• Title, Summary, Keyword: PI3K

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Cloning and Characterization of Phosphoinositide 3-Kinase γ cDNA from Flounder (Paralichthys olivaceus) (넙치에서 분리된 phosphoinositide 3-kinase γ 유전자의 클로닝 및 특성 연구)

  • Jeong, Tae Hyug;Youn, Joo Yeon;Ji, Keunho;Seo, Yong Bae;Kim, Young Tae
    • Journal of Life Science
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    • v.24 no.4
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    • pp.343-351
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    • 2014
  • Phosphoinositide 3-kinase (PI3K) plays a central role in cell signaling and leads to cell proliferation, survival, motility, exocytosis, and cytoskeletal rearrangements, as well as specialized cell responses, superoxide production, and cardiac myocyte growth. PI3K is divided into three classes; type I PI3K is preferentially expressed in leukocytes and activated by ${\beta}{\gamma}$ subunits of heterotrimeric G-proteins. In this study, the cDNAs encoding the $PI3K{\gamma}$ gene were isolated from a brain cDNA library constructed using the flounder (Paralichthys olivaceus). The sequence of the isolated $PI3K{\gamma}$ was 1341 bp, encoding 447 amino acids. The nucleotide sequence of the $PI3K{\gamma}$ gene was analyzed with that of other species, including Oreochromis niloticus and Danio rerio, and it turned out to be well conserved during evolution. The $PI3K{\gamma}$ gene was subcloned into the expression vector pET-44a(+), and expressed in the E. coli BL21 (DE3) codon plus cell. The resulting protein was expressed as a fusion protein of approximately 49 kDa containing a C-terminal six-histidine extension that was derived from the expression vector. The expressed protein was purified to homogeneity by His-tag affinity chromatography and showed enzymatic activity corresponding to $PI3K{\gamma}$. The binding of wortmannin to $PI3K{\gamma}$, as detected by anti-wortmannin antisera, closely followed the inhibition of the kinase activities. The results obtained from this study will provide a wider base of knowledge on the primary structure and characterization of the $PI3K{\gamma}$ at the molecular level.

Activation of Phosphatidylinositol 3-kinase(PI3K) is Required for Invasiveness and Motility in H-ras MCE10A Cells

  • Shin, Il-Chung;Aree Moon
    • Proceedings of the Korean Society of Applied Pharmacology
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    • pp.103-103
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    • 2001
  • 인간유방상피세포에서 H-ras가 침윤성과 세포 이동성을 유도한다는 것을 이 전연구에서 밝혔다. Phosphatidylinositol 3-kinase(PI3K)는 세포 이동성에서 중요한 역할을 하는 것으로 보고되고 있다. 본 연구에서 인간유방상피세포인 MCF10A에서 H-ras에 의해 유도된 침윤성에 PI3K가 어떠한 영향을 미치는지 살펴보고자 하였다. PI3K의 활성은 PI3K의 downstream molecule인 Akt의 인산화를 Western blot으로 확인하였다. Akt는 MCF10A, H-ras, N-ras MCF10A 세포에서 같은 정도로 발현되는 반면, 인산화된 Akt는 MCF10A 세포에 비해 H-ras MCF10A 세포와 N-ras MCF10A 세포에서 현저히 높게 나타났다. 이상의 결과로서 H-ras, N-ras 둘 다 PI3K를 활성화시키며, 침윤성과 세포이동성이 없는 N-ras MCF10A 세포에서도 PI3K가 활성화되었으므로, PI3K의 활성은 세포침윤성과 이동성을 유도하는데에 있어서 충분하지는 않음을 말해준다. PI3K의 저해제인 LY294002와 wortmannin을 세포에 처리하였을 때 세포침윤성과 이동성이 유의성 있게 감소되었다. 이상의 결과는 MCF10A 세포의 침윤성과 이동설에 있어서 PI3K의 활성이 충분하지는 않지만 반드시 필요하다는 것을 알 수 있었다.

