MapToCleave: High-throughput profiling of microRNA biogenesis in living cells
Wenjing Kang,
Bastian Fromm,
Anna J. Houben,
Eirik Høye,
Daniela Bezdan,
Carme Arnan,
Kim Thrane,
Michaela Asp,
Rory Johnson,
Inna Biryukova,
Marc R. Friedländer
Affiliations
Wenjing Kang
Science for Life Laboratory, Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden
Bastian Fromm
Science for Life Laboratory, Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden; The Arctic University Museum of Norway, UiT - The Arctic University of Norway, Tromsø, Norway
Anna J. Houben
Centre for Genomic Regulation (CRG), The Barcelona Institute for Science and Technology, Barcelona (BIST), Catalonia, Spain
Eirik Høye
Department of Tumor Biology, Oslo Institute for Cancer Research, Oslo University Hospital, Oslo, Norway
Daniela Bezdan
Centre for Genomic Regulation (CRG), The Barcelona Institute for Science and Technology, Barcelona (BIST), Catalonia, Spain; Institute of Medical Genetics and Applied Genomics, University of Tübingen, Tübingen, Germany; NGS Competence Center Tübingen (NCCT), University of Tübingen, Tübingen, Germany
Carme Arnan
Centre for Genomic Regulation (CRG), The Barcelona Institute for Science and Technology, Barcelona (BIST), Catalonia, Spain
Kim Thrane
Department of Gene Technology, School of Engineering Sciences in Chemistry, Biotechnology and Health, KTH Royal Institute of Technology, Science for Life Laboratory, Solna, Sweden
Michaela Asp
Department of Gene Technology, School of Engineering Sciences in Chemistry, Biotechnology and Health, KTH Royal Institute of Technology, Science for Life Laboratory, Solna, Sweden
Rory Johnson
Department of Medical Oncology, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland; Department for BioMedical Research, University of Bern, Bern, Switzerland; School of Biology and Environmental Science, University College Dublin, Dublin, Ireland; Conway Institute for Biomolecular and Biomedical Research, University College Dublin, Dublin, Ireland
Inna Biryukova
Science for Life Laboratory, Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden
Marc R. Friedländer
Science for Life Laboratory, Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden; Corresponding author
Summary: Previous large-scale studies have uncovered many features that determine the processing of microRNA (miRNA) precursors; however, they have been conducted in vitro. Here, we introduce MapToCleave, a method to simultaneously profile processing of thousands of distinct RNA structures in living cells. We find that miRNA precursors with a stable lower basal stem are more efficiently processed and also have higher expression in vivo in tissues from 20 animal species. We systematically compare the importance of known and novel sequence and structural features and test biogenesis of miRNA precursors from 10 animal and plant species in human cells. Lastly, we provide evidence that the GHG motif better predicts processing when defined as a structure rather than sequence motif, consistent with recent cryogenic electron microscopy (cryo-EM) studies. In summary, we apply a screening assay in living cells to reveal the importance of lower basal stem stability for miRNA processing and in vivo expression.
