• IMMUNE NETWORK
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• v.13 no.5
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• pp.213-217
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• 2013

Enterotoxigenic Bacteroides fragilis (ETBF) is a human gut commensal bacteria that causes inflammatory diarrhea and colitis. ETBF also promotes colorectal tumorigenesis in the Min mouse model. The key virulence factor is a secreted metalloprotease called B. fragilis toxin (BFT). BFT induces E-cadherin cleavage, cell rounding, activation of the ${\beta}$-catenin pathway and secretion of IL-8 in colonic epithelial cells. However, the precise mechanism by which these processes occur and how these processes are interrelated is still unclear. E-cadherin form homophilic interactions which tethers adjacent cells. Loss of E-cadherin results in detachment of adjacent cells. Prior studies have suggested that BFT induces IL-8 expression by inducing E-cadherin cleavage; cells that do not express E-cadherin do not secrete IL-8 in response to BFT. In the current study, we found that HT29/C1cells treated with dilute trypsin solution induced E-cadherin degradation and IL-8 secretion, consistent with the hypothesis that E-cadherin cleavage causes IL-8 secretion. However, physical damage to the cell monolayer did not induce IL-8 secretion. We also show that EDTA-mediated disruption of E-cadherin interactions without E-cadherin degradation was sufficient to induce IL-8 secretion. Finally, we determined that HT29/C1 cells treated with LiCl (${\beta}$-catenin activator) induced IL-8 secretion in a dose-dependent and time-dependent manner. Taken together, our results suggest that BFT induced IL-8 secretion may occur by the following process: E-cadherin cleavage, disruption of cellular interactions, activation of the ${\beta}$-catenin pathway and IL-8 expression. However, we further propose that E-cadherin cleavage per se may not be required for BFT induced IL-8 secretion.

• Biomolecules & Therapeutics
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• v.30 no.6
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• pp.593-602
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• 2022

The human papillomavirus (HPV)-18 E7 (E7) oncoprotein is a major transforming protein that is thought to be involved in the development of cervical cancer. It is well-known that E7 stimulates tumour development by inactivating pRb. However, this alone cannot explain the various characteristics acquired by HPV infection. Therefore, we examined other molecules that could help explain the acquired cancer properties during E7-induced cancer development. Using the yeast two-hybrid (Y2H) method, we found that the Elk-1 factor, which is crucial for cell proliferation, invasion, cell survival, anti-apoptotic activity, and cancer development, binds to the E7. By determining which part of E7 binds to which domain of Elk-1 using the Y2H method, it was found that CR2 and CR3 of the E7 and parts 1-206, including the ETS-DNA domain of Elk-1, interact with each other. As a result of their interaction, the transcriptional activity of Elk-1 was increased, thereby increasing the expression of target genes EGR-1, c-fos, and E2F. Additionally, the colony forming assay revealed that overexpression of Elk-1 and E7 promotes C33A cell proliferation. We expect that the discovery of a novel E7 function as an Elk-1 activator could help explain whether the E7 has novel oncogenic activities in addition to p53 inactivation. We also expect that it will offer new methods for developing improved strategies for cervical cancer treatment.

• Journal of Microbiology and Biotechnology
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• v.33 no.7
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• pp.964-972
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• 2023

Bacteriophage endolysins are peptidoglycan hydrolases composed of cell binding domain (CBD) and an enzymatically active domain. A phage endolysin CBD can be used for detecting bacteria owing to its high specificity and sensitivity toward the bacterial cell wall. We aimed to develop a method for detection of Enterococcus faecalis using an endolysin CBD. The gene encoding the CBD of ECP3 phage endolysin was cloned into the Escherichia coli expression vector pET21a. A recombinant protein with a C-terminal 6-His-tag (CBD) was expressed and purified using a His-trap column. CBD was adsorbed onto epoxy magnetic beads (eMBs). The bacterial species specificity and sensitivity of bacterial binding to CBD-eMB complexes were determined using the bacterial colony counting from the magnetic separations after the binding reaction between bacteria and CBD-eMB complexes. E. faecalis could bind to CBD-eMB complexes, but other bacteria (such as Enterococcus faecium, Staphylococcus aureus, Escherichia coli, Acinetobacter baumannii, Streptococcus mutans, and Porphyromonas gingivalis) could not. E. faecalis cells were fixed onto CBD-eMB complexes within 1 h, and >78% of viable E. faecalis cells were recovered. The E. faecalis recovery ratio was not affected by the other bacterial species. The detection limit of the CBD-eMB complex for E. faecalis was >17 CFU/ml. We developed a simple method for the specific detection of E. faecalis using bacteriophage endolysin CBD and MBs. This is the first study to determine that the C-terminal region of ECP3 phage endolysin is a highly specific binding site for E. faecalis among other bacterial species.

