RNA Sequencing Analysis of MC38 tumors with Glutamate decarboxylase 1 Knockdown

Various types of cancers, including colorectal cancer, have shown an unusual dependence on glutamine. Glutamine can undergo an alternate fate in neurons: conversion into a non-proteinogenic amino acid gamma-aminobutyric acid (GABA) by the glutamic acid decarboxylase (GAD), including two family members GAD1 and GAD2. Some studies have shown that GAD1 expression is dysregulated in certain cancer types, but the effect of GAD1 on the tumorigenic process remain largely unknown. Thus, we use mouse colon adenocarcinoma MC38 tumor samples to perform RNA-sequencing (RNA-seq) assay. The goal of this study is to investigate GAD1-mediated transcriptome changes within tumors. Overall design: MC38 cells stably expressing GAD1 shRNAs (GAD1-shA and GAD1-shB) or non-targeting shRNA control (NTC) were subcutaneously injected into C57BL/6 mice. All the tumors were extracted 20 days after implantation. Total RNA was extracted from GAD1-shA GAD1-shB and NTC MC38 tumor samples (3 replicates per group) using an RNeasy kit (Qiagen) and dissolved in RNase-free water. RNA samples were sequenced on Illumina novaseq.