Mature microRNA-binding protein QKI promotes microRNA-mediated gene silencing

ABSTRACT

Although Argonaute (AGO) proteins have been the focus of microRNA (miRNA) studies, we observed AGO-free mature miRNAs directly interacting with RNA-binding proteins, implying the sophisticated nature of fine-tuning gene regulation by miRNAs. To investigate microRNA-binding proteins (miRBPs) globally, we analyzed PAR-CLIP data sets to identify RBP quaking (QKI) as a novel miRBP for let-7b. Potential existence of AGO-free miRNAs were further verified by measuring miRNA levels in genetically engineered AGO-depleted human and mouse cells. We have shown that QKI regulates miRNA-mediated gene silencing at multiple steps, and collectively serves as an auxiliary factor empowering AGO2/let-7b-mediated gene silencing. Depletion of QKI decreases interaction of AGO2 with let-7b and target mRNA, consequently controlling target mRNA decay. This finding indicates that QKI is a complementary factor in miRNA-mediated mRNA decay. QKI, however, also suppresses the dissociation of let-7b from AGO2, and slows the assembly of AGO2/miRNA/target mRNA complexes at the single-molecule level. We also revealed that QKI overexpression suppresses cMYC expression at post-transcriptional level, and decreases proliferation and migration of HeLa cells, demonstrating that QKI is a tumour suppressor gene by in part augmenting let-7b activity. Our data show that QKI is a new type of RBP implicated in the versatile regulation of miRNA-mediated gene silencing.

KEYWORDS:

• QKI
• AGO2
• RNA-binding protein
• let-7b
• Tumour suppressor gene

Acknowledgments

We thank D. Patel (Memorial Sloan Kettering Cancer center, NY, USA) and MJ Gamble (Albert Einstein College of Medicine, NY, USA) for providing plasmids. We appreciate GM Wilson (University of Maryland School of Medicine, MD, USA) for providing critical comments. We appreciate Drs. David Corey and Joshua Mendell for providing us AGO TKO cells. We thank Dr. Narry Kim for kindly providing us AGO2 KO cells. This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Ministry of Science and ICT (NRF-2022R1A2C4001528). J.W.L., M.J.S., and K.M were supported by NRF (NRF-2022R1A2C4001528). H.B.P was supported by NRF (NRF-2022R1A2C1092803) and 2023 New Professor Support Program of Natural Science Research Institute in Gangneung-Wonju National University. S.K., J.-H.C., K.-W.M., H.M., and J.-H.Y. were supported by the Medical University of South Carolina Startup fund. S.K. and J.-H.Y. were supported by R01AA027532. H.M. and J.-H.Y. were supported by the University of Oklahoma Startup fund. A.P. and M.H. were supported by NIH IRP. D.K. and S.-U.K were supported by a grant from the NIH/NINDS R01NS123456. M.H.J., M.S. and S.H. were supported by Creative Research Initiatives (2009-0081562) of the National Research Foundation of Korea

Disclosure statement

No potential conflict of interest was reported by the author(s).

Data availability statement

The authors confirm that the data supporting the findings of this study are available within the article and its supplementary materials.

Supplementary material

Supplemental data for this article can be accessed online at https://doi.org/10.1080/15476286.2024.2314846.