• Proceedings of the Korea Society for Energy Engineering kosee Conference
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• 2004.05a
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• pp.103-108
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• 2004

• The Journal of Korean Society of Virology
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• v.26 no.1
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• pp.115-120
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• 1996

Human papillomavirus (HPV), especially type 16 and 18, has been closely associated with carcinomas and uterine cevical cancer, recently. From in vitro assays, E6 and E7 genes of HPV16 are closely linked with transformation of cell lines of rodent fibroplasts. However, the transforming activity of E6 and E7 genes of HPV type 16 in vivo has not been fully elucidated. For explaining this mechanism, we prepared a expression system with the promoter of mouse mammary tumorvirus long terminal repeat and E6E7's open reading frames. This expression system was introduced in rodent cell lines, No. 7, 3Y1 and shown normal transforming abilities. And, we produced transgenic mice with E6, E7 expression system. These transgenic mice were confirmed from Southern blot analysis. One male of them was observed enlargement of the testis after 5 months postdelivery.

• Journal of Microbiology and Biotechnology
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• v.34 no.3
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• pp.506-515
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• 2024

Primary human dermal papilla cells (HDPCs) are often preferred in studies on hair growth and regeneration. However, primary HDPCs are limited by their reduced proliferative capacity, decreased hair induction potential, and extended doubling times at higher passages. To overcome these limitations, pTARGET vectors containing human papillomavirus16 (HPV16) E6/E7 oncogenes were transfected into HDPCs and selected using G-148 to generate immortalized cells here. HPV16 E6/E7 oncogenes were efficiently transfected into primary HDPCs. Immortalized HDPC showed higher proliferative activity than primary HDPC, confirming an increased proliferation rate. Expression of p53 and pRb proteins was downregulated by E6 and E7, respectively. E6/E7 expressing HDPC cells revealed that cyclin-dependent kinase (CDK) inhibitor p21 expression was decreased, while cell cycle-related genes and proteins (CDK2 and cyclin E) and E2F family genes were upregulated. Immortalized HDPCs maintained their responsiveness to Wnt/β-catenin pathway and hair follicle formation capability, as indicated by their aggregative properties and stemness. E6/E7 immortalized HDPCs may facilitate in vitro hair growth and regeneration studies.

• Microbiology and Biotechnology Letters
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• v.22 no.4
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• pp.368-372
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• 1994

Three more unusual macrolides in addition to concnamycin B were isolated from the mycelium of Streptomyces sp. strain Bal6. These four compounds showed a potent cytotoxity to hunian cancer cell lines, SNU-1 (stomach cancer cell line), SNU-354 (liver cancer cell line), MCF- 7 (breast cancer cell line) and KB-3-1 (oral epidermoid carcinoma cell line). Interestingly, these compounds confered slight differential cytotoxity on RHEK-1, a human epidermal keratinocyte cell line immotalized by AD12-SV40 hybrid virus and RHEK-1/pSV$_{2}$ ras which was resulted from H-ras transfomation of RHEK-1. These compounds were determined to be concanamycin A, conca- namycin E and 0-methyl concanamycin B by NMR and other spectral analysis.

• Journal of Industrial and Engineering Chemistry
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• v.67
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• pp.231-235
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• 2018

In this study, we demonstrate the conversion of starch to (R,R)-2,3-butanediol (2,3-BD) in a hybrid cell-free synthesis system containing a mixture of lysates derived from Escherichia coli (E. coli) and cyanobacteria. A sufficient pool of pyruvate required for the synthesis of 2,3-BD was generated by combining metabolic pathways of cyanobacteria and E. coli. Successful synthesis of 2,3-BD was achieved by additional modifications of the hybrid cell-free system with the enzymes required to convert pyruvate to 2,3-BD. The results demonstrate a new approach to harness biological pathways to expand the scope of cell-free metabolic engineering by cross-species combinations of cell lysates.

