• BMB Reports
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• 제51권10호
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• pp.484-485
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• 2018

Receptor-interacting protein kinase-3 (RIP3 or RIPK3) is a serine-threonine kinase largely essential for necroptotic cell death; it also plays a role in some inflammatory diseases. High levels of RIP3 are likely sufficient to activate necroptotic and inflammatory pathways downstream of RIP3 in the absence of an upstream stimulus. For example, we have previously detected high levels or RIP3 in the skin of Toxic Epidermal Necrolysis patients; this correlates with increased phosphorylation of MLKL found in these patients. We have long surmised that there are molecular mechanisms to prevent anomalous activity of the RIP3 protein, and so prevent undesirable cell death and inflammatory effects when inappropriately activated. Recent discovery that Carboxyl terminus of Hsp 70-Interacting Protein (CHIP) could mediate ubiquitylation- and lysosome-dependent RIP3 degradation provides a potential protein that has this capacity. However, while screening for RIP3-binding proteins, we discovered that pellino E3 ubiquitin protein ligase 1 (PELI1) also interacts directly with RIP3 protein; further investigation in this study revealed that PELI1 also targets RIP3 for proteasome-dependent degradation. Interestingly, unlike CHIP, which targets RIP3 more generally, PELI1 preferentially targets kinase active RIP3 that has been phosphorylated on T182, subsequently leading to RIP3 degradation.

• BMB Reports
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• 제39권6호
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• pp.774-781
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• 2006

Human nucleolar phosphoprotein p140 (hNopp140) is a nucleolar phosphoprotein that can bind to doxorubicin, an anti-cancer agent. We have examined the interaction between hNopp140 and doxorubicin as well as the folding property of hNopp140. Also, the effects of ATP and phosphorylation on the affinity of hNopp140 to doxorubicin are investigated by affinity dependent co-precipitation and surface plasmon resonance methods. Doxorubicin preferentially binds to un-phosphorylated form of hNopp140 with a $K_D$ value of $3.3\;{\times}\;10^{-7}$ M. Furthermore, doxorubicin reduces the protein kinase CK2-dependent phosphorylation of hNopp140, indicating that doxorubicin may perturb the cellular function of hNopp140 by reducing the protein kinase CK2-dependent phosphorylation of hNopp140. Low contents of the secondary structures of hNopp140 and the fast rate of proteolysis imply that hNopp140 has a high percentage of flexible regions or extended loop structures.

• 한국생물물리학회:학술대회논문집
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• 한국생물물리학회 2001년도 학술 발표회 진행표 및 논문초록
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• pp.24-24
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• 2001

The present study was undertaken to determine whether p38 mitogen-activated protein kinase participates in the regulation of vascular smooth muscle contraction by endothelin-I (ET-1) in rat thoracic aorta. ET-1 induced a sustained contraction. In contrast, both the intracellular Ca$\^$2＋/ and myosin light chain (MLC) phosphorylations were not sustained.(omitted)

• 생명과학회지
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• 제32권11호
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• pp.882-889
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• 2022

Breast cancer anti-estrogen resistance 3 (BCAR3)는 유방암에서 항에스트로겐 내성을 유도하는 유전자들 중의 하나로 발견되었다. 우리는 이미 BCAR3가 c-jun, activator protein-1, serum response element의 promoter 등을 활성화하는 것을 보고하였다. 본 연구에서 우리는 정상 유방세포인 MCF-12A에서 estrogen response element (ERE) 활성에서의 BCAR3의 기능을 조사하였다. BCAR3의 발현이 ERE를 활성화하는 것을 발견하였다. 이 ERE 활성화는 17β-estradiol에 의해 더욱 증가하였고, 이는 항에스트론겐인 tamoxifen에 의해 억제되지 않았다. 다음으로 우리는 ERE 활성화를 이끄는 BCAR3의 신호전달 경로를 연구하였다. BCAR3에 의한 ERE 활성화는 phosphatidylinositol (PI) 3-kinase 경로 억제제인 LY294002와 AZD5363에 의해서는 억제되었으나, Mitogen-activated protein kinase 경로 억제제인 PD98059와 U0126에 의해서는 억제되지 않았다. ERE 활성화는 PI3-kinase의 catalytic subunit p110α와 Akt의 active mutant에 의해서는 유도되었고, 이 활성화는 추가적인 BCAR3에 의해서는 더욱 증가하지 않았다. 이러한 결과로부터 우리는 BCAR3가 PI3-kinase/Akt 신호전달경로를 통하여 ERE 활성화에 중요한 역할을 하는 것을 제시한다.

