探討Semaphorin 5A細胞膜外區域在非小細胞肺癌中抑制腫瘤的角色

Identification of the Tumor-Suppressing Roles of Semaphorin 5A Extracellular Domain in Non-Small-Cell Lung Cancer

陳姳蓁(Ming-Zhen Chen)

指導教授 ： 賴亮全

國立臺灣大學/醫學院/生理學研究所/碩士(2020年)

https://doi.org/10.6342/NTU202003914

摘要

Semaphorin 5A (SEMA5A)是semaphorin家族的一員，被視為是參與神經軸突發展的重要分子。在先前的研究中我們發現SEMA5A 可以做為非吸菸肺癌女性病患的生物性標誌物，並且在肺癌細胞中有抑制癌細胞生長及轉移的功能。然而，SEMA5A所參與的調控機制目前仍尚未明朗，因此本篇研究的目的便是探討SEMA5A不同的蛋白區域在抑制肺癌細胞的功能中所扮演的角色。首先，我們在大量表現SEMA5A以及SEMA5A細膜外區域的組別中，觀察到對於肺癌細胞A549及H1299都有顯著的抑制生長及轉移的效果。接著，我們以基因微陣列分析來檢驗大量表現不同SEMA5A蛋白區域後，肺癌細胞A549的基因表現情況。根據分析的結果，我們發現在干擾素(interferon)訊息傳遞途徑中的基因，例如STAT1, STAT2, IRF9, G1P2 以及 IFIT1，在大量表現SEMA5A以及SEMA5A細膜外區域的組別中有顯著上升的情況。另外，於先前的研究中發現，分泌型的SEMA5A細膜外區域與腫瘤的發生有關，然而在抑制脫落酶ADAM17使SEMA5A膜外區域的釋出減少後，並沒有減弱抑癌的效果，所以我們推論另有未發現的蛋白參與SEMA5A的抑癌機制。因此，我們想藉由質譜儀分析來找出與SEMA5A有交互作用的蛋白。然而目前只有在共同免疫沉澱法以及質譜儀分析中找到正控制的SEMA5A，後續仍然需要更多的研究來探討抑癌機制中會與SEMA5A有交互作用的蛋白。總結，在本篇研究中，我們發現SEMA5A膜外區域會透過增加干擾素傳遞途徑中的抑癌基因，來造成抑制肺癌細胞的效果。

關鍵字

肺癌 ； Semaphorin5A ； 膜外區域 ； 抑癌

並列摘要

Semaphorin 5A (SEMA5A) is one member of semaphorin family, which was identified as an axon guidance molecule in nervous systems. In our previous studies, SEMA5A was identified as a prognostic biomarker for lung cancer in nonsmoking women and acted as a tumor suppressor gene in inhibiting the proliferation and migration of lung cancer cells. Yet, the regulatory mechanism of SEMA5A was not clear. Therefore, the purpose of this study was to explore the roles of different domains of SEMA5A on the tumor suppressive effects in lung cancer cell lines. First, we found the overexpression of 5A-Full and 5A-ECD significantly inhibited the proliferation and migration in both A549 and H1299 cells. Next, microarray analyses were performed in lung cancer A549 cells expressing different domains. The results showed the expression levels of genes involved in interferon signaling pathway, such as STAT1, STAT2, IRF9, G1P2 and IFIT1, were significantly increased in overexpressing SEMA5A full length (5A-Full) and SEMA5A extracellular domain (5A-ECD) groups. The secreted form of SEMA5A was reported to be involved in tumor progression. However, tumor-suppressing effects were still observed in cells silencing the sheddase ADAM17 to decrease cleaving the extracellular domain of SEMA5A, suggesting the presence of unidentified proteins involved in the tumor suppressive effects. Therefore, mass spectrometry was used to identify the proteins interacting with SEMA5A. Currently, only the positive control SEMA5A was identified in co-immunoprecipitation and mass spectrometry assays. More experiments are still needed to explore the proteins interacting with SEMA5A and involved in the tumor-suppressing pathways. In summary, this study identified that the of extracellular domain of SEMA5A could upregulate genes in interferon signaling pathways and result in the suppressive effects of lung cancer cells.

並列關鍵字

Lung Cancer ； Semaphorin5A ； Extracellular Domain ； Tumor suppressor

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