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Role of p53-dependent PI3K in Radioresistance of Colon Cancer Cells (대장암 세포의 방사선저항성에 대한 p53의존성 PI3K의 역할)

  • Lee, Heui-Kwan;Kim, Jong-Suk;Kwon, Hyoung-Cheol
    • Journal of Food Hygiene and Safety
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    • v.25 no.3
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    • pp.258-262
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    • 2010
  • Radiotherapy is one of the major therapies for cancer treatment. p53 acts as a central mediator of the cellular response to stressful stimuli, such as radiation. Recently it has been known that activation of the phosphatidylinositol-3-kinase (PI3K) pathway is associated with radioresistance. In this study, we investigated whether X-irradiation up-regulates PI3K in a p53-dependent manner in human colon cancer cells. In order to study this phenomenon, we have treated p53-wild type and p53-mutant type HCT116 cells with X-ray. Treatment of wild type HCT116 cells with 8 Gy resulted in a marked increase in PI3K (p85), which paralleled an increase in PTEN, a counterpart of PI3K. However, these effects of X-rays in the p53-mutant cells were not observed. These results suggest that the X-irradiation-induced up-regulation of PI3K/PTEN pathway is p53-dependent.

Involvement of Phosphatidylinositol 3-Kinase in the Insulin Signaling in Preimplantation Mouse Embryos (생쥐 착상전 배아의 인슐린 신호전달 과정에 Phosphatidylinositol 3-Kinase의 관련성)

  • Gye, Myung-Chan;Nah, Hee-Young;Kim, Moon-Kyoo
    • Development and Reproduction
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    • v.4 no.1
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    • pp.29-35
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    • 2000
  • A phosphatidylinositol 3-kinase (PI3K) is a upstream component of insulin signaling by which protein synthesis can be stimulated in many systems. To elucidate involvement of PI3K and its downstream mammalian target of rapamycin (mTOR) in the insulin signaling in pleimplantation mouse embryos, 8-cell embryos were cultured to blastocysts in the presence or absence of insulin and/or inhibitor drugs. The number of blastomeres per blastocyst, protein synthesis, and protein phosphorylation were examined. There was significant difference in embryonic development to blastocyst stage and hatching was potentiated by the insulin supplementation. The increase in the mean celt numbers per blastocyst was apparent in the insulin culture. Wortmannin, a PI3K inhibitor and rapamycin, an inhibitor of mTOR abolished the stimulatory effect of insulin on morphological development mitosis and protein synthesis. In autoradiography, phosphoproteins pp22 and pp30 which undergo phosphorylation in response to insulin were identified. Taken together, it can be suggested that PI3K and mTOR engaged in insulin signaling in the mouse embryo 8-cell onward and mediate embryotropic offset of insulin.

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Inhibition of PI3K/AKT Signaling Pathway Enhances Cordycepin-Induced Apoptosis in Human Gastric Cancer Cells (인체위암 세포에서 PI3K/AKT 신호 전달계 차단에 의한 동충하초 유래 Cordycepin의 Apoptosis 유발 효과 증진)

  • Lee, Hye Hyeon;Jeong, Jin-Woo;Choi, Yung Hyun
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.45 no.6
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    • pp.835-842
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    • 2016
  • The phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway plays a crucial role in cancer occurrence by promoting cell proliferation and inhibiting apoptosis. In the present study, we evaluated the effect of a PI3K inhibitor, LY294002, on the chemosensitivity of gastric cancer cells to cordycepin, a predominant functional component of the fungus Cordyceps militaris, in AGS human gastric cancer cells and investigated possible underlying cellular mechanisms. Our results revealed that cordycepin inhibited viability of AGS cells in a concentration-dependent manner and induced apoptosis, as determined by apoptotic cell morphologies and fluorescence-activated cell sorting analysis associated with attenuated activation of the PI3K/Akt signaling pathway. Treatment with cordycepin in combination with a subtoxic concentration of LY294002 enhanced cordycepin-induced cytotoxicity and apoptotic potentials in AGS cells. Sensitization of LY294002 to cordycepin-induced apoptosis was accompanied by activation of caspases (caspases-3, -8, and -9) and was concomitant with poly(ADP-ribose) polymerase cleavage. Moreover, LY294002 up-regulated pro-apoptotic Bax and enhanced truncation of Bid in cordycepin-treated AGS cells, which was connected with increased loss of mitochondrial membrane potential and release of cytochrome c from mitochondria to the cytosol. Taken together, these results indicate that inhibition of the PI3K/Akt signaling pathway could augment cordycepin-induced apoptosis in human gastric cancer cells by up-regulating caspase activity through mitochondrial dysfunction.

