• Biomolecules & Therapeutics
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• 제25권1호
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• pp.26-43
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• 2017

Endocytosis is a process by which cells absorb extracellular materials via the inward budding of vesicles formed from the plasma membrane. Receptor-mediated endocytosis is a highly selective process where receptors with specific binding sites for extracellular molecules internalize via vesicles. G protein-coupled receptors (GPCRs) are the largest single family of plasma-membrane receptors with more than 1000 family members. But the molecular mechanisms involved in the regulation of GPCRs are believed to be highly conserved. For example, receptor phosphorylation in collaboration with ${\beta}$-arrestins plays major roles in desensitization and endocytosis of most GPCRs. Nevertheless, a number of subsequent studies showed that GPCR regulation, such as that by endocytosis, occurs through various pathways with a multitude of cellular components and processes. This review focused on i) functional interactions between homologous and heterologous pathways, ii) methodologies applied for determining receptor endocytosis, iii) experimental tools to determine specific endocytic routes, iv) roles of small guanosine triphosphate-binding proteins in GPCR endocytosis, and v) role of post-translational modification of the receptors in endocytosis.

• Journal of Pharmaceutical Investigation
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• 제33권4호
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• pp.281-286
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• 2003

To design gene deliver systems which can deliver higher amounts of genes into stem cells, we studied the expression of receptors involved in the receptor-mediated endocytosis of bone marrow stromal stem cells. Bone marrow was isolated from ICR mice, and bone marrow stromal stem cells were isolated based on their plastic adherence property. Several culture conditions were screened for effective and continuous culture of marrow stromal stem cells. MesenCult medium was finally used to cultivate marrow stromal stem cells in vitro. As candidate receptors, various chemokine receptors were studied. Both bone marrow cells ad marrow-derived stromal stem cells showed expression of CC chemokine receptors (CCR) and CXC chemokine receptors (CXCR). Marrow stromal stem cells showed higher expression of CCR5 ad CXCR4 chemokine receptors as compared to other types of chemokine receptors. Moreover, though the expression of chemokine receptors generally decreased in most chemokine receptors with the cultivaton of marrow stromal stem cells, CCR5 and CXCR4 chemokine receptors retained the higher level of receptor expressions over prolonged periods. These results suggest that the ligands exhibiting specific binding to CCR5 or CXCR4 might be used to modify gene delivery systems for increased levels of receptor-mediated gene delivery into stromal stem cells.

• 한국산학기술학회논문지
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• 제12권4호
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• pp.1770-1774
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• 2011

본 논문은 리포포린에 의해 디아실글리세리드(DAG)가 Manduca sexta 유충 지방체로 운반되는 과정을 조사하였다. $[^3H]$-DAG 표지 리포포린($[^3H]$-DAG-Lp)을 시간별로 유충 지방체와 배양하여 지방체로 운반되는 DAG의 방사능을 결정하였다. $[^3H]$-DAG-Lp와 지방체를 배양하면 지방체에 DAG가 축적되며 이의 일부는 지방체에서 트리아실글리세리드(TAG)로 전환되는 것을 알 수 있었다. Suramin과 표지되지 않은 리포포린(unlabeled Lp)의 존재 하에서는 지방체로 운반되는 DAG가 억제되는 데, 이는 DAG를 운반하는 데 리포포린 수용체가 관여한다는 사실을 알 수 있었다. Suramin의 효과는 다소 복잡하지만 리포포린 수용체에 결합하여 막의 특성을 변화시켜 DAG 운반속도에 영향을 주는 것 같다. 지질 운반과정이 수용체-매개 내포작용이라는 사실을 조사하기 위하여 내포작용 억제자인 ammonium chloride와 chloroquine을 처리하였다. 그 결과 리포포린과 지방체사이에서의 지질 운반 기작이 수용체-매개 과정이라는 사실을 보여준다.

• BMB Reports
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• 제36권3호
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• pp.265-268
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• 2003

Retinoic acid (RA) can transform the Golgi apparatus (GA) into a diffuse vacuolar aggregate and increase the toxicity of some immunotoxins that enter into cells by receptor-mediated endocytosis. An ultramorphological study of the RA-induced GA disruption was performed on F2000 fibroblasts. Cultures were treated with 0.11 to $30\;{\mu}M$ RA for 7 - 180 min. The endocytosis of Limax flavus agglutinin-peroxidase conjugate (LFA), and the interactions between a phorbol ester (PMA) and RA concerning GA disruption, were examined. Exposure to $0.33\;{\mu}M$ RA for 20 min transformed the GA into vacuolar aggregate. These vacuoles were not involved in endocytosis since they remained unstained after endocytosis of LFA. However, the lysosomes were involved in endocytosis, as they were strongly stained. Therefore, a RA-induced shift towards lysosomal routing of the entered LFA was presumed. Exposure to PMA made cells resistant to the Golgi-disturbing effects of RA, indicating that protein kinase C plays an important role in this process.

