RNA sequencing to identify differentially-expressed genes in HER3-depleted KatoII MET-amplified cancer cells

Receptor tyrosine kinases (RTKs) are recognized as targets of precision medicine in human cancer upon their gene amplification or constitutive activation, resulting in increased downstream signal complexity including heterotypic crosstalk with other RTKs. The Met hepatocyte growth factor RTK exhibits such reciprocal crosstalk with several members of the human EGFR (HER) family of RTKs when amplified in cancer cells. We observed that Met signaling converges on HER3 tyrosine phosphorylation across a panel of seven MET-amplified cancer cell lines and that HER3 is required for cancer cell expansion and oncogenic capacity in vitro and in vivo. In order to identify genes required for cancer cell proliferation in the MET-amplified setting whose expression, we stably transduced KatoII MET-amplified cells with shRNA targeting ERBB3, the gene encoding HER3. Stably-transduced cells were selected with puromycin, assayed for proliferative phenotype, and lysed for RNA extraction. Pooled libraries of total RNA were then sequenced using the Illumina HiSeq4000 platform, and differentially-expressed genes were identified from read counts normalized across samples. Differentially-expressed genes were further assayed for dependence on HER3 in other MET-amplified cell lines to identify genes recurrently dependent on HER3 in the MET-amplified setting. Overall design: MET-amplified KatoII cells were lentivirally transduced with shRNA targeting the human ERBB3 gene. Stably-transduced cells were selected with puromycin, assayed for proliferation phenotype and lysed in parallel. RNA sequencing of total RNA reads were mapped to genes, normalized across samples and assessed for differential gene expression.

搜集汇总

背景与挑战

背景概述

该数据集是一个RNA测序研究，包含9个人类样本的配对端测序数据，使用Illumina HiSeq 4000平台生成，旨在分析HER3缺失的KatoII MET扩增癌细胞中的差异表达基因。数据以fastq格式提供，每个样本的测序量在2.6Gb至3.6Gb之间，总计约8.6G至8.7G碱基，适用于基因表达谱和生物信息学分析。