Roles of PI3K and Rac Pathways in H-ras Induced Invasion and Motility

  • Ilchung Shin;Kim, Seonhoe;Aree Moon
    • Proceedings of the Korean Society of Toxicology Conference
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    • pp.144-144
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    • 2003
  • Phosphatidylinositol 3-kinase (PI3K) and Rac play important roles that regulate cellular functions including cell survival and .migration. In the present study, we investigated the functional roles of PI3K and Rac1 pathways in H-ras-induced invasive phenotype and motility of MCF10A cells.(omitted)

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Pharmacophore Based Screening and Molecular Docking Study of PI3K Inhibitors

  • Rupa, Mottadi;Madhavan, Thirumurthy
    • Journal of the Chosun Natural Science
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    • v.9 no.1
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    • pp.41-61
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    • 2016
  • Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related mortality worldwide. Phosphoinositide 3-kinases (PI3Ks) play important role in Non-Small Cell Lung Cancer. PI3Ks constitute a lipid kinase family which modulates the function of numerous substrates involved in the regulation of cell survival, cell cycle progression and cellular growth. Herein, we describe the ligand based pharmacophore combined with molecular docking studies methods to identify new potent PI3K inhibitors. Several pharmacophore models were generated and validated by Guner-Henry scoring Method. The best models were utilized as 3D pharmacophore query to screen against ZINC database (Chemical and Natural) and the retrieved hits were further validated by fitness score, Lipinski's rule of five. Finally four compounds were found to have good potential and they may act as novel lead compounds for PI3K inhibitor designing.

Opisthorchis viverrini Infection Activates the PI3K/AKT/PTEN and Wnt/β-catenin Signaling Pathways in a Cholangiocarcinogenesis Model

  • Yothaisong, Supak;Thanee, Malinee;Namwat, Nisana;Yongvanit, Puangrat;Boonmars, Thidarut;Puapairoj, Anucha;Loilome, Watcharin
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.23
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    • pp.10463-10468
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    • 2015
  • Opisthorchis viverrini (Ov) infection is the major etiological factor for cholangiocarcinoma (CCA), especially in northeast Thailand. We have previously reported significant involvement of PI3K/AKT/PTEN and $Wnt/{\beta}$-catenin in human CCA tissues. The present study, therefore, examined the expression and activation of PI3K/AKT/PTEN and $Wnt/{\beta}$-catenin signaling components during Ov-induced cholangiocarcinogenesis in a hamster animal model. Hamsters were divided into two groups; non-treated and Ov plus NDMA treated. The results of immunohistochemical staining showed an upregulation of PI3K/AKT signaling as determined by elevated expression of the $p85{\alpha}$-regulatory and $p110{\alpha}$-catalytic subunits of PI3K as well as increased expression and activation of AKT during cholangiocarcinogenesis. Interestingly, the staining intensity of activated AKT (p-AKT) increased in the apical regions of the bile ducts and strong staining was detected where the liver fluke resides. Moreover, PTEN, a negative regulator of PI3K/AKT, was suppressed by decreased expression and increased phosphorylation during cholangiocarcinogenesis. We also detected upregulation of $Wnt/{\beta}$-catenin signaling as determined by increased positive staining of Wnt3, Wnt3a, Wnt5a, Wnt7b and ${\beta}$-catenin, corresponded with the period of cholangiocarcinogenesis. Furthermore, nuclear staining of ${\beta}$-catenin was observed in CCA tissues. Our results suggest the liver fluke infection causes chronic inflammatory conditions which lead to upregulation of the PI3K/AKT and $Wnt/{\beta}$-catenin signaling pathways which may drive CCA carcinogenesis. These results provide useful information for drug development, prevention and treatment of CCA.