• 한국미생물학회:학술대회논문집
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• 한국미생물학회 2004년도 International Meeting of the Microbiological Society of Korea
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• pp.141-146
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• 2004

SPIN90 was identified to farm molecular complex with $\betaPIX$, WASP and Nck. This complex shows that SPIN90 interacts with Nck in a manner dependent upon cell adhesion to extracellular matrix, but $SPIN90{\cdot}{\beta}PIX{\cdot}WASP$ complex was stable even in suspended cells. This suggests that SPIN90 serves as an adaptor molecule to recruit other proteins to Nck at focal adhesions. SPIN90 was phosphorylated by ERK1, which was, itself, activated by cell adhesion and platelet-derived growth factor. Such phosphorylation of SPIN90 likely promotes the interaction of the $SPIN90{\cdot}{\beta}PIX{\cdot}WASP$ complex and Nck. It thus appears that the interaction of the $SPIN90{\cdot}{\beta}PIX{\cdot}WASP$ complex with Nck is crucial for stable cell adhesion and can be dynamically modulated by SPIN90 phosphorylation that is dependent on cell adhesion and ERX activation. SPIN90 directly binds syndapin I, syndapin isoform II-1 and II-s via its PRD region in vitro, in vivo and also associates with endocytosis core components such as clathrin and dynamin. In neuron and fibroblast, SPIN90 colocalizes with syndapins as puntate form, consistent with a role for SPIN90 in clathrin-mediated endocytosis pathway. Overexpression of SPIN90 N-term inhibits receptor-mediated endocytosis. Interestingly, SPIN90 PRD, binding interface of syndapin, significantly blocks internalization of transferrin, demonstrating SPIN90 involvement in endocytosis in vivo by interacting syndapin. Depletion of endogenous SPIN90 by introducing $\alpha-SPIN90$ also blocks receptor-mediated endocytosis. Actin polymerization could generate farce facilitating the pinch-out event in endocytosis, detach newly formed endocytic vesicle from the plasma membrane or push out them via the cytosol on actin tails. Here we found that SPIN90 localizes to high actin turn over cortical area, actin-membrane interface and membrane ruffle in PDGF treated cells. Overexpression of SPIN90 has an effect on cortical actin rearrangement as filopodia induction and it is mediated by the Arp2/3 complex at cell periphery. Consistent with a role in actin organization, CFP-SPIN90 present in actin comet tail generated by PIP5 $kinase\gamma$ overexpression. Therefore this study suggests that SPIN90 is functional linker between endocytosis and actin cytoskeleton.

• 한국동물학회지
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• 제39권3호
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• pp.292-298
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• 1996

안점의 꽃갯지렁이(Pseudopotamilla occelata)의 난자형성중 난황단백질의 전구체인 체강액 단백질(CP)은 수용체에 의해 중개되는데 이러한 수용체 중개에 의한 난모세포내로의 유입은 GTP-binding protein에 의해 조절되는 것으로 확인되었다. 체강액 단백질(CP)을 가장 활발히 투과시키는 중기 난모세포내로의 125 I-CP의 유입은 GTP에 의해 촉지되었고, GTP 유사체인 GTPrS나 GTP$\beta$S의 효과를 금입자로 표지된 체강액 단백질을 이용하여 전자현미경으로 확인해 본 결과, gold-labeled CP는 난황립에 집중되었고, 이러한 현상도 GTP에 의해서는 촉진되었고 GTPrS에 의해서는 억제되었다.

• Biomolecules & Therapeutics
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• 제32권1호
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• pp.56-64
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• 2024

