• 대한임상검사과학회지
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• 제49권4호
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• pp.460-469
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• 2017

열충격 단백질(heat shock proteins, HSPs)은 다양한 종양에서 과발현 되고, 종양이 형성되는 과정이나 그 예후에 영향을 주며, 분자량에 따라 HSP27, HSP60, HSP90, HSP100 등으로 구분한다. Heat shock protein90은 세포 내 불안정한 단백질을 보호하는 역할을 통해 질병의 유지에 기여하는데, 정상 조직에 비해 종양 세포에서 높은 수준으로 발현된다고 보고되었다. 이에 본 연구에서는 우리나라 사망원인 1위인 폐암 중 비소세포 폐암에서 Heat shock protein90 family 발현과 비소세포 폐암환자의 임상적 특징과의 상관관계를 분석하여 종양의 생물학적 표지자로서의 가능성을 조사하였다. HSP90 family의 발현과 임상병리학적 특성 및 생존율과의 상관관계를 분석한 결과 $HSP90{\alpha}$는 림프혈관강 침윤이 되지 않은 환자에서 높은 발현을 보였고(p=0.014), $HSP90{\beta}$ 은 조직학적 형태에서 편평상피세포 암종에서 높은 발현을 보였으며(p=0.003), GRP94 은 분화도가 낮을수록 높은 발현을 나타내었다(p=0.048). 생존율은 $HSP90{\alpha}$, $HSP90{\beta}$, GRP94 모두 발현 차이에 대한 유의성이 없었다. 본 연구를 통해 miRNA-126, miRNA-155, miRNA-200c의 발현은 비소세포 폐암의 진단을 위한 생물학적 표지자 및 예후 인자로서 사용될 수 있을 것으로 사료된다. 그리고 비소세포 폐암의 치료용으로 HSP90 family가 고려되어야 할 것이며, GRP94가 종양의 예후예측을 위한 중요한 인자라 사료된다.

• 대한의생명과학회지
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• 제12권4호
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• pp.419-425
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• 2006

Phospholipase $C-{\gamma}1\;(PLC-{\gamma}1)$ is an important signaling molecule for cell proliferation and differentiation. $PLC-{\gamma}1$ contains two pleckstrin homology (PH) domains, which are responsible for protein-protein interaction and protein-lipid interaction. $PLC-{\gamma}1$ also has two Src homology (SH)2 domains and a SH3 domain, which are responsible for protein- protein interaction. To identity proteins that specifically binds to PH domain of $PLC-{\gamma}1$, we prepared and incubated the glutathione S-transferase(GST)-fused PH domains of $PLC-{\gamma}1$ with COS7 cell lysate. We found that 90 kDa protein specifically binds to PH domain of $PLC-{\gamma}1$. By matrix-assisted laser desorption ionization time of flight-mass spectrometry, the 90 kDa protein revealed to be heat shock protein (Hsp) $90{\beta}$. Hsp $90{\beta}$ is a molecular chaperone that stabilizes and facilitates the folding of proteins that are involved in cell signaling, including receptors for steroids hormones and a variety of protein kinases. To know whether Hsp $90{\beta}$ affects on $PLC-{\gamma}1$ activity, we performed $PIP_2$ hydrolyzing activity of $PLC-{\gamma}1$ in the presence of purified Hsp $90{\beta}$ in vitro. Our results show that the Hsp $90{\beta}$ dose-dependently inhibits the enzymatic activity of $PLC-{\gamma}1$ and further suggest that Hsp $90{\beta}$ regulates cell growth and differentiation via regulation of $PLC-{\gamma}1$ activity.

