Abstract：
BACKGROUND: Our previous study showed that the abscisic acid receptor lanthionine synthetase C-like 2 (LanCL2) is a significant prognostic factor for overall survival in young glioblastoma patients. However, the role of LanCL2 in glioblastoma remains unclear yet.
OBJECTIVE: This study aims to investigate the role of LanCL2 in regulating in-vitro cell invasion and in-vivo tumor progression of glioblastoma and its underlying mechanism.
METHODS: Tyrosine 198 or 295 residue of LanCL2 was mutated using site-directed mutagenesis to block its phosphorylation. The role of LanCL2 in glioblastoma was investigated using transwell or 3D invasion assay, matrix degradation assay and intracranial xenograft model.
RESULTS: This study showed that nuclear transport of LanCL2 was enhanced by overexpression of LanCL2 or its ligand abscisic acid in glioblastoma cells. Knockdown of LanCL2 suppressed migration, invasion and invadopodia formation of glioblastoma cells, whereas overexpression of wild-type LanCL2 enhanced them. Blocking of Tyr295 residue phosphorylation of LanCL2 impeded its nuclear transport, retarded glioblastoma cell motility and invadopodia formation, and deceased the phosphorylation of Cortactin and STAT3. c-Met was identified as the upstream tyrosine kinase of Tyr295 residue of LanCL2, and inhibition of c-Met markedly suppressed the nuclear transport of LanCL2. Moreover, overexpression of wild-type LanCL2 significantly promoted orthotopic tumor growth of glioblastoma in vivo and led to poor survival of mice with median survival time of 33.5 days, whereas Tyr295 mutation rescued it with median survival time of 49 days.
CONCLUSIONS: Our findings suggested that Tyr295 phosphorylation is crucial to the activation and nuclear transport of LanCL2, as well as invadopodia formation and tumor progression of glioblastoma, providing the evidence of a novel signaling axis c-Met/LanCL2/STAT3/Cortactin and the first observation of the importance of Tyr295 phosphorylation to LanCL2.
OBJECTIVE: This study aims to investigate the role of LanCL2 in regulating in-vitro cell invasion and in-vivo tumor progression of glioblastoma and its underlying mechanism.
METHODS: Tyrosine 198 or 295 residue of LanCL2 was mutated using site-directed mutagenesis to block its phosphorylation. The role of LanCL2 in glioblastoma was investigated using transwell or 3D invasion assay, matrix degradation assay and intracranial xenograft model.
RESULTS: This study showed that nuclear transport of LanCL2 was enhanced by overexpression of LanCL2 or its ligand abscisic acid in glioblastoma cells. Knockdown of LanCL2 suppressed migration, invasion and invadopodia formation of glioblastoma cells, whereas overexpression of wild-type LanCL2 enhanced them. Blocking of Tyr295 residue phosphorylation of LanCL2 impeded its nuclear transport, retarded glioblastoma cell motility and invadopodia formation, and deceased the phosphorylation of Cortactin and STAT3. c-Met was identified as the upstream tyrosine kinase of Tyr295 residue of LanCL2, and inhibition of c-Met markedly suppressed the nuclear transport of LanCL2. Moreover, overexpression of wild-type LanCL2 significantly promoted orthotopic tumor growth of glioblastoma in vivo and led to poor survival of mice with median survival time of 33.5 days, whereas Tyr295 mutation rescued it with median survival time of 49 days.
CONCLUSIONS: Our findings suggested that Tyr295 phosphorylation is crucial to the activation and nuclear transport of LanCL2, as well as invadopodia formation and tumor progression of glioblastoma, providing the evidence of a novel signaling axis c-Met/LanCL2/STAT3/Cortactin and the first observation of the importance of Tyr295 phosphorylation to LanCL2.
摘要：
背景：脱落酸受体蓝硫氨酸合成酶C样2（LanCL2）是年轻胶质母细胞瘤患者总体生存的预后因素。然而,LanCL2在胶质母细胞瘤中的作用和潜在机制尚不清楚.
目的：本研究探讨LanCL2对胶质母细胞瘤体外细胞侵袭和体内肿瘤进展的调控作用及其机制。
方法:使用定点诱变使LanCL2的酪氨酸198或295残基突变以阻断其磷酸化。使用transwell或3D侵袭试验研究了LanCL2在胶质母细胞瘤中的作用，基质降解试验和颅内异种移植模型。
结果：这项研究显示了LanCL2的核定位信号，并通过脱落酸或LanCL2的过表达在胶质母细胞瘤细胞中增加了核转运。击倒LanCL2抑制迁移，胶质母细胞瘤细胞的侵袭和侵袭足形成，而野生型LanCL2的过表达增强了它们。LanCL2的Tyr295残基磷酸化的阻断阻碍了其核运输，胶质母细胞瘤细胞运动性和侵袭足形成迟缓，并降低了Cortactin和STAT3的磷酸化。c-Met被鉴定为LanCL2的Tyr295残基的上游酪氨酸激酶,并且c-Met的抑制显著抑制了LanCL2的核转运。此外，过表达野生型LanCL2显著促进胶质母细胞瘤的体内原位肿瘤生长,导致小鼠中位生存时间33.5天,而Tyr295突变拯救了它,中位生存时间为49天.
结论：我们的研究结果表明，Tyr295的磷酸化对LanCL2的激活和核运输以及GBM的侵袭足形成和肿瘤进展至关重要，提供新的信号轴c-Met/LanCL2/STAT3/Cortactin的证据，并首次观察到Tyr295磷酸化对LanCL2的重要性。
目的：本研究探讨LanCL2对胶质母细胞瘤体外细胞侵袭和体内肿瘤进展的调控作用及其机制。
方法:使用定点诱变使LanCL2的酪氨酸198或295残基突变以阻断其磷酸化。使用transwell或3D侵袭试验研究了LanCL2在胶质母细胞瘤中的作用，基质降解试验和颅内异种移植模型。
结果：这项研究显示了LanCL2的核定位信号，并通过脱落酸或LanCL2的过表达在胶质母细胞瘤细胞中增加了核转运。击倒LanCL2抑制迁移，胶质母细胞瘤细胞的侵袭和侵袭足形成，而野生型LanCL2的过表达增强了它们。LanCL2的Tyr295残基磷酸化的阻断阻碍了其核运输，胶质母细胞瘤细胞运动性和侵袭足形成迟缓，并降低了Cortactin和STAT3的磷酸化。c-Met被鉴定为LanCL2的Tyr295残基的上游酪氨酸激酶,并且c-Met的抑制显著抑制了LanCL2的核转运。此外，过表达野生型LanCL2显著促进胶质母细胞瘤的体内原位肿瘤生长,导致小鼠中位生存时间33.5天,而Tyr295突变拯救了它,中位生存时间为49天.
结论：我们的研究结果表明，Tyr295的磷酸化对LanCL2的激活和核运输以及GBM的侵袭足形成和肿瘤进展至关重要，提供新的信号轴c-Met/LanCL2/STAT3/Cortactin的证据，并首次观察到Tyr295磷酸化对LanCL2的重要性。
