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5-Lipoxygenase and cysteinyl leukotriene receptor 1 regulate epidermal growth factor-induced cell migration through Tiam1 upregulation and Rac1 activation.

Magi S, Takemoto Y, Kobayashi H, Kasamatsu M, Akita T, Tanaka A, Takano K, Tashiro E, Igarashi Y, Imoto M - Cancer Sci. (2014)

Bottom Line: In this study, we found that 5-lipoxygenase (5-LOX) is activated in the process of EGF-induced cell migration, and that leukotriene C4 (LTC4 ) produced by 5-LOX mediated the second wave of Rac1 activation, as well as cell migration.Furthermore, these effects caused by LTC4 were found to be blocked in the presence of the antagonist of cysteinyl leukotriene receptor 1 (CysLT1).We also found that 5-LOX inhibitors, CysLT1 antagonists and the knockdown of CysLT1 inhibited EGF-induced T cell lymphoma invasion and metastasis-inducing protein 1 (Tiam1) expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Biosciences and Informatics, Faculty of Science and Technology, Keio University, Yokohama, Japan.

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Cysteinyl leukotriene receptor 1 (CysLT1) signaling regulates expression levels of T cell lymphoma invasion and metastasis-inducing protein 1 (Tiam1) in A431 cells. (a, b) Time-course data of epidermal growth factor (EGF)-induced Tiam1 (a) and mRNA (b) in A431 cells. (c) The effect of CysLT1 antagonists on the expression level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to western blotting using the indicated antibodies. (d) The effect of CysLT1 antagonists on the mRNA level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to real-time RT-PCR analysis. (e) The effect of 5-lipoxygenase inhibitors and leukotrienes on the expression level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to western blotting using the indicated antibodies.
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fig04: Cysteinyl leukotriene receptor 1 (CysLT1) signaling regulates expression levels of T cell lymphoma invasion and metastasis-inducing protein 1 (Tiam1) in A431 cells. (a, b) Time-course data of epidermal growth factor (EGF)-induced Tiam1 (a) and mRNA (b) in A431 cells. (c) The effect of CysLT1 antagonists on the expression level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to western blotting using the indicated antibodies. (d) The effect of CysLT1 antagonists on the mRNA level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to real-time RT-PCR analysis. (e) The effect of 5-lipoxygenase inhibitors and leukotrienes on the expression level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to western blotting using the indicated antibodies.

Mentions: We previously demonstrated that Tiam1 might function as the RacGEF responsible for the second EGF-induced wave of Rac1 activation by interacting with 14-3-3ζ. As shown in Figure4(a) and in our previous report,6 the intracellular expression levels of Tiam1 were quite low in unstimulated A431 cells, but gradually increased within 9 h after EGF stimulation. Furthermore, an elevated level of the Tiam1 mRNA was detected by real time RT-PCR analysis in EGF-stimulated A431 cells (Fig.4b); this observation suggests that Tiam1 expression was increased at the level of transcription. However, the regulatory mechanisms underlying EGF-increased Tiam1 expression have not yet been discovered. Therefore, we examined the possibility that CysLT1 signaling regulates the expression level of Tiam1. As shown in Figure4(c,d), the CysLT1 antagonists MK-571 and montelukast suppressed the EGF-induced increase in Tiam1 expression at both the mRNA and protein levels. Similar results were obtained when TT cells (a human thyroid cancer cell line) or EC109 cells (a human esophageal cancer cell line), were used in place of A431 cells (Fig. S3). The 5-LOX inhibitors BU-4664L and AA-861 also inhibited expression, and addition of LTD4 ified this inhibition (Figs4e,S4). Taken together, 5-LOX/LTC4(D4)/CysLT1 signaling regulates the second EGF-induced wave of Rac1 activation and lamellipodia formation by modulating the expression level of Tiam1 protein.


