• BMB Reports
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• 제34권6호
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• pp.517-525
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• 2001

Six renaturable protein kinases that utilize the myelin basic protein (MBP) as a substrate were activated during prolonged exposure of cardiac myocytes to okadaic acid (OA). We characterized the substrate preference and activation of these kinases, with particular emphasis on 3 novel kinases-MBPK-55, MBPK-62 and MBPK-87. The transcription factors c-Jun, Elk, ATF2, and c-Fos that are used to assess mitogen-activated protein kinase activation were all poor substrates for these three kinases. MAPKAPK2 was also not phosphorylated. In contrast, Histone IIIS was phosphorylated by MBPK-55 and MBPK-62. These protein kinases were activated in cultured cardiac fibroblasts, H9c2 cardiac myoblasts, and Cos cells. High concentrations (0.5 to $1\;{\mu}M$) of OA were essential for the activation of the protein kinases in all of the cell types examined, whereas calyculin A [an inhibitor of protein phosphatase 1 (PP1) and PP2A], cyclosporin A (a PP2B inhibitor), and an inactive OA analog all failed to activate these kinases. The high dose of okadaic acid that is required for kinase activation was also required for phosphatase inhibition, as assessed by immunoblotting whole cell lysates with anti-phosphothreonine antibodies. A variety of chemical inhibitors, including PD98059 (MEK-specific), genistein (tyrosine kinase-specific) and Bisindolylmaleimide I (protein kinase C-specific), failed to inhibit the OA activation of these kinases. Thus, MBPK-55 and MBPK-62 are also Histone IIIS kinases that are widely expressed and specifically activated upon exposure to high OA concentrations.

• BMB Reports
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• 제56권11호
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• pp.584-593
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• 2023

Hypoxia, a widespread occurrence observed in various malignant tumors, results from rapid tumor growth that outpaces the oxygen supply. Tumor hypoxia precipitates several effects on tumor biology; these include activating angiogenesis, intensifying invasiveness, enhancing the survival of tumor cells, suppressing anti-tumor immunity, and fostering resistance to therapy. Aligned with the findings that correlate CMGC kinases with the regulation of Hypoxia-Inducible Factor (HIF), a pivotal modulator, reports also indicate that hypoxia governs the activity of CMGC kinases, including DYRK1 kinases. Prolyl hydroxylation of DYRK1 kinases by PHD1 constitutes a novel mechanism of kinase maturation and activation. This modification "primes" DYRK1 kinases for subsequent tyrosine autophosphorylation, a vital step in their activation cascade. This mechanism adds a layer of intricacy to comprehending the regulation of CMGC kinases, and underscores the complex interplay between distinct post-translational modifications in harmonizing precise kinase activity. Overall, hypoxia assumes a substantial role in cancer progression, influencing diverse aspects of tumor biology that include angiogenesis, invasiveness, cell survival, and resistance to treatment. CMGC kinases are deeply entwined in its regulation. To fathom the molecular mechanisms underpinning hypoxia's impact on cancer cells, comprehending how hypoxia and prolyl hydroxylation govern the activity of CMGC kinases, including DYRK1 kinases, becomes imperative. This insight may pave the way for pioneering therapeutic approaches that target the hypoxic tumor microenvironment and its associated challenges.

• Bulletin of the Korean Chemical Society
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• 제34권2호
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• pp.437-442
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• 2013

A new series of 4-(2-(substituted)pyridin-4-yl)-3-(3-methoxy-5-methylphenyl)-1H-pyrazoles (4a-f) and their 1,2-isoxazole analogues (5a-f) has been rationally designed, synthesized and screened against both ROS and MAPK14 kinases. Compounds 4b, 4c and 4e showed moderate inhibitions against both ROS and MAPK14 kinases. Compound 4e has showed the strongest inhibitions with IC50 values of 1.25 ${\mu}M$ and 3.00 ${\mu}M$ against ROS and MAPK14 kinases, respectively. A brief structure-activity relationship study and a molecular modeling study were made revealing a group of essential structural features for good kinase inhibitory activity within this new class of kinase inhibitors.

• 한국미생물·생명공학회지
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• 제35권2호
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• pp.112-117
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• 2007

Protein kinase는 진핵생물과 원핵생물을 포함하는 모든 생명체에서 세포생존에 절대적으로 중요한 조절 기능을 담당한다. 일반적으로 원핵생물은 histidine 과 aspartic acid kinase로 구성된 bacterial two-component regulatory system에 의하여 환경변화에 따른 유전자의 발현이 조절되지만, 방선균을 비롯한 고등 원핵생물에서는 진핵생물성의 serine/threonine kinase들이 세포분화와 같은 분화과정을 조절하고 있다. Streptomycin 생산균인 Streptomyces griseus 균주에서도 다양한 serine/threonine kinase들이 존재하는 것으로 추정되며, 이들의 기능을 밝히는 것은 생명현상을 이해하는 중요한 열쇠를 제공해 줄 것으로 기대된다. 따라서, S. griseus로부터 protein kinase 를 동정하는 연구를 실시하였으며, 기존의 복잡한 chromatography법의 단점을 보완하기 위해 anti-phosphothreonine, anti-phosphoserine, anti-phosphotyrosine antibody를 이용한 immunoaffinity column chromatography 방법을 도입하였다. 실험 결과 약 14, 29, 31, 35, 40, 52, 56, 60 kDa의 단백질을 효과적으로 동정 할 수 있었으며, nonradioactive protein kination assay 방법으로 이들의 인산화능을 확인하였다.

