Fig. 4: Frameshift mutation of RIG-I triggers circRIG-I generation.
From: Mutant RIG-I enhances cancer-related inflammation through activation of circRIG-I signaling
a RT-qPCR analysis of Ddx58 mRNA in virus-infected WT and Rig-ifs/fs MEFs (n = 3 cell cultures, mean ± s.e.m., *** P < 0.0001, two-tailed unpaired Student’s t-test). b-e RT-qPCR analysis of Ddx58 mRNA (b-c) and pre-mRNA (d-e) in indicated mice infected with VSV (n = 4 mice, mean ± s.e.m., ****P < 0.0001, **P = 0.001046 (c), *P = 0.0182, **P = 0.0058 (d), two-tailed unpaired Student’s t-test). f RNA secondary structures of RIG-I pre-mRNA (exon 3 and intron 3) were predicted by RNAfold web server. Frameshift mutation and pre-mRNA structural change were highlighted in red and blue, respectively. g Integrative Genomics Viewer analysis of CLIP-seq coverage, DHX9 peaks at Alu elements of Ddx58 were highlighted. h Interaction between DHX9 and WT or frameshift mutant RIG-I pre-mRNA. Data are collected from 2 independent experiments. i Schematic diagram illustrated the formation of circRIG-I (mmu-DDX58_0004) originated from Rig-i pre-mRNA (exon 5 and 12). Sequencing analysis of circRIG-I junction site was shown. Data are collected from 2 independent experiments. j PCR analysis of circRIG-I in WT and Rig-ifs/fs MEFs treated by RNase R or not with specific divergent primers. Data are collected from 2 independent experiments. k Northern blotting analysis of circRIG-I and RIG-I mRNA levels in MEFs by hybridization with exon 5 (upper) and exon 5-exon 12 junction (lower) probes with and without RNase R treatment. Data are collected from 2 independent experiments. l Subcellular localization of circRIG-I in iBMDM by fluorescence in situ hybridization. The scale bars represent 20 μm. Data are collected from 2 independent experiments. m RT-qPCR analysis of circRIG-I expression in indicated MEFs with VSV or LCMV Cl13 infection (n = 3 cell cultures, mean ± s.e.m., *P = 0.045, **P = 0.002226 (Control), **P = 0.002421 (VSV), two-tailed unpaired Student’s t-test). n RT-qPCR analysis of circRIG-I expression in indicated tissues from indicated mice infected with VSV (n = 5 mice, mean ± s.e.m., **P = 0.006939 (Liver), **P = 0.008278 (Lung), two-tailed unpaired Student’s t-test). Source data are provided as a Source Data file.
