• BMB Reports
• /
• 제36권3호
• /
• pp.288-293
• /
• 2003

Low molecular weight phosphotyrosine protein phosphatase (LMW-PTP) has been implicated in modulating the EphB1-mediated signaling pathway. In this study, we demonstrated that the EphA8 receptor phosphorylates LMW-PTP in vitro. In addition, we discovered that mixing these two proteins leads to EphA8 dephosphorylation in the absence of phosphatase inhibitors. Finally, we demonstrated that LMW-PTP, modified by the EphA8 autokinase activity, possesses enhanced catalytic activity in vitro. These results suggest that LMW-PTP may also participate in a feedback-control mechanism of the EphA8 receptor autokinase activity in vivo.

• Molecules and Cells
• /
• 제38권4호
• /
• pp.349-355
• /
• 2015

EphA7 has been implicated in the regulation of apoptotic cell death in neural epithelial cells. In this report, we provide evidence that EphA7 interacts with caspase-8 to induce apoptotic cell signaling. First, a pull-down assay using biotinylated ephrinA5-Fc showed that EphA7 co-precipitated with wild type caspase-8 or catalytically inactive caspase-8 mutant. Second, co-transfection of EphA7 with caspase-8 significantly increased the number of cleaved caspase-3 positive apoptotic cells under an experimental condition where transfection of EphA7 or caspase-8 alone did not affect cell viability or apoptosis. EphA4 also had a causative role in inducing apoptotic cell death with caspase-8, whereas EphA8 did not. Third, caspase-8 catalytic activity was essential for the apoptotic signaling cascade, whereas tyrosine kinase activity of the EphA4 receptor was not. Interestingly, we found that kinase-inactive EphA4 was well co-localized at the plasma membrane with catalytically inactive caspase-8, suggesting that an interaction between these mutant proteins was more stable. Finally, we observed that the extracellular region of the EphA7 receptor was critical for interacting with caspase-8, whereas the cytoplasmic region of EphA7 was not. Therefore, we propose that Eph receptors physically associate with a transmembrane protein to form an apoptotic signaling complex and that this unidentified receptor-like protein acts as a biochemical linker between the Eph receptor and caspase-8.

• Genomics & Informatics
• /
• 제21권3호
• /
• pp.30.1-30.13
• /
• 2023

Ephs belong to the largest family of receptor tyrosine kinase and are highly conserved both sequentially and structurally. The structural organization of Eph is similar to other receptor tyrosine kinases; constituting the extracellular ligand binding domain, a fibronectin domain followed by intracellular juxtamembrane kinase, and SAM domain. Eph binds to respective ephrin ligand, through the ligand binding domain and forms a tetrameric complex to activate the kinase domain. Eph-ephrin regulates many downstream pathways that lead to physiological events such as cell migration, proliferation, and growth. Therefore, considering the importance of Eph-ephrin class of protein in tumorigenesis, 7,620 clinically reported missense mutations belonging to the class of variables of unknown significance were retrieved from cBioPortal and evaluated for pathogenicity. Thirty-two mutations predicted to be pathogenic using SIFT, Polyphen-2, PROVEAN, SNPs&GO, PMut, iSTABLE, and PremPS in-silico tools were found located either in critical functional regions or encompassing interactions at the binding interface of Eph-ephrin. However, seven were reported in nonsmall cell lung cancer (NSCLC). Considering the relevance of receptor tyrosine kinases and Eph in NSCLC, these seven mutations were assessed for change in the folding pattern using molecular dynamic simulation. Structural alterations, stability, flexibility, compactness, and solvent-exposed area was observed in EphA3 Trp790Cys, EphA7 Leu749Phe, EphB1 Gly685Cys, EphB4 Val748Ala, and Ephrin A2 Trp112Cys. Hence, it can be concluded that the evaluated mutations have potential to alter the folding pattern and thus can be further validated by in-vitro, structural and in-vivo studies for clinical management.

• Animal cells and systems
• /
• 제8권1호
• /
• pp.57-63
• /
• 2004

This study provides evidence that expression of EphA8 receptor in NG108-15 cells results in a substantial increase in the number of neurite-bearing cells. However, the EphA8-induced neurite outgrowth does not require either ephrin-A5 stimulation or ectopic expression of $p110{\gamma}$ PI-3 kinase. In contrast, co-expression of a lipid kinase-inactive $p110{\gamma}$ mutant together with EphA8 causes neurite retraction in the presence of ephrin-A5 stimulation. This effect was not observed in the absence of ephrin-A5 stimulation. Significantly, the tyrosine kinase activity of EphA8 is not important for either of these processes. Taken together, our results strongly suggest that $p110{\gamma}$ PI-3 kinase is critically involved in the regulatory process by which ephrin-A5 exerts effects on the EphA8-induced neurite outgrowth.