• 한국미생물·생명공학회지
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• 제32권4호
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• pp.297-306
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• 2004

열 충격단백질(Heat shock protein : HSP)은 온도 스트레스에 대하여 세포 내에서 발현되는 단백질이다. HSP의 중요 분류군의 하나가 HSP90 family 이다. 여러 종류의 포유동물과 조류에서 HSP 유전자 특성에 대한 연구가 많이 진행되었다. 본 연구에서는 넙치(Paralichthys olivaceus)로부터 제조한 넙치 뇌 cDNA 유전자 은행을 이용하여 넙치 HSP90 cDNA 유전자를 분리하여 구성 염기서열의 특성을 밝혀 내었다. 염기서열의 분석결과 넙치의 hsp90$\beta$ 유전자는 2,791 개의 뉴클레오타이드로 구성되어 있고, 726개의 아미노산 잔기가 암호화되어 있었다. 넙치 hsp90$\beta$ 유전자는 European sea bass와 96.6% zebrafish와 92.9%, Atlantic salmon와 92.0%, 그리고 사람과는 89.5%의 염기서열 상동성을 지니고 있었다. 또한 HSP90 아미노산 서열을 바탕으로 척추동물 종들과의 진화계통수를 구축하였다. 넙치 hsp90$\beta$ 유전자의 mRNA의 분포 정도를 RT-PCR를 이용하여 조사하였다. hsp90$\beta$ 유전자는 조사한 모든 조직(뇌, 간, 신장, 근육, 비장)에서 높은 수준으로 발현이 되고 있었다. 또한, 넙치 hsp90$\beta$ 단백질을 대량발현하기 위하여 대장균에서 발현을 유도하였다.

• 한국물환경학회지
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• 제29권2호
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• pp.232-238
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• 2013

Water temperature is key factor influencing growth and reproduction of fish and its increase give rise to various physiological changes including gene expression. Heat shock protein (Hsp), one of the molecular chaperones, is highly conserved throughout evolution and its expression is induced by various stressors such as temperature, oxidative, physical and chemical stresses. Here, we isolated partial cDNA clones encoding 70-kDa Hsp (Hsp70) and $\beta$-actin using reverse transcriptase-PCR (RT-PCR) from gut of Rhynchocypris kumgangensis, a Korean indigenous species and cold-water fish, and investigated expression profiles of Hsp70 under an increase of water temperature using $\beta$-actin as an internal control for RT-PCR. Cloned Hsp70 cDNA of R. kumgangensis showed homology to Ctenopharyngodon idella (96%), Hypophthalmichthys molitrix (96%), Danio rerio (93%) and Oncorhynchus mykiss (81%) Hsp70. Cloned $\beta$-actin cDNA of R. kumgangensis showed homology to D. rerio (98%), H. molitrix (97%), C. idella (97%) and O. mykiss (90%) $\beta$-actin. Both mRNA of Hsp70 and $\beta$-actin were expressed in gut, brain, and liver in R. kumgangensis. Futhermore, expression of Hsp70, in brain, was highly augmented by an increase of water temperature. These results suggest that Hsp70 mRNA expression level in brain can be used as a biological molecular marker to represent physiological stress against an increase of water temperature.

• 한국해양생명과학회지
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• 제3권2호
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• pp.51-58
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• 2018

수온의 변화는 어류의 거의 모든 생리학적 부분에 영향을 미친다. 기후 변화로 인한 수온의 상승은 어류에게 물리적 피해를 줄 수 있다. 이 연구는 최적의 수온(15℃)보다 높은 수온(20℃)에서의 대서양 연어의 건강상태를 평가하기 위해 수행하였다. 간 조직은 열 적응에 중요한 대사기능을 발휘하기에 본 연구에 간 조직을 사용하였다. 생체지표유전자의 개발을 위한 분석 방법으로는 NGS RNAseq 방법을 사용하였고, 생체지표유전자의 발현 양상을 관찰하기 위한 분석 방법으로는 RT-qPCR을 사용하였다. NGS RNAseq 분석을 통해 1,366개의 차별적 발현 유전자를 확인하였으며, 그 중에서 880개의 증가하는 유전자와 486개의 감소하는 유전자를 확인하였다. 생체지표유전자로는 heat shock protein 90 alpha (Hsp90α), heat shock protein 90 beta (Hsp90β) 및 cytochrome P450 1A (CYP1A)을 선정하였는데 이들 유전자는 NGS RNAseq 분석에서 수온의 변화에 민감하게 반응하는 유전자들이었다. 이들 유전자의 RT-qPCR을 통한 발현 양상은 NGS RNAseq 분석과 유사하게 나타났다. 이 연구의 결과는 다른 어종에도 적용할 수 있으며, 산업적으로도 유용하다고 생각된다.