• Journal of Ginseng Research
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• 제39권3호
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• pp.279-285
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• 2015

Background: Korean Red Ginseng has been used as a traditional oriental medicine to treat illness and to promote health for several thousand years in Eastern Asia. It is widely accepted that ginseng saponins, ginsenosides, are the major active ingredients responsible for Korean Red Ginseng's therapeutic activity against many kinds of illness. Although the crude saponin fraction (CSF) displayed antiplatelet activity, the molecular mechanism of its action remains to be elucidated. Methods: The platelet aggregation was induced by collagen, the ligand of integrin ${\alpha}_{II}{\beta}_I$ and glycoprotein VI. The crude saponin's effects on granule secretion [e.g., calcium ion mobilization and adenosine triphosphate (ATP) release] were determined. The activation of mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated protein kinase 1/2 (ERK1/2), c-Jun N-terminal kinases (JNKs), and p38 MAPK, and phosphoinositide 3-kinase (PI3K)/Akt was analyzed by immunoblotting. In addition, the activation of integrin ${\alpha}_{II}b{\beta}_{III}$ was examined by fluorocytometry. Results: CSF strongly inhibited collagen-induced platelet aggregation and ATP release in a concentration-dependent manner. It also markedly suppressed $[Ca^{2+}]_i$ mobilization in collagen-stimulated platelets. Immunoblotting assay revealed that CSF significantly suppressed ERK1/2, p38, JNK, PI3K, Akt, and mitogen-activated protein kinase kinase 1/2 phosphorylation. In addition, our fraction strongly inhibited the fibrinogen binding to integrin ${\alpha}_{IIb}{\beta}_3$. Conclusion: Our present data suggest that CSF may have a strong antiplatelet property and it can be considered as a candidate with therapeutic potential for the treatment of cardiovascular disorders involving abnormal platelet function.

• The Korean Journal of Physiology and Pharmacology
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• 제3권1호
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• pp.11-18
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• 1999

Nuclear factor $_{\kappa}B\;(NF-_{\kappa}B)$ activation is modulated by various protein kinases. Activation of $NF-_{\kappa}B$ is known to be important in the regulation of cell viability. The present study investigated the effect of inhibitors of protein tyrosine kinase (PTK), protein kinase C (PKC) and protein kinase A (PKA) on $NF-_{\kappa}B$ activity and the viability of bovine cerebral endothelial cells (BCECs). In serum-deprivation-induced BCEC death, low doses of $TNF{\alpha}$ showed a protective effect. $TNF{\alpha}$ induced $NF-_{\kappa}B$ activation within 4 h in serum-deprivation. PTK inhibitors (herbimycin A and genistein) and PKC inhibitor (calphostin C) prevented $NF-_{\kappa}B$ activation stimulated by $TNF{\alpha}.$ Likewise, these inhibitors prevented the protective effect of $TNF{\alpha}.$ In contrast to $TNF{\alpha}-stimulated\;NF-_{\kappa}B$ activity, basal $NF-_{\kappa}B$ activity of BCECs in media containing serum was suppressed only by calphostin C, but not by herbimycin A. As well BCEC death was also induced only by calphostin C in serum-condition. H 89, a PKA inhibitor, did not affect the basal and $TNF{\alpha}-stimulated\;NF-_{\kappa}B$ activities and the protective effect of $TNF{\alpha}$ on cell death. These data suggest that modulation of $NF-_{\kappa}B$ activation could be a possible mechanism for regulating cell viability by protein kinases in BCECs.

• Animal cells and systems
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• 제11권2호
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• pp.205-213
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• 2007

Nek6 belongs to NIMA1 (never in mitosis, gene A) related kinase, which was originally identified in Aspergillus nidulans as a serine/threonine kinase critical for cell cycle progression. We noticed that the putative SUMOylation site is localized on the $K^{252}$ residue in $^{251}FKsD^{254}$ of Nek6, based on the consensus sequence ${\Phi}KxE$; where ${\Phi}$ represents L, I, V or F and x is any amino acid. We observed that the Nek6 SUMO mutant (K252R) has decreased protein kinase activity, nuclear speckle localization and protein stability, compared with that of the Nek6 wild type. However, the Nek6 SUMO mutant increased the cell survival rate of COS-1 cells as determined by FACS analysis. Therefore, our data suggest that SUMOylation on the $K^{252}$ residue of Nek6 is required for its normal functions, such as proper nuclear localization, kinase activity and protein stability, to control cell cycle.

