Journal Article

CLK-dependent exon recognition and conjoined gene formation revealed with a novel small molecule inhibitor

Iwatani, Misa, Morin, Gregg B., Sano, Masaya, Ogasawara-Shimizu, Mari, Araki, Shinsuke, Cheng, S.-W. Grace, Wan, Adrian, Morishita, Daisuke, Nakao, Shoichi, Aparicio, Samuel, Shumansky, Karey, Uchiyama, Noriko, Tasaki, Shinya, Sasaki, Satoshi, Rosner, Jamie, Oloumi, Arusha, Tozaki, Hirokazu, Hughes, Christopher S., Morishita, Nao, Nomura, Toshiyuki, Ohashi, Tomohiro, Miyazaki, Tohru, Kawamoto, Tomohiro, Kong, Esther, Ohori, Momoko, Nakayama, Yusuke, Nakanishi, Osamu, Karashima, Masatoshi, Funnell, Tyler, Yap, Damian, Hara, Ryujiro, Murai, Aiko, McKinney, Steven, Shah, Sohrab, Nakanishi, Atsushi, Toyoshiba, Hiroyoshi

2017

Overview

CDC-like kinase phosphorylation of serine/arginine-rich proteins is central to RNA splicing reactions. Yet, the genomic network of CDC-like kinase-dependent RNA processing events remains poorly defined. Here, we explore the connectivity of genomic CDC-like kinase splicing functions by applying graduated, short-exposure, pharmacological CDC-like kinase inhibition using a novel small molecule (T3) with very high potency, selectivity, and cell-based stability. Using RNA-Seq, we define CDC-like kinase-responsive alternative splicing events, the large majority of which monotonically increase or decrease with increasing CDC-like kinase inhibition. We show that distinct RNA-binding motifs are associated with T3 response in skipped exons. Unexpectedly, we observe dose-dependent conjoined gene transcription, which is associated with motif enrichment in the last and second exons of upstream and downstream partners, respectively. siRNA knockdown of CLK2-associated genes significantly increases conjoined gene formation. Collectively, our results reveal an unexpected role for CDC-like kinase in conjoined gene formation, via regulation of 3′-end processing and associated splicing factors. The phosphorylation of serine/arginine-rich proteins by CDC-like kinase is a central regulatory mechanism for RNA splicing reactions. Here, the authors synthesize a novel small molecule CLK inhibitor and map CLK-responsive alternative splicing events and discover an effect on conjoined gene transcription.

• Publisher: Nature Publishing Group UK,Nature Publishing Group,Nature Portfolio
• Subject: 631/337/1645/1792 / 631/337/2019 / 631/92/500 / 631/92/613 / Alternative splicing / Alternative Splicing - drug effects / Arginine / Chemical synthesis / Exons / Gene Expression Profiling / Genome, Human / HCT116 Cells / Humanities and Social Sciences / Humans / Imidazoles - chemical synthesis / Imidazoles - pharmacology / Inhibitors / Kinases / multidisciplinary / Phosphorylation / Phosphorylation - drug effects / Protein Kinase Inhibitors - chemical synthesis / Protein Kinase Inhibitors - pharmacology / Protein-Serine-Threonine Kinases - antagonists & inhibitors / Protein-Serine-Threonine Kinases - genetics / Protein-Serine-Threonine Kinases - metabolism / Protein-Tyrosine Kinases - antagonists & inhibitors / Protein-Tyrosine Kinases - genetics / Protein-Tyrosine Kinases - metabolism / Proteins / Pyrimidines - chemical synthesis / Pyrimidines - pharmacology / Ribonucleic acid / RNA / RNA, Messenger - antagonists & inhibitors / RNA, Messenger - genetics / RNA, Messenger - metabolism / RNA, Small Interfering - genetics / RNA, Small Interfering - metabolism / RNA-Binding Proteins - antagonists & inhibitors / RNA-Binding Proteins - genetics / RNA-Binding Proteins - metabolism / Science / Science (multidisciplinary) / Serine / Splicing / Structure-Activity Relationship / Transcription / Transcription, Genetic