5-Lipoxygenase and cysteinyl leukotriene receptor 1 regulate epidermal growth factor-induced cell migration through Tiam1 upregulation and Rac1 activation.

Magi S, Takemoto Y, Kobayashi H, Kasamatsu M, Akita T, Tanaka A, Takano K, Tashiro E, Igarashi Y, Imoto M - Cancer Sci. (2014)

Cysteinyl leukotriene receptor 1 (CysLT1) signaling regulates expression levels of T cell lymphoma invasion and metastasis-inducing protein 1 (Tiam1) in A431 cells. (a, b) Time-course data of epidermal growth factor (EGF)-induced Tiam1 (a) and mRNA (b) in A431 cells. (c) The effect of CysLT1 antagonists on the expression level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to western blotting using the indicated antibodies. (d) The effect of CysLT1 antagonists on the mRNA level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to real-time RT-PCR analysis. (e) The effect of 5-lipoxygenase inhibitors and leukotrienes on the expression level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to western blotting using the indicated antibodies.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig04: Cysteinyl leukotriene receptor 1 (CysLT1) signaling regulates expression levels of T cell lymphoma invasion and metastasis-inducing protein 1 (Tiam1) in A431 cells. (a, b) Time-course data of epidermal growth factor (EGF)-induced Tiam1 (a) and mRNA (b) in A431 cells. (c) The effect of CysLT1 antagonists on the expression level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to western blotting using the indicated antibodies. (d) The effect of CysLT1 antagonists on the mRNA level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to real-time RT-PCR analysis. (e) The effect of 5-lipoxygenase inhibitors and leukotrienes on the expression level of Tiam1. A431 cells were treated with the indicated concentrations of inhibitors and stimulated with EGF. After 6 h, the cells were subjected to western blotting using the indicated antibodies.
Mentions: We previously demonstrated that Tiam1 might function as the RacGEF responsible for the second EGF-induced wave of Rac1 activation by interacting with 14-3-3ζ. As shown in Figure4(a) and in our previous report,6 the intracellular expression levels of Tiam1 were quite low in unstimulated A431 cells, but gradually increased within 9 h after EGF stimulation. Furthermore, an elevated level of the Tiam1 mRNA was detected by real time RT-PCR analysis in EGF-stimulated A431 cells (Fig.4b); this observation suggests that Tiam1 expression was increased at the level of transcription. However, the regulatory mechanisms underlying EGF-increased Tiam1 expression have not yet been discovered. Therefore, we examined the possibility that CysLT1 signaling regulates the expression level of Tiam1. As shown in Figure4(c,d), the CysLT1 antagonists MK-571 and montelukast suppressed the EGF-induced increase in Tiam1 expression at both the mRNA and protein levels. Similar results were obtained when TT cells (a human thyroid cancer cell line) or EC109 cells (a human esophageal cancer cell line), were used in place of A431 cells (Fig. S3). The 5-LOX inhibitors BU-4664L and AA-861 also inhibited expression, and addition of LTD4 ified this inhibition (Figs4e,S4). Taken together, 5-LOX/LTC4(D4)/CysLT1 signaling regulates the second EGF-induced wave of Rac1 activation and lamellipodia formation by modulating the expression level of Tiam1 protein.

Bottom Line: In this study, we found that 5-lipoxygenase (5-LOX) is activated in the process of EGF-induced cell migration, and that leukotriene C4 (LTC4 ) produced by 5-LOX mediated the second wave of Rac1 activation, as well as cell migration.Furthermore, these effects caused by LTC4 were found to be blocked in the presence of the antagonist of cysteinyl leukotriene receptor 1 (CysLT1).We also found that 5-LOX inhibitors, CysLT1 antagonists and the knockdown of CysLT1 inhibited EGF-induced T cell lymphoma invasion and metastasis-inducing protein 1 (Tiam1) expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Biosciences and Informatics, Faculty of Science and Technology, Keio University, Yokohama, Japan.

Show MeSH
Related in: MedlinePlus