SRC抑制物對β3GnT家族的調控研究

Preliminary studies of β3GnT families expression regulated by a SRC inhibitor

劉軒辰(Hsuan-Chen Liu)

指導教授 ： 華國媛

國立臺北科技大學/工程學院/有機高分子研究所/碩士(2012年)

摘要

β1,3-N-acetylglucosaminyltransferase(β3GnT)蛋白質家族的功能是將GlcNAc的醣基以β1→3的鍵結方式與細胞中特定的醣鏈結合。在醣化作用中，β3GnT蛋白扮演著重要的角色，β3GnT的基因也廣泛的表現於人類以及老鼠的組織細胞中。目前已知β3GnT 可參與poly-N-acetyllactosamine聚醣結構的合成，而這個結構已被指出與癌症轉移有關。此外，SRC是一個已知的致癌基因，此基因與調節細胞的增生、腫瘤的形成、侵入和轉移有關。因此，本篇論文研究的目標是了解是否SRC會調控老鼠和人的β3GnT。首先，我們研究一株穩定轉殖v-Src基因的老鼠細胞，我們發現在過度表現Src基因的老鼠細胞中其poly-N-acetyllactosamine表現量與正常的NIH/3T3老鼠細胞相較之下有增加的現象；當加入SRC抑制物bosutinib時，正常的NIH/3T3老鼠細胞以及過度表現SRC的老鼠細胞其poly-N-acetyllactosamine表現量皆下降。為了更深入了解SRC在poly-N-acetyllactosamine和β3GnT基因所扮演的調控角色，我們在人類結腸癌細胞(SW620)以及肺癌細胞(NCI-H23)加入了SRC 抑制物(bosutinib)後，觀察細胞中β3GnT表現量變化。由即時定量分析結果顯示，當癌細胞加入bosutinib後，NCI-H23細胞株的β3GnT2、β3GnT3、β3GnT4、β3GnT5、β3GnT6、β3GnT8以及SW620細胞株的β3GnT3、β3GnT6、β3GnT8的基因表現量會受到影響。此外，由流式細胞儀的分析顯示，細胞表面上的poly-N-acetyllactosamine表現量會因SRC抑制物的加入而改變，此結果與β3GnT2和β3GnT8基因表現量減少有一致性。總結來說，我們發現SRC至少對於β3GnT2和β3GnT8有調控關係因而導致poly-N-acetyllactosamine表現的改變。

關鍵字

poly-N-acetyllactosamine ； β1,3-N-acetylglucosamonyltransferase ； Src

並列摘要

The β1,3-N-acetylglucosaminyltransferase(β3GnT) protein family is responsible for the synthesis of β1,3-Linked GlcNAc residues to different specific types of glycoconjugates of cells. The β3GnT protein family is one of the important enzyme family in glycosylation, and the β3GnT genes were widely expressed in human and mouse tissues. Some members of the protein family are involved in the biosynthesis of poly-N-acetyllactosamine chains, which has been implicated for their metastatic potential in tumor cells. In addition, SRC is a well-known oncogene, which has been implicated in pathways regulating cell proliferation, angiogenesis, invasion and metastasis. Therefore, the goal of my thesis is to study if mouse/and human β3GnT genes is under the control of SRC. First, we had used a stable mouse cell line expressing v-Src, we found SRC overexpressed mouse cell line displayed the increase of poly-N-acetyllactosamine in comparing with the normal NIH/3T3 cell line. To further study the regulatory role of SRC on the poly-N-acetyllactosamine and β3GnT genes, we had studied the expressions of β3GnTs in human colon cancer SW620 cells and lung cancer NCI-H23 cells by treating the cells with bosutinib, a SRC inhibitor. The real-time qPCR results showed that when cancer cell line was treated with bosutinib affected the expression of the transcripts of the β3GnT2, β3GnT3, β3GnT4, β3GnT5, β3GnT6 and β3GnT8 in NCI-H23, and the β3GnT3, β3GnT6 and β3GnT8 in SW620. Furthermore, the FACS analysis revealed that the surface expression of poly-N-acetyllactosamine was altered by the SRC inhibitor, which was consistent with the results from the gene expression of β3GnT2 and β3GnT8. As a summary, we had demonstrated at least β3GnT2 and β3GnT8 were under the control of SRC pathway, which resulted in alteration of expression of poly-N-acetyllactosamine.

並列關鍵字

poly-N-acetyllactosamine ； β1,3-N-acetylglucosamonyltransferase ； Src

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