Identification of signaling pathways modulated by kiaa0101 in HepG2 cells. Homo sapiens

Kiaa0101 (p15PAF) was previously observed overexpressed in hepatocelluler carcinoma tissues and promoted tumorigenesis. In order to identify potential biological pathways modulated by Kiaa0101, a lentivirus vector with Kiaa0101 was constructed and transfected into an HepG2 cell line to obtain a stable cell line with high expression of Kiaa0101. Interestingly, all the protein levels of P21, phosphorylated P21, Akt and phosphorylated Akt were induced. Further, gene expression profiles of Kiaa0101 overexpressed hepatocelluler cancer cells were analyzed using the DNA microarray technique. Differentially expressed genes (DEGs) were screened and followed by Gene Ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. 1361 genes and 942 genes were upregulated and downregulated, respectively. The most represented enriched GO biological process, cellular component and molecular function categories included DNA-dependent regulation of transcripton, plasma membrane and DNA binding, respectively. KEGG pathway analysis showed that the most significantly expressed genes were linked to pathways in cancer, MAPK signaling pathway and Wnt signaling pathway. Our study provides a comparative transcriptome analysis and revealed the downstream genes and signaling pathways that are possibly involved in Kiaa0101-induced hepatocelluler carcinoma. Overall design: Three kiaa0101-overexpressing stable cells and three control cells were chosen for microarray analysis. Subsequently, the RNAs were analyzed using microarray chips (Agilent Technologies) according to the manufacturer's protocol.Gene Ontology (GO) and KEGG pathways were performed to investigate the involved pathways

Kiaa0101（p15PAF）此前已被证实可在肝细胞癌（hepatocellular carcinoma）组织中过表达，并可促进肿瘤发生。为鉴定Kiaa0101调控的潜在生物学通路，研究人员构建了携带Kiaa0101的慢病毒载体（lentivirus vector），并转染至HepG2细胞系，以获得稳定高表达Kiaa0101的细胞株。有趣的是，P21、磷酸化P21、Akt及磷酸化Akt的蛋白水平均被诱导上调。进一步通过DNA微阵列（DNA microarray）技术分析Kiaa0101过表达肝癌细胞的基因表达谱，筛选得到差异表达基因（differentially expressed genes, DEGs）后，开展基因本体（Gene Ontology, GO）功能富集分析与京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes, KEGG）通路富集分析。结果显示，共有1361个基因上调、942个基因下调。富集程度最高的GO类别分别为：生物学过程大类为转录的DNA依赖性调控，细胞组分大类为质膜，分子功能大类为DNA结合。KEGG通路富集分析显示，差异表达基因最显著富集于癌症通路、丝裂原活化蛋白激酶（mitogen-activated protein kinase, MAPK）信号通路与Wnt信号通路。本研究通过比较转录组分析，揭示了可能参与Kiaa0101诱导肝细胞癌发生的下游基因与信号通路。

实验设计：选取3株Kiaa0101过表达稳定细胞株与3株对照细胞株进行微阵列分析。随后，按照制造商的操作流程，使用安捷伦科技（Agilent Technologies）的微阵列芯片对RNA进行检测，并通过GO与KEGG通路分析探究其参与的调控通路。