RanBPM Protein Acts as a Negative Regulator of BLT2 Receptor to Attenuate BLT2-mediated Cell Motility

Wei, Jun-Dong,Kim, Joo-Young,Kim, Ae-Kyoung,Jang, Sung Key,Kim, Jae-Hong American Society for Biochemistry and Molecular Bi 2013 The Journal of biological chemistry Vol.288 No.37

<P>BLT2, a low affinity receptor for leukotriene B<SUB>4</SUB> (LTB<SUB>4</SUB>), is a member of the G protein-coupled receptor family and is involved in many signal transduction pathways associated with various cellular phenotypes, including chemotactic motility. However, the regulatory mechanism for BLT2 has not yet been demonstrated. To understand the regulatory mechanism of BLT2, we screened and identified the proteins that bind to BLT2. Using a yeast two-hybrid assay with the BLT2 C-terminal domain as bait, we found that RanBPM, a previously proposed scaffold protein, interacts with BLT2. We demonstrated the specific interaction between BLT2 and RanBPM by GST pulldown assay and co-immunoprecipitation assay. To elucidate the biological function of the RanBPM-BLT2 interaction, we evaluated the effects of RanBPM overexpression or knockdown. We found that BLT2-mediated motility was severely attenuated by RanBPM overexpression and that knockdown of endogenous RanBPM by shRNA strongly promoted BLT2-mediated motility, suggesting a negative regulatory function of RanBPM toward BLT2. Furthermore, we observed that the addition of BLT2 ligands caused the dissociation of BLT2 and RanBPM, thus releasing the negative regulatory effect of RanBPM. Finally, we propose that Akt-induced BLT2 phosphorylation at residue Thr<SUP>355</SUP>, which occurs after the addition of BLT2 ligands, is a potential mechanism by which BLT2 dissociates from RanBPM, resulting in stimulation of BLT2 signaling. Taken together, our results suggest that RanBPM acts as a negative regulator of BLT2 signaling to attenuate BLT2-mediated cell motility.</P>

BLT1 and BLT2: Leukotriene B4 Receptors, Their Characteristics and Role in Various Diseases

박혜준(Hyejun Park),최용석(Yongseok Choi) 고려대학교 생명자원연구소 2021 생명자원연구 Vol.29 No.-

LTB4는 호중구를 포함한 림프구의 모집 및 활성화에 중요한 역할을 하는 지질 매개체이다. LTB4는 두 가지 수용체를 통해 면역세포를 모집하거나 사이토카인을 분비함으로써 면역 반응을 조절한다. LTB4는 염증성 질환과 밀접한 관련이 있다. BLT1 저해제 개발에 대한 연구가 활발히 진행 중이지만 성공한 사례가 거의 없다. BLT2의 역할은 많이 연구되지 않았지만 최근 연구에 따르면 기도 염증과 암에서 염증을 유도하는 데 중요한 역할을 하는 것으로 나타났다. 본 논문에서는 다양한 질병에서의 LTB4와 LTB4 수용체인 BLT1과 BLT2의 특성과 역할에 대한 연구결과들을 살펴보고자 한다. LTB4 is a lipid mediator which plays a principal role in the recruitment and activation of lymphocytes, including neutrophils. LTB4 modulates immune responses by recruiting immune cells or secreting cytokines through two receptors. In addition, LTB4 is closely related to inflammatory diseases. Research on the development of BLT1 inhibitors is actively underway, but with little success. Although the role of BLT2 has not been studied much, recent studies have shown that it plays an important role in inducing inflammation in airway inflammation and cancer. In this paper, a review of the studies on the characteristics and roles of LTB4 and LTB4 receptors, BLT1 and BLT2, in various diseases would be described.

