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標題: 新肺癌轉移及腫瘤抑制基因,HLJ1,之功能定性
HLJ1 is a novel invasion and tumor suppressor
作者: 陳健尉
關鍵字: 基礎醫學類, 醫學技術, 生物技術, 臨床醫學類;NSCLC;非小細胞肺癌;heat shock protein HLJ1;invasion;yeast two-hybrid;RNA interference;microarray;熱休克蛋白HLJ1;侵入作用;酵母菌雙雜合法;RNA干擾技術;基因微陣列;基礎研究
Lung cancer, predominantly non-small-cell lung cancer (NSCLC), is the most common cause of cancer deaths worldwide. In the study of last two years, we have demonstrated that heat shock protein HLJ1 is a novel tumor/metastasis suppressor, as well as a significantly prognostic predictor of recurrence and overall survival in NSCLC patients. We also identified a novel mechanism that up-regulates tumor suppressor HLJ1 expression by enhancer AP-1 binding with promoter YY1 and p300 through DNA bending and multi-protein complex formation, which further inhibits in vitro cancer cell invasion. However, the global understanding of HLJ1 functions and regulatory mechanisms are still unclear. To further disclose the true face of HLJ1, in the project of this fiscal year, we will characterize the interaction partners of HLJ1 protein and the distribution of HLJ1 and its associated proteins in lung cancer cell by immuno-precipitation and yeast two-hybrid approaches. In addition, we will employ RNA interference and microarray technologies to identify the downstream genes of HLJ1, which are then subjected to the analysis of comprehensive bioinformatics tool we developed (CRSD, All these efforts may assist us in exploring the putative mechanisms of HLJ1 involved in tumor/metastasis suppression and orchestrating the process of cancer metastasis. In the preliminary results, we have constructed all the expression vector needed in this project and identified 10 HLJ1-silenced clones and 3 scramble cell controls, and subsequently were subjected to cellular functions. The results revealed that knockdown of HLJ1 expression could promote cancer cell invasion and migration. In addition, the microarray results showed that several pathways are affected by HLJ1 silence, including MAPK signaling, focal adhesion, regulation of actin cytoskeleton, Jak-STAT signaling, Wnt, and EMT pathways. Among these pathways, we picked up 43 genes to verify their expression patterns by using real-time PCR. The results revealed that 34 genes are coincident with biochip, and 9 genes are no difference or opposite. Further characterization of genes identified in this study is currently in progress.

肺癌,主要是非小細胞肺癌,為當今世上最常見的致死癌症之ㄧ。在前兩年的研究中,我們發現熱休克蛋白HLJ1是一個新的腫瘤及轉移抑制基因,表現HLJ1可降低肺癌的復發率並延長病人的存活率,是疾病復發及存活預後的重要預測因子。此外,也發現其促進子上的轉錄因子AP-1與啟動子上的轉錄因子YY1會經由DNA彎曲的機制(DNA bending),並藉由P300蛋白質的媒介,而形成蛋白質複合物,以進一步共同調節HLJ1的表現並抑制肺癌細胞的侵襲能力。然而,其完整的功能及調控機制仍不清楚。為進一步揭露此基因的詳細面貌,在今年度的計畫中,將進一步以免疫共沉澱法及酵母菌雙雜合法來篩選與HLJ1進行交互作用的蛋白分子,及其在細胞內產生作用的區域;此外,也將利用RNA干擾及微陣列技術研究其下游基因的表現情形,並以我們所發展出來的廣泛型生物資訊工具進行分析(CRSD,,以建構出HLJ1基因參與腫瘤及癌轉移抑制的可能機制。目前已構築所有需使用的基因表現載體,並已獲得10株HLJ1靜默細胞株及3株對照組細胞株(scramble control)。將上述靜默HLJ1表現的細胞株進行試管內侵入能力(in vitro invasion assay)及移動能力(migration)分析,結果顯示降低HLJ1表現量會增加細胞的侵入能力及移動能力。以微陣列分析HLJ1基因靜默後細胞的基因表現,結果顯示與MAPK signaling、Focal adhesion、Regulation of actin cytoskeleton、Jak-STAT signaling、Wnt及EMT等生物途徑有關。在這些路徑中,篩選出43個差異表現基因,利用及時定量聚合酶連鎖反應偵測基因表現趨勢是否和晶片一致,結果有34個基因趨勢與晶片一致,9個基因無差異或是相反。後續進一步的分析工作正在進行中。
其他識別: DOH97-TD-G-111-008
Appears in Collections:生命科學系所

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