• Proceedings of the KSAR Conference
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• 2001.03a
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• pp.21-21
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• 2001

일반적으로 세포는 방사능이나 항암제 등의 자극에 의해 DNA가 손상받았을 때 DNA를 합성하기 전, DNA변이를 복구하기 위해 cell cycle을 정지시키게 된다. pRB(retinoblatoma protein)는 이러한 cell cycle의 조절기작에서 중요한 역할을 담당하는 것으로 알려져 있다. G1기에서 S기로 진행하는 것을 조절하는 단백질인 pRB 은 E2F(cell cycle transcription factor)와 상호작용하여 cell cycle 진행에 필요한 전사활성을 억제, PCNA (proliferating cell nuclear antigen)의 합성을 저해한다. 또한, E2F와 결합된 pRB는 apoptosis를 제어하는 유전자를 조절하는 것으로 알려져 있다. Cell cycle에 영향을 미치는 항암제의 일종인 busulfan을 처리하면, 정소 내에 존재하는 대부분의 생식세포들은 사멸되고 spermatogonia만 남는 것으로 알려져 있다. 그러나 그 기작에 대해서는 자세히 연구된 바가 없다. 본 연구에서는 busulfan처리시 spermatogonial stem cell이 어떤 기작에 의해 손상받지 않고 유지되는지를 알아보고자 실험을 수행하였다. Busulfan을 처리한 마우스 (항암제 투여 후 5주)와 정상적인 13주령의 마우스의 정소로부터 각각 세포를 분리하였다. LSC (laser scanning cytometry)를 이용하여 처리군(busulfan treated mice)과 대조군(normal mature mice)에 대해 각각 DNA함량을 비교ㆍ분석한 결과 G0/G1(2N)에 머물러 있는 세포비율이 처리군에서 현저하게 증가했다 (79.3$\pm$5.5%:8.1$\pm$1.3%). Cell cycle의 G1/S check point인 pRB와 PCNA 발현을 Western blot과 면역조직학적인 방법(immunohisto-chemistry)을 이용하여 조사하였다 PCNA는 대조군과 비교해, 처리군에서 매우 낮은 수준으로 발현되었다. 면역염색된 정소단면을 살펴보면, 대조군에서는 모든 세정관에서 PCNA를 발현하는 세포가 높은 비율로 검출되었고, 처리군에서는 소수의 세정관에서 세포들이 낮은 수준으로 검출되었다. 반면에, pRB의 경우 PCNA와는 상반된 결과를 나타내어, 대조군에서는 거의 발현이 되지 않는 반면, 처리군에서는 대부분의 세정관내, 기저막을 따라 위치한 세포들에서 발현되었다. 이상의 결과는 busulfan에 의해 pRB의 인산화가 억제, pRB 와 결합된 E2F는 전사 활성이 억제되어, PCNA 합성을 저해하는 것으로 설명되어질 수 있다. 결론적으로, 인산화가 억제된 pRB (underphosphorylated RB protein)이 quiescent spermatogonial stem cell에서만 특이하게 발현하는 단백질이며, 이러한 pRB의 발현은 apoptosis를 제어하는 역할을 담당해 busulfan처리에 의해 손상받지 않고 남아있는 것으로 시사된다.

• BMB Reports
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• v.42 no.11
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• pp.719-724
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• 2009

Recent studies have revealed that endoplasmic reticulum (ER) disturbance is involved in the pathophysiology of neurodegenerative disorders, contributing to the activation of the ER stress-mediated apoptotic pathway. Therefore, we investigated here the molecular mechanisms underlying the ER-mitochondria axis, focusing on calcium as a potential mediator of cell death signals. Using NT2 cells treated with brefeldin A or tunicamycin, we observed that ER stress induces changes in the mitochondrial function, impairing mitochondrial membrane potential and distressing mitochondrial respiratory chain complex Moreover, stress stimuli at ER level evoked calcium fluxes between ER and mitochondria. Under these conditions, ER stress activated the unfolded protein response by an overexpression of GRP78, and also caspase-4 and-2, both involved upstream of caspase-9. Our findings show that ER and mitochondria interconnection plays a prominent role in the induction of neuronal cell death under particular stress circumstances.