• Journal of Life Science
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• v.33 no.2
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• pp.129-137
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• 2023

Reactive oxygen species (ROS) play an essential role in a variety of cellular physiological phenomena. The present study assessed the signaling axis that mediates the lysophosphatidic acid (LPA)-induced migration of SKOV-3 cells. Insulin-like growth factor-1 (IGF-1) stimulated SKOV-3 cell migration in a time- and dose-dependent manner. Similarly, LPA stimulated SKOV-3 cell migration and the phosphorylation of Akt in a time- and dose-dependent manner. The pharmacological inhibition of LPA receptors (LPA₁/LPA₃) significantly suppressed LPA-induced SKOV-3 cell migration. However, IGF-1-induced SKOV-3 cell migration was not affected by the inhibition of LPA1 and LPA3. Pharmacological inhibition of phosphoinositide 3-kinase (PI3K) or Rho-associated kinase (ROCK) significantly suppressed LPA-induced migration, whereas the inhibition of MAPK kinase (MEK) had no effect. Inhibition of PI3K or ROCK completely suppressed LPA-induced ROS generation, and suppression of nicotinamide adenine dinucleotide phosphate oxidase (NOX) or chelation of ROS by N-acetylcysteine (NAC) blocked LPA-induced SKOV-3 cell migration. LPA-induced ROS generation was suppressed by silencing Rictor or Akt1 but not Raptor or Akt2. Silencing Rictor or Akt1 significantly suppressed LPA-induced SKOV-3 cell migration, whereas silencing Raptor or Akt2 had no effect. Finally, the overexpression of the constitutively active form Akt1 (CA-Akt1) significantly enhanced the LPA-induced migration of SKOV-3 cells. Given these results, we suggest that LPA stimulates SKOV-3 cell migration by ROS generation, which is mediated by the mTORC2/Akt1/NOX signaling axis.

• Applied Biological Chemistry
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• v.13 no.2
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• pp.111-129
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• 1970

As a series of studies on the nucleic acids and their related substances 210 samples were collected from 76 places such as farm soil, compost of heap, nuruk and meju to obtain microbial strains which produce 5'-phosphodiesterase. From these samples total of 758 strains were isolated by the use of dilution pour plate method. For all isolated strains primary screening of the productivity of RNA depolymerase was performed and useful strains with regard to 5'-phosphodiesterase productivities were identified. For these useful strains optimum condition, the effect of various compounds on the activity of 5'-phosphodiesterase, and the optimum condition for enzyme reaction were discussed. The quantitative of 5'-mononucleotides produced by the action of 5'-phosphodiesterase was performed using anion-exchange column chromatography and their identified was done by paper chromatography, thinlayer chromatography, ultra violet spectrophotometry, and characteristic color reaction using carbazole and schiff's reagent. (1) Penicillium citreo-viride PO 2-11 and Streptomyces aureus SOA 4-21 from soil were identified as a potent 5'-phosphodiesterase producing strains. (2) Optimum culture conditions for Penicillium citreo-viride PO 2-11 strain isolated were found to be pH 5.0 and $30^{\circ}C$, and the optimum conditions for enzyme action of 5'-phosphodiesterase were pH 4.2 and $60^{\circ}C$. Best carbon source for the production of 5'-phosphodiesterase was found to be sucrose and ammonium nitrate for nitrogen source. Addition of 0.01% corn steep liquor or yeast extract exhibited 20% increase in the amount of 5'-phosphodiesterase production compared to the control. 5'-phosphodiesterase produced by this strain was activated by $Mg^{++},\;Ca^{++},\;Zn^{++},\;Mn^{++}$ and was inhibited by EDTA, citrate, $Cu^{++},\;CO^{++}$. 5'-phosphodiesterase produced 5'-mononucleotide from RNA at a rate of 65.81%, and among the 5'-mononucleotides accumulated 5'-GMP only was found to have flavorous and the strain was also found lack of 5'-AMP deaminase. Productivity of flavorous 5'-GMP was found to be 186.7mg per gram of RNA. (3) Optimum culture canditions for the isolated Streptomyces aureus SOA 4-21 strain were pH 7.0 and $28^{\circ}C$, and the optimum conditions for the action of 5'-phosphodiesterase were pH 7.3 and $50^{\circ}C$. The best carbon source for 5'-phosphodiesterase production was found to be glucose and that of nitrogen was asparagine. Addition of 0.01% yeast extract exhibited increased productivity of 5'-phosphodiesterase by 40% compared to the non-added control. 5'-phosphodiesterase produced by this strain was activated by $Ca^{++},\;Zn^{++},\;Mn^{++}$ and was inhibited by citrate, EDTA, $Cu^{++}$. It was also found that the strain produce 5'-AMP deaminase in addition to 5'-phosphodiesterase. For this reason although decomposition rate was 63.58% the accumulation of 5'-AMP, 5'-CMP, 5'-GMP and 5'-UMP occurred by the breakdown of RNA. In the course of these reaction 5'-AMP deaminase converted 60% of 5'-AMP thus produced into 5'-IMP and flavorous 5'-mono nucleotide production was significantly increased by this strain over the above mentioned one. Production rates were found to be 171.8mg per grain of RNA for 5'-IMP and 148.2mg per gram of RNA for 5'-GMP, respectively.