• Journal of Gastric Cancer
• /
• 제6권1호
• /
• pp.25-30
• /
• 2006

목적: EphB2 수용체는 티로신 키나제 수용체 군에 속하면서 Wnt 신호 전달계의 대상 유전자로 세포의 성장 및 이동을 조절함으로써 종양의 발생과 진행을 억제한다. 연구자들은 한국인 위암에서 EphB2 단백의 발현 변화가 위암의 발생 및 진행에 관여하는지와 위암의 병리 지표들과의 연관성을 알아보고자 하였다. 대상 및 방법: 83예의 파라핀 포매된 위암조직에서 암세포들을 각각 3군데에서 펀치하여 새로운 파라핀 블록으로 옳겨 위암의 고집적 tissue microarray를 제작하였다. Tissue microarray 절편에서 EphB2 단백에 대한 항체로 면역조직화 학염색을 실시한 후 그 발현 양상을 병리 지표들인 조직학적 분류, 침습 정도, 림프절 전이 및 복막 파종과의 연관성을 조사하였다. 결과: EphB2 단백은 정상 위 점막 상피세포의 세포막과 세포질에서 발현되고 있었고 조사된 83예의 위암 중 30예 (36.1%)에서 발현이 소실되었다. 흥미롭게도 EphB2 단백의 발현 소실은 위암의 림프절 전이와 통계적으로 연관성이 있었으나 조직학적 분류, 침습 정도 및 복막 파종과는 연관성이 없었다. 결론: 이러한 소견들은 EphB2 수용체 단백의 발현 소실이 위암의 발생 및 진행에 있어 중요한 과정 중의 하나임을 의미한다.

• Genomics & Informatics
• /
• 제6권4호
• /
• pp.192-201
• /
• 2008

Erythropoietin-producing human hepatocellular carcinoma receptor B1 (EPHB1) is a member of the Eph family of receptor tyrosine kinases that mediate vascular system development. Eph receptor overexpression has been observed in various cancers and is related to the malignant transformation, metastasis, and differentiation of cancers, including hepatocellular carcinoma (HCC). Eph receptors regulate cell migration and attachment to the extracellular matrix by modulating integrin activity. EphrinB1, the ligand of EPHB1, has been shown to regulate HCC carcinogenesis. Here, we sought to determine whether EPHB1 polymorphisms are associated with hepatitis B virus (HBV)-infected liver diseases, including chronic liver disease (CLD) and HCC. We genotyped 26 EPHB1 single nucleotide polymorphisms (SNPs) in 399 Korean CLD, HCC, and LD (CLD+HCC) cases and seroconverted controls (HBV clearance, CLE) using the GoldenGate assay. Two SNPs (rs6793828 and rs11717042) and 1 haplotype that were composed of these SNPs were associated with an increased risk for CLD, HCC, and LD (CLD+HCC) compared with CLE. Haplotypes that could be associated with HBV-infected liver diseases by affecting downstream signaling were located in the Eph tyrosine kinase domain of EPHB1. Therefore, we suggest that EPHB1 SNPs, haplotypes, and diplotypes may be genetic markers for the progression of HBV-associated acute hepatitis to CLD and HCC.

• BMB Reports
• /
• 제41권6호
• /
• pp.479-484
• /
• 2008

In the present study, we demonstrate that ephrin-A5 is able to induce a transient increase of MAP kinase activity in PC12 cells. However, the effects of ephrin-A5 on the MAP kinase signaling pathway are about three-fold less than that of EGF. In addition, we demonstrate that EphA4 is the only Eph member expressed in PC12 cells, and that tyrosine phosphorylation induced by ephrin-A5 treatment is consistent with the magnitude and longevity of MAP kinase activation. Experiments using the Ras dominant negative mutant N17Ras reveal that Ras plays a pivotal role in ephrin-A5-induced MAP kinase activation in PC12 cells. Importantly, we found that the EphA4 receptor is rapidly internalized by endocytosis upon engagement of ephrin-A5, leading to a subsequent reduction in the MAP kinase activation. Together, these data suggest a novel regulatory mechanism of differential Ras-MAP kinase signaling kineticsexhibited by the forward signaling of EphA4 in PC12 cells.