• Journal of Plant Biotechnology
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• 제42권4호
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• pp.277-283
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• 2015

Plant genome analyses, including Arabidopsis thaliana showed a large gene family of plant receptor kinases with various extracellular ligand-binding domain. Now intensively studies to understand physiological and cellular functions for higher plant receptor kinases in diverse and complex biological processes including plant growth, development, ligands perception including steroid hormone and plant-microbe interactions. Brassinosteroids (BRs) as a one of well know steroid hormone are plant growth hormones that control biomass accumulation and also tolerance to many biotic and abiotic stress conditions and hence are of relevance to agriculture. BRI1 receptor kinase, which is localized in plasma membrane in the cell sense BRs and it bind to a receptor protein known as BRASSINOSTEROID INSENSITIVE 1 (BRI1). Recently, we reported that BRI1 and its co-receptor, BRI1-ASSOCIATED KINASE (BAK1) autophosphorylated on tyrosine residue (s) in vitro and in vivo and thus are dual-specificity kinases. Other plant receptor kinases are also phosphorylated on tyrosine residue (s). Post-translational modifications (PTMs) can be studied by altering the residue modified by directed mutagenesis to mimic the modified state or to prevent the modification. These approaches are useful to not only characterize the regulatory role of a given modification, but may also provide opportunities for plant improvement.

• Journal of Plant Biotechnology
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• 제45권4호
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• pp.315-321
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• 2018

Molecular and functional characterization of proteins and their levels is of great interest in understanding the mechanism of diverse cellular processes. In this study, we report on the convenient Escherichia coli-based protein expression system that allows recombinant of soluble proteins expression and cytosolic domain of membrane-localised kinases, followed by the detection of autophosphorylation activity in protein kinases. This approach is applied to regulatory proteins of Arabidopsis thaliana, including 14-3-3, calmodulin, calcium-dependent protein kinase, TERMINAL FLOWER 1(TFL1), FLOWERING LOCUS T (FT), receptor-like cytoplasmic kinase and cytoplasmic domain of leucine-rich repeat-receptor like kinase proteins. Our Western blot analysis which uses phospho-specific antibodies showed that five putative LRR-RLKs and two putative RLCKs have autophosphorylation activity in vitro on threonine and/or tyrosine residue(s), suggesting their potential role in signal transduction pathways. Our findings were also discussed in the broader context of recombinant expression and biochemical analysis of soluble and membrane-localised receptor kinases in microbial systems.

• 한국독성학회:학술대회논문집
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• 한국독성학회 2001년도 International Symposium on Dietary and Medicinal Antimutgens and Anticarcinogens
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• pp.29-30
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• 2001

MAP kinase/ AP-1 signal transduction components are rapidly initiated by many extracellular stimuli, especially environmental carcinogens. We have investigated the role of MAP kinases (Erks, JNKs, and p38 kinases) and AP-1 signal transduction pathways in the process of cell transformation and carcinogenesis. Incubation of Cl 41 cells with tumor promoters such as TPA, EGF, arsenic, or TNF-$\alpha$ led to cell transformation and activation of MAP kinases.(omitted)

• 대한의생명과학회지
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• 제14권1호
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• pp.59-62
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• 2008

This experiment was performed to study the effect of TSH (thyroid-stimulating hormone) on the expression of endoplasmic reticulum (ER) chaperones in the rat thyrocytes FRTL-5 cells. Although the expressions of ER membrane kinases (ATF6, IRE1 and PERK) were specially enhanced under absence of TSH, no remarkable up- or down regulations of ER chaperones (BiP, CHOP and Calnexin) were detected by TSH. We firstly report here that TSH by dose up-regulated expression of ER membrane kinases in FRTL-5 culture thyrocytes.

• BMB Reports
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• 제46권6호
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• pp.289-294
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• 2013

To maintain cellular homeostasis against the demands of the extracellular environment, a precise regulation of kinases and phosphatases is essential. In cell cycle regulation mechanisms, activation of the cyclin-dependent kinase (CDK1) and cyclin B complex (CDK1:cyclin B) causes a remarkable change in protein phosphorylation. Activation of CDK1:cyclin B is regulated by two auto-amplification loops-CDK1:cyclin B activates Cdc25, its own activating phosphatase, and inhibits Wee1, its own inhibiting kinase. Recent biological evidence has revealed that the inhibition of its counteracting phosphatase activity also occurs, and it is parallel to CDK1:cyclin B activation during mitosis. Phosphatase regulation of mitotic kinases and their substrates is essential to ensure that the progression of the cell cycle is ordered. Outlining how the mutual control of kinases and phosphatases governs the localization and timing of cell division will give us a new understanding about cell cycle regulation.

• 한국동물학회지
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• 제37권1호
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• pp.130-136
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• 1994

Although alteration in protein phosphorylation by specific protein kinases is of importance in transducing cellular signals in a variety of neural/endocrine systems, little is known about protein phosphorylation in the hvpothalamus. The present study aims to explore whether activation of the second messenger-dependent protein kinases affects phosphorylation of specific proteins using a cell free phosphorylation system followed by SDS-polvacrylamide gel electrophoresis. Cytoplasmic fractions derived from hvpothalami of immature rats were used as substrates and several activators and/or inhibitors of CAMP-, phosphatidylinositol- and Ca2+-calmodulin-dependent protein kinases were assessed. Many endogenous proteins were extensively phosphorylated and depending on the signal transduction pathways, phosphorvlation profiles were markedly different. The present data indicate that extracellular signals may affect cellular events through protein phosphorylation by second messengers-protein kinases in the rat hypothalamus.