Leukotriene B₄ receptors contribute to house dust mite-induced eosinophilic airway inflammation via T_H2 cytokine production

( Donghwan Park ),( Dong-wook Kwak ),( Jae-hong Kim ) 생화학분자생물학회 2021 BMB Reports Vol.54 No.3

Leukotriene B₄ (LTB₄) is a lipid mediator of inflammation that is generated from arachidonic acid via the 5-lipoxygenase pathway. Previous studies have reported that the receptors of LTB₄, BLT1, and BLT2 play mediatory roles in the allergic airway inflammation induced by ovalbumin (OVA). However, considering that house dust mites (HDMs) are the most prevalent allergen and well-known risk factor for asthmatic allergies, we are interested in elucidating the contributory roles of BLT1/2 in HDM-induced allergic airway inflammation. Our aim in this study was to investigate whether BLT1/2 play any roles in HDM-induced allergic airway inflammation. In this study, we observed that the levels of ligands for BLT1/2 [LTB₄ and 12(S)-HETE (12(S)-hydroxyeicosatetraenoic acid)] were significantly increased in bronchoalveolar lavage fluid (BALF) after HDM challenge. Blockade of BLT1 or BLT2 as well as of 5-lipoxygenase (5-LO) or 12-lipoxygenase (12-LO) markedly suppressed the production of T_H2 cytokines (IL-4, IL-5, and IL-13) and alleviated lung inflammation and mucus secretion in an HDM-induced eosinophilic airway-inflammation mouse model. Together, these results indicate that the 5-/12-LO-BLT1/2 cascade plays a role in HDM-induced airway inflammation by mediating the production of T_H2 cytokines. Our findings suggest that BLT1/2 may be a potential therapeutic target for patients with HDM-induced allergic asthma. [BMB Reports 2021; 54(3): 182-187]

The barley lipid transfer protein, BLT101, enhances cold tolerance in wheat under cold stress

최창현,황철호 한국식물생명공학회 2015 Plant biotechnology reports Vol.9 No.4

To understand the mechanism by which the barley plants tolerate overwintering, we analyzed the BLT101 gene expression in four different cultivars including two each of spring and winter barleys, where the BLT101 transcript level varied among the cultivars with a long-term cold acclimation. The BLT101 transcript was induced at a higher level in the crown tissues in response to low temperature treatment. The BLT101 transcript level correlated with those of the corresponding protein. BLT101- overexpressing transgenic wheat lines (BLT101ox’s) grew slowly and showed decreased expression of the genes induced by hormones (such as auxin and cytokinin), compared to non-transgenic (NT) plants. After a prolonged cold treatment, the BLT101ox leaves showed normal phenotypes, while the NT plants displayed blighted and withered leaves. A substances leakage test showed that the BLT101ox wheat retains enhanced freezing tolerance, compared to the NT plants. The transgenic lines lost less water than the NT plants, under a cold-stressed condition. Our findings suggest that tolerance to water stress and the retardation of growth in cold conditions, the essential features in overwintering barleys, involve BLT101.

Contributory Role of BLT2 in the Production of Proinflammatory Cytokines in Cecal Ligation and Puncture-Induced Sepsis

박동환,노명자,이아진,곽동욱,정윤노,김재홍 한국분자세포생물학회 2021 Molecules and cells Vol.44 No.12

BLT2 is a low-affinity receptor for leukotriene B4, a potent lipid mediator of inflammation generated from arachidonic acid via the 5-lipoxygenase pathway. The aim of this study was to investigate whether BLT2 plays any role in sepsis, a systemic inflammatory response syndrome caused by infection. A murine model of cecal ligation and puncture (CLP)-induced sepsis was used to evaluate the role of BLT2 in septic inflammation. In the present study, we observed that the levels of ligands for BLT2 (LTB4 [leukotriene B4] and 12(S)-HETE [12(S)-hydroxyeicosatetraenoic acid]) were significantly increased in the peritoneal lavage fluid and serum from mice with CLP-induced sepsis. We also observed that the levels of BLT2 as well as 5-LO and 12-LO, which are synthesizing enzymes for LTB4 and 12(S)-HETE, were significantly increased in lung and liver tissues in the CLP mouse model. Blockade of BLT2 markedly suppressed the production of sepsis-associated cytokines (IL-6 [interleukin-6], TNF-α [tumor necrosis factor alpha], and IL-1β [interleukin-1β] as well as IL-17 [interleukin-17]) and alleviated lung inflammation in the CLP group. Taken together, our results suggest that BLT2 cascade contributes to lung inflammation in CLP-induced sepsis by mediating the production of inflammatory cytokines. These findings suggest that BLT2 may be a potential therapeutic target for sepsis patients.