• Tuberculosis and Respiratory Diseases
• /
• 제78권3호
• /
• pp.218-226
• /
• 2015

Background: Eph receptors and ephrin ligands have several functions including angiogenesis, cell migration, axon guidance, fluid homeostasis, oncogenesis, inflammation and injury repair. The EphA2 receptor potentially mediates the regulation of vascular permeability and inflammation in response to lung injury. Methods: Mice were divided into 3 experimental groups to study the role of EphA2 signaling in the lipopolysaccharide (LPS)-induced lung injury model i.e., IgG+phosphate-buffered saline (PBS) group (IgG instillation before PBS exposure), IgG+LPS group (IgG instillation before LPS exposure) and EphA2 monoclonal antibody (mAb)+LPS group (EphA2 mAb pretreatment before LPS exposure). Results: EphA2 and ephrinA1 were upregulated in LPS-induced lung injury. The lung injury score of the EphA2 mAb+LPS group was lower than that of the IgG+LPS group ($4.30{\pm}2.93$ vs. $11.45{\pm}1.20$, respectively; p=0.004). Cell counts (EphA2 mAb+LPS: $11.33{\times}10^4{\pm}8.84{\times}10^4$ vs. IgG+LPS: $208.0{\times}10^4{\pm}122.6{\times}10^4$; p=0.018) and total protein concentrations (EphA2 mAb+LPS: $0.52{\pm}0.41mg/mL$ vs. IgG+LPS: $1.38{\pm}1.08mg/mL$; p=0.192) were decreased in EphA2 mAb+LPS group, as compared to the IgG+LPS group. In addition, EphA2 antagonism reduced the expression of phospho-p85, phosphoinositide 3-kinase $110{\gamma}$, phospho-Akt, nuclear factor ${\kappa}B$, and proinflammatory cytokines. Conclusion: This results of the study indicated a role for EphA2-ephrinA1 signaling in the pathogenesis of LPS-induced lung injury. Furthermore, EphA2 antagonism inhibits the phosphoinositide 3-kinase-Akt pathway and attenuates inflammation.

• Animal cells and systems
• /
• 제13권1호
• /
• pp.9-15
• /
• 2009

Eph receptors and their ephrin ligands have been implicated in a variety of cellular processes such as cellular morphogenesis and motility. Our previous studies demonstrated that Odin, one of the Anks family proteins, functions as a scaffolding protein of the EphA8 signaling pathway leading to modulation of cell migration or axonal outgrowth. Here we show that WDR7 is associated with Odin and that it is possibly implicated in the EphA8 signaling pathway. WD40 repeats present in the COOH-terminal region of WDR7 appear to be crucial for its association with Odin, whereas the binding motif of Odin is located in between ankyrin repeats and PTB domain. Co-immunoprecipitation experiments revealed that association of WDR7 with Odin is enhanced by ephrin ligand treatment, possibly through forming large protein complexes including both EphA8 and ephrin-A5. Consistently, immunofluorescence staining experiments suggested that WDR7 constitute a component of the large protein complexes containing Odin, EphA8 and ephrin-A5. Taken together, our results suggest the WDR7-Odin complexes might be involved in the signaling pathway downstream of the EphA8 receptor.

• BMB Reports
• /
• 제49권12호
• /
• pp.651-652
• /
• 2016

EphA2 has been implicated in amplifying ErbB2 tumorigenic signaling. One protein that interacts with EphA2 is the Anks1a PTB adaptor. However, the precise role of Anks1a in EphA2-mediated tumorigenesis is unclear. We demonstrated that Anks1a localizes to the ER upon phosphorylation and that the Ankyrin repeats and PTB of Anks1a bind to EphA2 and Sec23, respectively. Thus, Anks1a facilitates the selective packaging of EphA2 into COPII vesicles. Additionally, Anks1a knockout mice, a phenocopy of EphA2 knockout mice, exhibited markedly reduced ErbB2-induced breast tumorigenesis. Strikingly, ErbB2 did not localize to the cell surface following Anks1a knockdown in primary mammary tumor cells over-expressing ErbB2. Importantly, EphA2 was critical for stabilizing ErbB2 through complex formation, but its interaction with Anks1a also facilitated ErbB2 loading into COPII carriers. These findings suggest a novel role for Anks1a in the molecular pathogenesis of breast tumors and possibly other human diseases.