• The Korean Journal of Zoology
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• v.38 no.2
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• pp.204-211
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• 1995

In an effort to develop a new therapeutic strategy for human gene therapy of solid ovarian tumor, we studied the expression of the p53 tumor suppressor Sene as well as regulatory subunits of cyclic AMP (cAMP)-dependent protein kinase in human ovarian carcinoma cells. Four cell lines (2774, Caov-3, SK-OV-3 and OVCAR-3) were selected for the analyses. The p53 transcript and protein were detected only in the 2774 cell line by Northern and Western Bnalysis. In the relatively fast growing cell line, SK-OV-3, the %rope 1 a regulstorv subunit (RIA of CAMP-dependent protein kinase was the highest among the four cell lines. The expression level of $RII\beta$ protein was low in the four cell lines examined. These results maw point to a direction to select the target gene(sl to be employed for gene therapy to control the ovarian cancer.

• Journal of Applied Biological Chemistry
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• v.64 no.4
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• pp.323-331
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• 2021

Lincomycin is a lincosamide antibiotic isolated from the actinomycete Streptomyces lincolnensis. Moreover, it has been found to be effective against infections caused by Staphylococcus, Streptococcus, and Bacteroides fragillis. To identify the melanin-inducing properties of lincomycin, we used B16F10 melanoma cells in this study. The melanin content and intracellular tyrosinase activity in the cells were increased by lincomycin, without any cytotoxicity. Western blot analysis indicated that the protein expressions of tyrosinase, tyrosinase related protein 1 (TRP1) and TRP2 increased after lincomycin treatment. In addition, lincomycin enhanced the expression of master transcription regulator of melanogenesis, a microphthalmia-associated transcription factor (MITF). Lincomycin also increased the phosphorylation of p38 mitogen-activated protein kinase (MAPK) and decreased the AKT phosphorylation. Moreover, the activation of tyrosinase activity by lincomycin was inhibited by the treatment with SB203580, which is p38 inhibitor. Furthermore, we also found that lincomycin-induced tyrosinase expression was reduced by H-89, a specific protein kinase A (PKA) inhibitor. These results indicate that lincomycin stimulate melanogenesis via MITF activation via p38 MAPK, AKT, and PKA signal pathways. Thus, lincomycin can potentially be used for treatment of hypopigmentation disorders.

• International Journal of Industrial Entomology and Biomaterials
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• v.22 no.2
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• pp.59-64
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• 2011

Protein kinase C (PKC) is involved in many cellular signaling pathways, it participates in many physiological processes, such as cell cycle, growth, proliferation, differentiation and apoptosis. To investigate the effect of PKC on the silkworm midgut tissue infection of Bombyx mori parvo-like virus (BmPLV), a B. mori atypical protein kinase C (BmaPKC) gene was cloned from larval midgut tissue, expressed in E. coli and purified. Additionally, the BmPLV susceptible silkworm strain and resistant silkworm strain were used to test the effect of the B. mori infection on BmPLV. The result showed that BmaPKC encodes a predicted 586 amino acid protein, which contains a C-terminal kinase domain and an N-terminal regulatory domain. The maximum expression amount of the soluble (His)6-tagged fusion protein was detected after 0.8 mmol/L IPTG was added and cultured at $21^{\circ}C$. The (His) 6-tagged fusion protein revealed about 73 kDa molecular weight which confirmed by western blot and mass spectrography. Furthermore BmaPKC protein were detected at 0-72 h post-infection in BmPLVinfected larval midgut tissue, western blot showed that as time went on, the expression of BmaPKC increased gradually in susceptible strain, the expression quantity on 72 h is 5 times of 0 h. However, in resistant strain, the expression quantity is slightly lower than susceptible strain. But no significant change in resistant strain was observed as time went on. The available data suggest that BmaPKC may involve in the regulation of BmPLV proliferation.

• Toxicological Research
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• v.17
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• pp.251-256
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• 2001

The expression of phase II detoxifying enzymes is affected by a variety of compounds and the induction of the enzymes plays an essential role in chemoprevention. A variety of phytochemicals such as sulfur-containing chemoprotective agents (SCC) may trigger cellular signals and activate phase II gene expression through ARE activation. see induces glutathione S-transferases. Studies were conducted to investigate the role of mitogen-activated protein (MAP) kinase and phosphatidylinositol 3-kinase (PI3-kinase) in the induction of GST (e.g. rGSTA2) by sec. We also studied the MAP kinase pathway responsible for the GST expression by see and compared that with the pathway activated by oxidative stress as a result of sulfur amino acids deprivation (SAAD). see inhibited phosphorylation of ERK1/2 although the effect of see on JNK and p38 MAP kinase was minimal. Wortmannin and LY294002. PI3-kinase inhibitors. abolished the increases in rGSTA2 mRNA and protein levels by SCC. Deprivation of cystine and methionine caused oxidative stress in H4IIE cells. as evidenced by a decrease in the reduced glutathione and an increase in prooxidant production. Electrophoretic mobility shift assay revealed that the ARE complex consisting of Nrf-1/2 and Maf proteins was activated 12~48 h. The rGSTA2 mRNA and protein levels were increased by SAAD. Activation of ARE and induction of rGSTA2 were both completely inhibited by PI3-kinase inhibitors. Inhibition of p38 MAP kinase by SB203580 prevented the ARE-mediated rGSTA2 induction. The results of this study showed that PI3-kinase might play an essential role in the ARE-mediated rGSTA2 induction by see or SAAD and that the dual MAP kinase pathways were responsible for the enzyme induction.