The 5-/12-lipoxygenase-BLT2 cascade induces elevated expression of c-Myc, thus mediating the proliferation and migration of KRAS mutant colorectal cancer cells

박재인,장재현,Jun-Dong Wei,김재홍 한국통합생물학회 2023 Animal cells and systems Vol.27 No.1

Colorectal cancer (CRC) is the most common malignant tumor and one of the leading causes ofcancer-related death worldwide. Oncogene KRAS is a commonly mutated in CRC and playscrucial roles in the colorectal tumorigenesis. Emerging evidence suggests that increasedexpression levels of c-Myc are critical for KRAS-mediated CRC progression. However, it is unclearexactly how c-Myc expression is regulated in CRC. In the present study, we found that blockadeof 5/12-lipoxygenase (5/12-LO) or leukotriene B4 receptor (BLT2) markedly reduced c-Mycexpression in KRAS-mutant LOVO cells, while the control COLO 320DM cells were not affected. When the 5/12-LO products LTB4 and 12(S)-HETE, ligands of BLT2, were added to LOVO cells,c-Myc expression levels were restored, together suggesting that ‘5/12-LO-BLT2 cascade’regulates c-Myc expression in KRAS-mutant CRC cells. We also observed that ‘5/12-LO-BLT2’-mediated c-Myc upregulation contributes to cell proliferation by regulating the expression ofcyclin D1 in LOVO cells. Moreover, ‘5/12-LO-BLT2-cMyc’ cascade regulates the expression ofEMT-related proteins in KRAS-mutant CRC cells, and depletion of both c-Myc and BLT2 usingsiRNA significantly enhanced the level of E-cadherin and reduced the level of Vimentin in LOVOcells. Taken together, our findings suggest that the BLT2-linked cascade promotes KRAS-mutantCRC cell proliferation and migration through the elevated expression of c-Myc. Thus, our resultssuggest that BLT2 appears to be an effective therapeutic target for KRAS-mutant CRCs.

A leukotriene B ₄ receptor-2 is associated with paclitaxel resistance in MCF-7/DOX breast cancer cells

Kim, H,Park, G-S,Lee, J E,Kim, J-H Nature Publishing Group 2013 The British journal of cancer Vol.109 No.2

<P><B>Background:</B></P><P>Breast cancer is the most common malignancy in women. Although chemotherapeutic agents, such as paclitaxel, are effective treatments for the majority of breast cancer patients, recurrence is frequent and often leads to death. Thus, there is an urgent need to identify novel therapeutic targets that sensitise tumour cells to existing chemotherapy agents.</P><P><B>Methods:</B></P><P>The levels of leukotriene B<SUB>4</SUB> receptor-2 (BLT2) in multidrug-resistant MCF-7/DOX cells were determined using quantitative PCR and FACS analysis. The potential role of BLT2 in the paclitaxel resistance of MCF-7/DOX cells was assessed using a pharmacological inhibitor and small interfering RNA knockdown, and the BLT2-associated resistance mechanism was assessed.</P><P><B>Results:</B></P><P>The expression levels of BLT2 were markedly upregulated in MCF-7/DOX cells. The inhibition of BLT2 by pre-treatment with LY255283 or siBLT2 knockdown significantly sensitised MCF-7/DOX cells to paclitaxel and induced significant levels of apoptotic death, suggesting that BLT2 mediates paclitaxel resistance. We also demonstrated that BLT2-induced paclitaxel resistance was associated with the upregulation of P-glycoprotein. Finally, co-treatment with a BLT2 inhibitor and paclitaxel markedly reduced tumour growth in an MCF-7/DOX <I>in vivo</I> model.</P><P><B>Conclusion:</B></P><P>Together, our results demonstrate that BLT2 is a novel therapeutic target that sensitises drug-resistant breast cancer cells to paclitaxel.</P>

RanBPM inhibits BLT2-mediated IL-8 production and invasiveness in aggressive breast cancer cells

Wei, Jun-Dong,Jang, Jae-Hyun,Kim, Jae-Hong Academic Press 2017 Biochemical and biophysical research communication Vol. No.

