胰腺癌细胞中 AP-2α 剪接变异体的异常表达。
Aberrant expressions of AP-2α splice variants in pancreatic cancer.
机构信息
Department of Medicine, Dartmouth Medical School, Hanover, NH, USA.
出版信息
Pancreas. 2011 Jul;40(5):695-700. doi: 10.1097/MPA.0b013e31821f2715.
OBJECTIVES
The present study was conducted to evaluate the expression and function of AP-2α isoforms in pancreatic ductal adenocarcinoma.
METHODS
The expression of AP-2α was evaluated at the RNA level by reverse transcription-polymerase chain reaction and at the protein level by Western blotting and immunofluorescence. Its function as a transcription factor was evaluated in transient transfection experiments: DNA binding properties by electromobility shift assay and transactivation capabilities by luciferase assay.
RESULTS
Multiple alternative splicing events of AP-2α messenger occurred in all human pancreatic cancer cell lines, including a novel isoform, termed variant 6, which was not present in HeLa cells. At the protein level, except for 1 cell line, all pancreatic cancer cell lines expressed high nuclear levels of AP-2α. We also showed that AP-2α expressed by the pancreatic cancer cell lines could bind its cognate recognition site and activate transcription. However, variant 6, although not able to activate transcription, did not act in a dominant negative manner when cotransfected with the full-length protein.
CONCLUSIONS
Multiple isoforms of AP-2α are highly expressed in pancreatic cancer cell lines including a new isoform, AP-2α variant 6, which seems to be pancreatic cancer specific and is deprived of transcriptional activity.
目的
本研究旨在评估 AP-2α 异构体在胰腺导管腺癌中的表达和功能。
方法
通过逆转录-聚合酶链反应（RT-PCR）评估 AP-2α 在 RNA 水平的表达，通过 Western blot 和免疫荧光评估其在蛋白质水平的表达。通过瞬时转染实验评估其作为转录因子的功能：通过电泳迁移率变动分析评估 DNA 结合特性，通过荧光素酶测定评估转录激活能力。
结果
所有人类胰腺癌细胞系中均发生了 AP-2α 信使的多种选择性剪接事件，包括一种新的异构体，称为变体 6，其不存在于 HeLa 细胞中。在蛋白质水平上，除了 1 种细胞系外，所有胰腺癌细胞系均表达高水平的核 AP-2α。我们还表明，胰腺癌细胞系表达的 AP-2α 能够与其同源识别位点结合并激活转录。然而，变体 6 虽然不能激活转录，但在与全长蛋白共转染时，并不表现出显性负性作用。
结论
多种 AP-2α 异构体在胰腺癌细胞系中高度表达，包括一种新的异构体 AP-2α 变体 6，其似乎是胰腺癌细胞特异性的，并且缺乏转录活性。
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