• The Korean Journal of Physiology
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• 제29권1호
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• pp.39-50
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• 1995

This study was designed to clarify the action of cyclic nucleotides, cyclic AMP and cyclic GMP, on phorbol 12,13-dibutyrate (PDBu)-induced contraction in rings isolated from rabbit carotid artery. Arterial rings, 2 mm in width, were myographied isometrically in an isolated organ bath. PDBu produced slowly developing, sustained contraction in rabbit carotid artery, in a dose dependent manner, which was independent of extracellular $Ca^{2+}$ PDBu-induced contraction was relaxed by staurosporine, which suggests that PDBu-induced contraction is mediated by protein kinase C (PKC). $^{45}Ca^{2+}$ uptake by rabbit carotid artery was increased by PDBu during depolarization, but not in control. Isoproterenol and sodium nitroprusside (SNP) relaxed phenylephrine-induced contraction. However, SNP but not isoproterenol relaxed the contraction induced by PDBu. Acetylcholine relaxed PDBu-induced contraction in the presence of the endothelium. 8-bromo-cyclic AMP, a permeable analogue of cyclic AMP, suppressed phenylephrine-induced contraction but not PDBu-induced contraction. 8-bromo cyclic GMP relaxed both of them with dose dependency. A large dose of forskolin relaxed PDBu-induced contraction. PDBu increased cyclic AMP without considerable change in the level of cyclic GMP. Based on these findings, PDBu-induced contraction of rabbit carotid artery was relaxed by cyclic GMP more effectively than cyclic AMP, and the action of cyclic AMP could be mediated by cyclic GMP dependent protein kinase. Therefore it is suggested that the antagonistic action between protein kinase C and cyclic GMP-dependent protein kinase plays a major role in the regulation of vascular tone.

• Bulletin of the Korean Chemical Society
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• 제18권4호
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• pp.428-432
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• 1997

The dephosphorylation specificity of protein phosphatase 2A (PP2A), calcineurin (PP2B) and protein phosphatase 2C (PP2C) were studied in vitro using myelin basic protein (MBP) as a model substrate which was fully phosphorylated at multiple sites by protein kinase C (PKC) or cyclic AMP-dependent protein kinase (PKA). In order to determine the site specificity of phosphates in myelin basic protein, the protein was digested with trypsin and the radioactive phosphopeptide fragments were isolated by high performance liquid chromatography (HPLC) on reversed-phase column. Subsequent analysis and/or sequential manual Edman degradation of the purified phosphopeptides revealed that Thr-65 and Ser-115 were most extensively phophorylated by PKA and Ser-55 by PKC. For the dephosphorylation kinetics, the phosphorylated MBP was treated with calcineurin or PP2C with various time intervals and the reaction was terminated by direct tryptic digest. Both Thr-65 and Ser-115 residues were dephosphorylated more rapidly than any other ones by phosphatases. However it can be differentiated further by first-order kinetics that the PP2B dephosphorylated both Thr-65 and Ser-115 with almost same manner, whereas PP2C dephosphorylated somewhat preferentially the Ser-115.

• BMB Reports
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• 제35권3호
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• pp.343-347
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• 2002

The human protein tyrosine kinase-6 (PTK6) polypeptide that is deduced from the cDNA sequence contains a Src homology (SH) 3 domain, SH2 domain, and catalytic domain of tyrosine kinase. We initiated biochemical and NMR characterization of PTK6 SH3 domain in order to correlate the structural role of the PTK6 using circular dichroism and heteronuclear NMR techniques. The circular dichroism data suggested that the secondary structural elements of the SH3 domain are mainly composed of $\beta$-sheet conformations. It is most stable when the pH is neutral based on the pH titration data. In addition, a number of cross peaks at the low-field area of the proton chemical shift of the NMR spectra indicated that the PTK6 SH3 domain retains a unique and folded conformation at the neutral pH condition. For other pH conditions, the SH3 domain became unstable and aggregated during NMR measurements, indicating that the structural stability is very sensitive to pH environments. Both the NMR and circular dichroism data indicate that the PTK6 SH3 domain experiences a conformational instability, even in an aqueous solution.