RNA-Seq of SMG1-SMG8-SMG9 complex alterations in human colorectal adenocarcinoma cell line HCT116
Nonsense-mediated mRNA decay (NMD) is a translation-dependent mRNA turnover pathway, which degrades transcripts containing premature termination codons. The execution of NMD requires the phosphorylation of N- and C-terminal tails of the key NMD factor UPF1, which thereby serve as binding platforms for the degradation factors SMG5, SMG6 and SMG7.  UPF1 phosphorylation is mediated by the kinase SMG1, whose activity is regulated by a heterodimer consisting of SMG8 and SMG9. Recent work indicated that SMG9 functions as a bridge between SMG1 and SMG8, allowing the C-terminus of SMG8 to elicits its role of stabilizing the autoinhibitory state of SMG1. Here, we deleted the C-terminus of endogenous SMG8 in human colorectal adenocarcinoma cell line HCT116 via CRISPR-Cas9. In addition, we established SMG8- and SMG9-depleted cells. With these cell lines we wanted to explore the regulatory role of SMG8 and SMG9 for NMD execution. Furthermore, we tested the transcriptomic changes upon treatment of cells with the SMG1 inhibitor SMG1i, which functions as an ATP-competitive inhibitor and binds to the active site of SMG1. Cells were treated with 0, 0.1 or 1 µM SMG1i for 24 h. As controls, the HCT116 wildtype cells were treated with DMSO.