observation area, we used image-stitching software to increase the microscope field of view to a $2{\times}2$ stitched area from the $4{\times}$ object lens. Our advanced tube formation assay system successfully demonstrated the time-dependent dynamic progression of tube formation in the presence and absence of VEGF and FGF-2. Vatalanib, VEGF inhibitor, was tested by our assay system. Of note, $IC_{50}$ values of vatalanib was different at each observation time point. Collectively, these results indicate that our advanced tube formation assay system replicates the dynamic progression of tube formation in response to angiogenic modulators. Therefore, this new system provides a sensitive and versatile assay model for evaluating pro- or anti-angiogenic drugs.

CT Examinations for COVID-19: A Systematic Review of Protocols, Radiation Dose, and Numbers Needed to Diagnose and Predict

Lee Jong Hyuk,Hong Hyunsook,Kim Hyungjin,Lee Chang Hyun,Goo Jin Mo,Yoon Soon Ho 대한영상의학회 2021 대한영상의학회지 Vol.82 No.6

Purpose Although chest CT has been discussed as a first-line test for coronavirus disease 2019 (COVID-19), little research has explored the implications of CT exposure in the population. To review chest CT protocols and radiation doses in COVID-19 publications and explore the number needed to diagnose (NND) and the number needed to predict (NNP) if CT is used as a first-line test. Materials and Methods We searched nine highly cited radiology journals to identify studies discussing the CT-based diagnosis of COVID-19 pneumonia. Study-level information on the CT protocol and radiation dose was collected, and the doses were compared with each national diagnostic reference level (DRL). The NND and NNP, which depends on the test positive rate (TPR), were calculated, given a CT sensitivity of 94% (95% confidence interval [CI]: 91%–96%) and specificity of 37% (95% CI: 26%–50%), and applied to the early outbreak in Wuhan, New York, and Italy. Results From 86 studies, the CT protocol and radiation dose were reported in 81 (94.2%) and 17 studies (19.8%), respectively. Low-dose chest CT was used more than twice as often as standarddose chest CT (39.5% vs.18.6%), while the remaining studies (44.2%) did not provide relevant information. The radiation doses were lower than the national DRLs in 15 of the 17 studies (88.2%) that reported doses. The NND was 3.2 scans (95% CI: 2.2–6.0). The NNPs at TPRs of 50%, 25%, 10%, and 5% were 2.2, 3.6, 8.0, 15.5 scans, respectively. In Wuhan, 35418 (TPR, 58%; 95% CI: 27710–56755) to 44840 (TPR, 38%; 95% CI: 35161–68164) individuals were estimated to have undergone CT examinations to diagnose 17365 patients. During the early surge in New York and Italy, daily NNDs changed up to 5.4 and 10.9 times, respectively, within 10 weeks. Conclusion Low-dose CT protocols were described in less than half of COVID-19 publications, and radiation doses were frequently lacking. The number of populations involved in a first-line diagnostic CT test could vary dynamically according to daily TPR; therefore, caution is required in future planning.

Deep Seawater Increases Dendritic Branches of Cultured Rat Hippocampal Neurons

HyunSook Lee(이현숙),Kyung Soo Nam(남경수),Yun Hee Shon(손윤희),Il Soo Moon(문일수) 한국생명과학회 2008 생명과학회지 Vol.18 No.6

해양심층수(deep seawater, DSW)는 청정성과 무기물질의 풍부함 때문에 여러 분야에 응용하기 위하여 최근 많은 관심을 받고 있다. 본 연구에서는 동해 양양 부근의 해저 1,100 m에서 취수하여 역삼투압 시스템으로 탈염과 농축을 한 심층수가 배양한 흰쥐해마신경세포의 형태적 분화에 미치는 영향을 조사하였다. 10%(v/v) fetal bo-vine serum이 첨가된 MEM 배지에서 키운 세포와 비교할 때 25%(v/v) DSW이 포함될 경우 배양 17시간째에는 차이가 없었다. 그러나 DIV3, 7, 14, 및 17에 관찰하면 경도 0 및 200의 DSW가 포함된 배지에서 자란 신경세포는 가지돌기의 수가 현저히 줄었다. 반면에 경도 600의 DSW에서 자란 신경세포는 그 가지돌기의 수가 대조군과 비슷하였으며, 경도 1000의 경우는 대조군에 비하여 거의 2배 증가하였다. 이 결과는 적당한 경도의 DSW는 신경세포의 성장 및 건강을 돕는 것으로 해석된다. Deep seawater (DSW; deep ocean water) is pure, rich in inorganic materials which have attracted attention for various applications. In this study we investigated the effects of the DSW upwelled from the East Sea, offshore Yang Yang (Korea) on the morphological differentiation of cultured rat hippocampal neurons, which were grown in the minimal essential medium containing 10% (v/v) fetal bo-vine serum and 25% (v/v) DSW with various hardness. DSW had no effect on initial morphological differentiation (17 hr post-plating). When observed on DIV3, 7, 14, and 17, low hardness (0 and 200) DSW reduced dendritic branching. However, dendritic branches within 80 μm diameter from the center of soma nearly doubled in neurons grown in hardness 1,000 DSW-containing media. DSW with hardness 600 was more or less same as control groups. These results indicate that DSW with appropriate hardness ameliorates neuronal health.