Odorant Stimulation Promotes Survival of Rodent Olfactory Receptor Neurons via PI3K/Akt Activation and Bcl-2 Expression

  • Kim, So Yeun;Yoo, Seung-Jun;Ronnett, Gabriele V;Kim, Eun-Kyoung;Moon, Cheil
    • Molecules and Cells
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    • v.38 no.6
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    • pp.535-539
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    • 2015
  • Olfactory stimulation activates multiple signaling cascades in order to mediate activity-driven changes in gene expression that promote neuronal survival. To date, the mechanisms involved in activity-dependent olfactory neuronal survival have yet to be fully elucidated. In the current study, we observed that olfactory sensory stimulation, which caused neuronal activation, promoted activation of the phosphatidylinositol 3'-kinase (PI3K)/Akt pathway and the expression of Bcl-2, which were responsible for olfactory receptor neuron (ORN) survival. We demonstrated that Bcl-2 expression increased after odorant stimulation both in vivo and in vitro. We also showed that odorant stimulation activated Akt, and that Akt activation was completely blocked by incubation with both a PI3K inhibitor (LY294002) and Akt1 small interfering RNA. Moreover, blocking the PI3K/Akt pathway diminished the odorantinduced Bcl-2 expression, as well as the effects on odorant-induced ORN survival. A temporal difference was noted between the activation of Akt1 and the expression of Bcl-2 following odorant stimulation. Blocking the PI3K/Akt pathway did not affect ORN survival in the time range prior to the increase in Bcl-2 expression, implying that these two events, activation of the PI3K pathway and Bcl-2 induction, were tightly connected to promote post-translational ORN survival. Collectively, our results indicated that olfactory activity activated PI3K/Akt, induced Bcl-2, and promoted long term ORN survival as a result.

Curcumin targets vascular endothelial growth factor via activating the PI3K/Akt signaling pathway and improves brain hypoxic-ischemic injury in neonatal rats

  • Li, Jia;An, Yan;Wang, Jia-Ning;Yin, Xiao-Ping;Zhou, Huan;Wang, Yong-Sheng
    • The Korean Journal of Physiology and Pharmacology
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    • v.24 no.5
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    • pp.423-431
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    • 2020
  • This study aimed to evaluate the effect of curcumin on brain hypoxic-ischemic (HI) damage in neonatal rats and whether the phosphoinositide 3-kinase (PI3K)/Akt/vascular endothelial growth factor (VEGF) signaling pathway is involved. Brain HI damage models were established in neonatal rats, which received the following treatments: curcumin by intraperitoneal injection before injury, insulin-like growth factor 1 (IGF-1) by subcutaneous injection after injury, and VEGF by intracerebroventricular injection after injury. This was followed by neurological evaluation, hemodynamic measurements, histopathological assessment, TUNEL assay, flow cytometry, and western blotting to assess the expression of p-PI3K, PI3K, p-Akt, Akt, and VEGF. Compared with rats that underwent sham operation, rats with brain HI damage showed remarkably increased neurological deficits, reduced right blood flow volume, elevated blood viscosity and haematocrit, and aggravated cell damage and apoptosis; these injuries were significantly improved by curcumin pretreatment. Meanwhile, brain HI damage induced the overexpression of p-PI3K, p-Akt, and VEGF, while curcumin pretreatment inhibited the expression of these proteins. In addition, IGF-1 treatment rescued the curcumin-induced down-regulated expression of p-PI3K, p-Akt, and VEGF, and VEGF overexpression counteracted the inhibitory effect of curcumin on brain HI damage. Overall, pretreatment with curcumin protected against brain HI damage by targeting VEGF via the PI3K/Akt signaling pathway in neonatal rats.