• Molecules and Cells
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• 제20권3호
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• pp.378-384
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• 2005

Estrogen metabolites are carcinogenic. The comparative mitogenic activities of $17{\beta}$-estradiol (E2) and four metabolites, 2-hydroxyestradiol (2-OHE2), 4-hydroxyestradiol (4-OHE2), $16{\alpha}$-hydroxyestrone ($16{\alpha}$-OHE1) and 2-methoxyestradiol (2-ME), were determined in estrogen receptor(ER)-positive MCF-7 human breast cancer cells. Each of the E2 metabolites caused proliferation of the MCF-7 cells, but only E2 and $16{\alpha}$-OHE1 induced a greater than 20-fold increases in transcripts of the progesterone receptor (PR) gene, a classical ER-mediated gene. This suggests that the mitogenic action of E2 and $16{\alpha}$-OHE1 could result from their effects on gene expression via the ER. E2 metabolites altered the expression of E2-regulated proteins including heat shock proteins (Hsps). $16{\alpha}$-OHE1 and 2-ME as well as E2 increased levels of Hsp56, Hsp60, $Hsp90{\alpha}$ and Hsp110 transcripts, and the patterns of these inductions resembled that of PR. Hsp56 and Hsp60 protein levels were increased by all the E2 metabolites. Levels of the transcripts of 3 E2-upregulated proteins (XTP3-transactivated protein A, protein disulfide isomerase-associated 4 protein and stathmin 1) and an E2-downregulated protein (aminoacylase 1) were also affected by the E2 metabolites. These results suggest that the altered expression of Hsps (especially Hsp56 and Hsp60) by E2 metabolites such as E2, $16{\alpha}$-OHE1 and 2-ME could be closely linked to their mitogenic action.

• 대한약침학회지
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• 제22권1호
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• pp.28-34
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• 2019

Background: Breast cancer is a complex, heterogeneous disease and one of the most common malignancies in women worldwide. The efficacy of chemotherapy as an important breast cancer treatment option has been severely limited because of the inherent or acquired resistance of cancer cells. The molecular chaperone heat shock protein 90 (HSP90) upregulated in response to cellular stress is required for functions such as conformational maturation, activation and stability in more than 200 client proteins, mostly of the signaling type. In this study, the expression of HSP90 isoforms including $HSP90{\alpha}$ and $HSP90{\beta}$ in breast cancer cell lines before and after treatment with doxorubicin (DOX) was assessed. Material and Methods: The cell cytotoxicity of DOX in MDA-MB-231 and MCF-7 cell lines was determined using the MTT assay. immunofluorescence and western blotting techniques were used to determine the expression of $HSP90{\beta}$ in the cell lines before and after DOX treatment. Immunofluorescence was also conducted to ascertain the expression of $HSP90{\alpha}$. Results: The MTT assay results showed that the MDA-MB-231 cells ($IC_{50}=14.521{\mu}M$) were more sensitive than the MCF-7 cells ($IC_{50}=16.3315{\mu}M$) to DOX. The immunofluorescence results indicated that the expression of $HSP90{\alpha}$ in both cell lines decreased after exposure to DOX. The western blot and immunofluorescence analyses showed that $HSP90{\beta}$ expression decreased in the MCF-7 cells but increased in the MDA-MB-231 cells after DOX treatment. Conclusion: The obtained results suggested that $HSP90{\alpha}$ and $HSP90{\beta}$ expression levels were reduced in the MCF-7 cells after exposure to DOX. In the MDA-MB-231 cells, $HSP90{\alpha}$ expression was reduced while $HSP90{\beta}$ was found to be overexpressed following DOX treatment.