Biased signaling or functional selectivity refers to the ability of an agonist or receptor to selectively activate a subset of transducers such as G protein and arrestin in the case of G protein-coupled receptors (GPCRs). Although signaling through arrestin has been reported from various GPCRs, only a few studies have examined side-by-side how it differs from signaling via G protein. In this study, two signaling pathways were compared using dopamine D₂ receptor (D₂R) mutants engineered via the evolutionary tracer method to selectively transduce signals through G protein or arrestin (D₂G and D₂Arr, respectively). D₂G mediated the inhibition of cAMP production and ERK activation in the cytoplasm. D₂Arr, in contrast, mediated receptor endocytosis accompanied by arrestin ubiquitination and ERK activation in the nucleus as well as in the cytoplasm. D₂Arr-mediated ERK activation occurred in a manner dependent on arrestin3 but not arrestin2, accompanied by the nuclear translocation of arrestin3 via importin1. D₂R-mediated ERK activation, which occurred in both the cytosol and nucleus, was limited to the cytosol when cellular arrestin3 was depleted. This finding supports the results obtained with D₂Arr and D₂G. Taken together, these observations indicate that biased signal transduction pathways activate distinct downstream mechanisms and that the subcellular regions in which they occur could be different when the same effectors are involved. These findings broaden our understanding on the relation between biased receptors and the corresponding downstream signaling, which is critical for elucidating the functional roles of biased pathways.

• BMB Reports
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• 제44권2호
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• pp.96-101
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• 2011

Calcineurin is a serine/threonine protein phosphatase controlled by $Ca^{2+}$ and calmodulin that has been implicated in various signaling pathways. Previously, we reported that calcineurin regulates coelomocyte endocytosis in Caenorhabditis elegans. So far, simple and powerful in vivo approaches have been developed to study various endocytic processes in C. elegans. Using these in vivo assays, we further analyzed the endocytic phenotypes of calcineurin mutants. We observed that the calcineurin mutants were defective in apical endocytosis in the intestine as well as synaptic vesicle recycling in the nerve cord. However, we found that calcineurin mutants displayed normal receptor-mediated endocytosis in oocytes. Therefore, our results suggest that calcineurin may regulate specific sets of endocytic processes in nematode.

• IMMUNE NETWORK
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• 제1권3호
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• pp.196-202
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• 2001

Backgorund: WEHI-231 B cell line is a representative model for $IgM^+$ mature B cells. To understand the signaling differences between mature and immature B cells, we compared the responsiveness of WEHI-231 and Bal 17 B cell lines to BCR cross-linking. Methods: The extents of tyrosine phosphorylation, ligand-induced internalization, and activation-induced cell death upon BCR cross-linking were compared in two cell lines. Results: Despite a higher expression of BCR, cross-linking of BCR on WEHI-231 cell evoked a weaker level of tyrosine phosphorylation and BCR endocytosis than Bal 17 cells. Furthermore, the endocytosed BCR could not enter the lysosomal compartment and stayed as peripheral spots in WEHI-231 cells. Conclusion: WEHI-231 cell showed preferred BCR-mediated signaling pathways leading to a reduced capability of antigen presentation as well as the enhanced apoptosis in comparision with Bal 17 cells. These results might reflect the signaling differences between mature and immature B cells.

• 생명과학회지
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• 제25권10호
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• pp.1103-1109
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• 2015

뇌 해마의 콜린성 신경분포는 학습과 기역에 연관성이 있는 것으로 알려져 있으며 이의 작용제인 carbachol 투여 시 장기기억 저하가 유도됨이 알려져 왔다. 그러나 이러한 콜린성 자극에 의한 해마 신경세포의 시냅스 내 변화기작은 완전히 알려지지 않고 있다. 본 연구에서는 아세틸콜린 수용체의 활성에 의하여 유도되는 장기기억 저하 현상에 있어 alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) 수용체가 후시냅스 표면으로부터 사라지는 현상과 이의 조절기작에 대하여 알아보고자 한다. 이를 위하여 쥐 해마의 일차세포를 추출하고 체외에서 배양한 성숙 신경세포에 carbachol 을 투여하여 장기기억 저하를 유도 하였으며, 후시냅스의 표면으로부 터 AMPA 수용체의 아단위체인 GluA2가 M1 무스카린 수용체의 길항제에 의하여 저해 되었다. 또한 콜린성 자극 에 의한 GluA2의 내재화 현상의 작용기작 연구를 위하여 쥐 해마 절편에 carbachol 투여 후 GluA2와 직접적인 상호작용을 하는 Glutam내재화 되었음을 확인하였다. 이러한 현상은 ate receptor-interacting protein 1 (GRIP1) 과 clathrine 단백질이 매개하는 세포내이입 작용을 하는 adaptin-α 단백질의 결합 변화를 관찰하였다. GluA2는 carbachol 자극에 의해 세포내이입 과정에서 adaptin-α 와의 결합이 증가하였으며 반대로 GRIP1과는 해리되었다. 이는 아세틸콜린의 수용체의 자극에 의하여 GluA2의 내제화 작용이 수반되며, 이의 작용기작으로 GluA2의 후시 냅스 표면 발현시에 결합하고 있는 GRIP1과 해리 되면서 장기기억 저하 현상이 유도됨을 의미한다.