Alexander Hirth1,2,10, Edoardo Fatti1,2,10,11, Eugen Netz3,4,10, Sergio P. Acebron1,12, Dimitris Papageorgiou5,6, Andrea Švorinić1,2,7,13, Cristina-Maria Cruciat1,14, Emil Karaulanov8, Alexandr Gopanenko8, Tianheng Zhu1,2, Irmgard Sinning7, Jeroen Krijgsveld5,6, Oliver Kohlbacher3,4,9 and Christof Niehrs1,8 1Division of Molecular Embryology, DKFZ-ZMBH Alliance, Deutsches Krebsforschungszentrum (DKFZ), 69120 Heidelberg, Germany; 2Faculty of Biosciences, Ruprecht-Karls University of Heidelberg, 69120 Heidelberg, Germany; 3Applied Bioinformatics, Department of Computer Science, University of Tübingen, 72076 Tübingen, Germany; 4Institute for Bioinformatics and Medical Informatics, University of Tübingen, 72076 Tübingen, Germany; 5Proteomics of Stem Cells and Cancer, German Cancer Research Center (DKFZ), 69120 Heidelberg, Germany; 6Medical Faculty, Heidelberg University, 69120 Heidelberg, Germany; 7Heidelberg University Biochemistry Center (BZH), 69120 Heidelberg, Germany; 8Institute of Molecular Biology (IMB), 55128 Mainz, Germany; 9Translational Bioinformatics, University Hospital Tübingen, 72076 Tübingen, Germany

↵10 These authors contributed equally to this work.

↵Present addresses: 11Department of Biology, Institute of Biochemistry, ETH (Eidgenössische Technische Hochschule) Zürich, 8093 Zürich, Switzerland; 12Centre for Organismal Studies (COS), Heidelberg University, 69120 Heidelberg, Germany; 13Department of Molecular and Cellular Biology, University of Geneva, 1211 Geneva, Switzerland; 14Hochschule Esslingen, University of Applied Sciences, Department of Biotechnology, 73728 Esslingen, Germany

Corresponding author: niehrsdkfz-heidelberg.de Abstract

DEAD box (DDX) RNA helicases are a large family of ATPases, many of which have unknown functions. There is emerging evidence that besides their role in RNA biology, DDX proteins may stimulate protein kinases. To investigate if protein kinase–DDX interaction is a more widespread phenomenon, we conducted three orthogonal large-scale screens, including proteomics analysis with 32 RNA helicases, protein array profiling, and kinome-wide in vitro kinase assays. We retrieved Ser/Thr protein kinases as prominent interactors of RNA helicases and report hundreds of binary interactions. We identified members of ten protein kinase families, which bind to, and are stimulated by, DDX proteins, including CDK, CK1, CK2, DYRK, MARK, NEK, PRKC, SRPK, STE7/MAP2K, and STE20/PAK family members. We identified MARK1 in all screens and validated that DDX proteins accelerate the MARK1 catalytic rate. These findings indicate pervasive interactions between protein kinases and DEAD box RNA helicases, and provide a rich resource to explore their regulatory relationships.

Received July 10, 2023. Accepted June 12, 2024.