• 약학회지
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• 제51권4호
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• pp.264-269
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• 2007

Alzheimer’s disease (AD) is characterized pathologically by the presence of intracellular neurofibrillary tangles and deposition of ${\beta}-amyloid $ (A ${\beta}$) peptides, which are generated by processing of amyloid precursor protein (APP). It is urgent to develop effective therapies for the treatment of AD, since our society rapidly accelerate aging. A${\beta}$ peptides have been believed to be neurotoxic and now are also considered to have effects on the mechanism of memory formation. In this study, effects of radicicol on the metabolism of APP were analyzed. Radicicol inhibited the secretion of A${\beta}$ from the Neuro2a cell line (APPswe cell) expressing APPswe. Beta-site APP cleaving enzyme (BACE) fluorescence resonance energy transfer (FRET) assay revealed that it inhibited BACE activity in a dose dependently manner. Immunoblotting study showed that it inhibited intracellular heat shock protein (HSP)90 and it increased the secretion of HSP90 from the APPswe cells. We suggest that radicicol inhibits APP metabolism and Ap generation by the means of HSP90 inhibitory mechanism and partially BACE inhibitory mechanism. This is the first report that radicicol inhibits the secretion of A${\beta}$ peptides from neuroblastoma cells.

• BMB Reports
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• 제46권1호
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• pp.47-52
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• 2013

Intestinal epithelial cell (IEC) apoptosis induced by hypoxia compromise intestinal epithelium barrier function. Both Akt and Hsp90 have cytoprotective function. However, the specific role of Akt and $Hsp90{\beta}$ in IEC apoptosis induced by hypoxia has not been explored. We confirmed that hypoxia-induced apoptosis was reduced by $Hsp90{\beta}$ overexpression but enhanced by decreasing $Hsp90{\beta}$ expression. $Hsp90{\beta}$ overexpression enhanced BAD phosphorylation and thus reduced mitochondrial release of cytochrome C. Reducing $Hsp90{\beta}$ expression had opposite effects. The protective effect of $Hsp90{\beta}$ against apoptosis was negated by LY294002, an Akt inhibitor. Further study showed that Akt phosphorylation was enhanced by $Hsp90{\beta}$, which was not due to the activation of upstream PI3K and PDK1 but because of stabilization of pAkt via direct interaction between $Hsp90{\beta}$ and pAkt. These results demonstrate that $Hsp90{\beta}$ may play a significant role in protecting IECs from hypoxia-induced apoptosis via stabilizing pAkt to phosphorylate BAD and reduce cytochrome C release.

• 한국가금학회지
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• 제46권2호
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• pp.77-86
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• 2019

본 연구는 겨울철에 발생하여 육성된 닭들과 여름철에 발생하여 육성된 닭들 간의 열 스트레스 반응 정도와 생산능력을 비교 분석하고자 하였다. 공시계로는 겨울철에 발생된 한국토종종계 초생추 1,156수와 여름철에 발생된 초생추 934수로 총 2,090수를 분석 대상으로 하였다. 스트레스 반응정도와 생산능력을 비교하기 위하여 텔로미어의 함량과 heat shock proteins(HSPs)의 유전자 발현율을 분석하고, 생존율, 산란율 및 체중을 조사하였다. 분석 결과, HSP-70, $HSP-90{\alpha}$ 및 $HSP-90{\beta}$ 유전자 발현율은 겨울철에 발생하여 육성된 닭들이 여름철에 발생하여 육성된 닭들에 비하여 모두 유의하게 높은 발현값을 나타내었다. 텔로미어 함량은 겨울철과 여름철에 발생한 닭들 간에 유의한 차이가 없었다. 생존율에서는 여름철 발생하여 육성된 닭들이 겨울철에 발생하여 육성된 닭들에 비해 유의하게 높았고, 산란율 및 난중 또한 여름철 발생 계군이 높게 나타났다. 반면, 초산일령은 겨울철 발생 계군이 여름철 발생 계군에 비해 빨랐다. 체중에 있어서 24주까지는 겨울철 발생 계군이 여름철 발생 계군에 비해 높았으나, 28주 이후부터 발생 계군 간 역전된 결과를 보였다. 결론적으로 여름철에 발생하여 육성된 닭들이 겨울철에 발생하여 육성된 닭들에 비해 열 스트레스에 대한 저항성이 높고 생산성이 우수함을 보였다. 이는 발생 및 육성 초기에 고온에 노출된 닭들이 상대적으로 높은 열적응성을 습득한 결과로 사료된다.