• The Korean Journal of Pharmacology
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• v.31 no.1 s.57
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• pp.115-122
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• 1995

Effects of staurosporine, genistein and pertussis toxin on superoxide and HOCl production in C5a- or PMA-activated neutrophils were investigated. A C5a-induced superoxide and $H_2O_2$ production was inhibited by staurosporine, genistein and pertussis toxin. The stimulatory effect of PMA was inhibited by staurosporine but was not affected by pertussis toxin, whereas it was further promoted by genistein. Staurosporine and genistein inhibited superoxide production by sodium fluoride, but pertussis toxin did not affect it. PMA-induced $H_2O_2$ production was inhibited by staurosporine but was not affected by pertussis toxin. Genistein did not show a stimulatory effect on PMA-induced $H_2O_2$ production. Staurosporine and pertussis toxin inhibited HOCl production by C5a- or PMA, whereas genistein stimulated it. C5a-or PMA-induced myeloperoxidase release was inhibited by genistein, in this response the effect of pertussis toxin was not detected. Staurosporine did not affect the stimulatory effect of PMA on the release. Myeloperoxidase activity was markedly increased by genistein but was not affected by staurosporine and pertussis toxin. These results indicate that the respiratory burst of neutrophils may be regulated by protein kinase C and protein tyrosine kinase. Superoxide production induced by the direct activation of protein kinase C might be affected by protein tyrosine kinase oppositely. Genistein probably pro-motes HOCl production by activating myeloperoxidase.

• BMB Reports
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• v.42 no.1
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• pp.35-40
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• 2009

Protein kinase D2 (PKD2) is a member of the PKD serine/threonine protein kinase family that has been implicated in the regulation of a variety of cellular processes including proliferation, survival, protein trafficking and immune response. In the present study, we report a novel interaction between PKD2 and Lck, a member of the Src tyrosine protein kinase family that is predominantly expressed in T cells. This interaction involved the C-terminal kinase domains of both PKD2 and Lck. Moreover, co-expression of Lck enhanced the tyrosine phosphorylation of PKD2 and increased its kinase activity. Finally, we report that PKD2 enhanced T cell receptor (TCR)-induced nuclear factor of T cell (NFAT) activity in Jurkat T cells. These results suggested that Lck regulated the activity of PKD2 by tyrosine phosphorylation, which in turn may have modulated the physiological functions of PKD2 during TCR-induced T cell activation.

• Archives of Pharmacal Research
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• v.29 no.12
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• pp.1114-1118
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• 2006

(+)-Eudesmin [4,8-bis(3,4-dimethoxyphenyl)-3,7 -dioxabicyclo[3.3.0]octane] was isolated from the stem bark of Magnolia kobus DC. var. borealis Sarg. and found to have neuritogenic activity. $50\;{\mu}M$ (+)-eudesmin induced neurite outgrowth and enhanced nerve growth factor (NGF)-mediated neurite outgrowth from PC12 cells. At this concentration, (+)-eudesmin also enhanced NGF-induced neurite-bearing activity and this activity was partially blocked by various protein kinase inhibitors. These included PD98059, a mitogen-activated protein kinase (MAPK) kinase inhibitor. GF109203X, a protein kinase C (PKC) inhibitor and H89, a protein kinase A (PKA) inhibitor. These results suggest that (+)-eudesmin can induce neurite outgrowth from PC12 cells by stimulating up-stream MAPK, PKC and PKA pathways.

• Journal of Plant Biotechnology
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• v.45 no.4
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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.

• Proceedings of the PSK Conference
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• 2002.10a
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• pp.297.2-297.2
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• 2002

Previous report showed that histamine release by HCI was mediated via reactive oxygen species (ROS) generation in RBL 2H3 cells. To investigate action of protein kinase on histamine release and ROS generation. we observed effects of protein kinase inhibitors on histamine release and ROS generation in RBL 2H3 cells stimulated by HCI HCI dose-dependently increased both histamine release and ROS generation. HCI-induced histamine release was significantly inhibited by bisindolmaleimide (10 ${\mu}$M). DHC (10 ${\mu}$M). , and wortmannin (10 ${\mu}$M), but not by PD098059 (10 ${\mu}$M). ON the other hand. HCI-induced ROS generation was significantly inhibited by DHC (10 ${\mu}$M). but not by bisindolmaleimide(10 ${\mu}$M). wortmannin (10 ${\mu}$M). and PD098059 (10 ${\mu}$M). However KN-62 did not inhibited both. These results showed that involvement of protein kinase in regulation of histamine release and ROS generation may be different and only tyrosine kinase may be associated with regulation of both histamine release and ROS generation in RBL 2H3 cells.

• Journal of Life Science
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• v.25 no.5
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• pp.507-514
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• 2015

D-Xylulose is phosphorylated to D-xylulose-5-phosphate by D-xylulose kinase before it enters glycolysis via the nonoxidative pentose phosphate pathway. A gene encoding a novel D-xylulose kinase (XK) from K. gwangalliensis strain SJ2 was sequenced and expressed in E. coli. The sequence of the isolated XK gene was 1,419 bp, encoding 472 amino acids. The XK protein was more closely related to the Arthrobacter phenanthrenivorans XK than to the Bifidobacterium catenulatum one, as reflected in the sequence identity (54.9% vs. 38.7%). The XK gene was subcloned into the pCold-II expression vector. The resulting plasmid was transformed into E. coli strain BL21 (DE3) cells and the expression of the recombinant XK protein was induced by the addition of IPTG. The resulting protein was expressed as a fusion protein of approximately 48 kDa containing a N-terminal six-histidine extension that was derived from the expression vector. The expressed protein was homogenized by affinity chromatography and showed enzymatic activity corresponding to D-xylulose kinase. XK enzyme kinetic studies with D-xylulose and ATP showed a Km of 250±20 μM and 1,300±50 μM, respectively. The results obtained from this study will provide a wider knowledge base for the characterization of D-xylulose kinase at the molecular level.