<P><B>Abstract</B></P> <P>RanBPM is a scaffolding protein that regulates several cellular processes by interacting with various proteins. Previously, we reported that RanBPM acts as a negative regulator of BLT2, a low-affinity leukotriene B<SUB>4</SUB> receptor; thus, it interferes with BLT2-mediated cell motility. In the present study, we observed that the expression levels of RanBPM were markedly reduced in the highly aggressive MDA-MB-435 and MDA-MB-231 human breast cancer cell lines compared with those in non-invasive MCF-7 cells. Additionally, we found that the restoration of RanBPM levels suppressed the invasiveness of these aggressive breast cancer cells in a manner dependent on BLT2 activation. In contrast, the knockdown of endogenous RanBPM by shRNA strongly promoted invasiveness in non-invasive MCF-7 cells. We also observed that RanBPM suppressed the invasiveness of aggressive breast cancer cells by inhibiting BLT2-mediated reactive oxygen species (ROS) generation and IL-8 production. Taken together, our results suggest that RanBPM acts as a negative regulator of BLT2, thus attenuating the invasiveness of aggressive breast cancer cells.</P> <P><B>Highlights</B></P> <P> <UL> <LI> RanBPM suppresses the invasiveness of aggressive breast cancer cells by inhibiting BLT2-mediated ROS generation and IL-8 production. </LI> <LI> The expression levels of RanBPM are reduced in the aggressive breast cancer cells. </LI> <LI> Restoration of RanBPM levels suppressed the invasiveness of aggressive breast cancer cells in a manner dependent on the LTB4-BLT2 axis. </LI> </UL> </P>

Leukotriene B4 Regulates Proliferation and Differentiation of Cultured Rat Myoblasts via the BLT1 Pathway

Ru Sun,Xueqing Ba,Lingling Cui,Yan Xue,Xianlu Zeng 한국분자세포생물학회 2009 Molecules and cells Vol.27 No.4

Skeletal muscle regeneration is a highly orchestrated process initiated by activation of adult muscle satellite cells. Upon muscle injury, the inflammatory process is always accompanied by muscle regeneration. Leukotriene B4 is one of the essential inflammatory mediators. We isolated and cultured primary satellite cells. RT-PCR showed that myoblasts expressed mRNA for LTB4 receptors BLT1 and BLT2, and LTB4 promoted myoblast proliferation and fusion. Quantitative real-time PCR and immunoblotting showed that LTB4 treatment expedited the expression process of differentiation markers MyoD and M-cadherin. U-75302, a specific BLT1 inhibitor, but not LY2552833, a specific BLT2 inhibitor, blocked proliferation and differentiation of myoblasts induced by LTB4, which implies the involvement of the BLT1 pathway. Overall, the data suggest that LTB4 contributes to muscle regeneration by accelerating proliferation and differentiation of satellite cells.

BLT2, a leukotriene B4 receptor 2, as a novel prognostic biomarker of triple-negative breast cancer

( Jaein Park ),( Jae-hyun Jang ),( Geun-soo Park ),( Yunro Chung ),( Hye Jin You ),( Jae-hong Kim ) 생화학분자생물학회 2018 BMB Reports Vol.51 No.8

Triple-negative breast cancer (TNBC) is considered to be a notorious type of cancer due to its aggressive metastatic potential and poor prognosis. Recent evidence suggests that BLT2, a low-affinity LTB₄ receptor is critically associated with the phenotypes of TNBC cells, including invasion, metastasis, and survival. Furthermore, in a group of 545 breast cancer patients with metastasis, we observed that the high-BLT2 subgroup had a lower disease-free-survival rate than the low-BLT2 subgroup. Thus, we theorized that anti-BLT2 strategies could facilitate the development of new therapies used for TNBC. This review focuses on recent discoveries regarding BLT2 and its roles in as a novel prognostic biomarker in TNBC. [BMB Reports 2018; 51(8): 373-377]