Determine the effects of QKI7 on translation efficiency via ribosome profiling (Ribo-seq)

N7-methylguanosine (m7G) modification, routinely occurring at the 5’ cap of mRNA or within tRNA and rRNA, also exists internally in mRNA. Although essential for mRNA translation as well as stress response, the “reader” protein for mRNA internal m7G modification is still unrevealed. Here, we reported that Quaking protein (QKI), especially QKI7, can selectively recognize the internal mRNA m7G decoration in the cytosol of various cell types. We identified over 1000 confident m7G-modified and QKI binding RNA targets with a conserved motif, “GANGAN (N=A/U/G)”. More strikingly, internal m7G reader QKI7 directly interacts with the SG core protein G3BP1 and can shuttle a subset of m7G-modified transcripts into SG mRNA pool under oxidative stress condition. Additionally, by sequestering mRNA within SGs, QKI7 modulates the translation efficiency of selected transcripts. Moreover, in line with the observation that doxorubicin triggers the assembly of SGs, QKI7 mediates the sensitivity of cancer cells to chemotherapy drug treatment. Two 15-cm dishes empty vector or QKI7 overexpression U2OS cells were treated with 0.5mM NaAsO2 for 1 hour before harvesting the cells. The samples were subjected to ribosome profiling and both the input RNA and footprint RNAs were used for sequencing.