NTR1 is required for transcription elongation checkpoints at alternative exons in Arabidopsis

• Published in: The EMBO journal 2015-02, Vol.34 (4), p.544-558
• Main Authors: Dolata, Jakub, Guo, Yanwu, Kołowerzo, Agnieszka, Smoliński, Dariusz, Brzyżek, Grzegorz, Jarmołowski, Artur, Świeżewski, Szymon
• Format: Article
• Language: English
• Subjects: alternative splicing; Arabidopsis - genetics; Arabidopsis - metabolism; Botany; elongation checkpoint; EMBO09; EMBO30; EMBO44; Exons - genetics; Genetics; Mutants; Mutation; Periplasmic Binding Proteins - genetics; Periplasmic Binding Proteins - metabolism; RNA polymerase; RNA Splice Sites - genetics; Site selection; transcription; Transcription factors; Transcription, Genetic - genetics
• ISSN: 0261-4189; 1460-2075
• DOI: 10.15252/embj.201489478

The interconnection between transcription and splicing is a subject of intense study. We report that Arabidopsis homologue of spliceosome disassembly factor NTR1 is required for correct expression and splicing of DOG1 , a regulator of seed dormancy. Global splicing analysis in atntr1 mutants revealed a bias for downstream 5′ and 3′ splice site selection and an enhanced rate of exon skipping. A local reduction in PolII occupancy at misspliced exons and introns in atntr1 mutants suggests that directionality in splice site selection is a manifestation of fast PolII elongation kinetics. In agreement with this model, we found AtNTR1 to bind target genes and co‐localise with PolII. A minigene analysis further confirmed that strong alternative splice sites constitute an AtNTR1‐dependent transcriptional roadblock. Plants deficient in PolII endonucleolytic cleavage showed opposite effects for splice site choice and PolII occupancy compared to atntr1 mutants, and inhibition of PolII elongation or endonucleolytic cleavage in atntr1 mutant resulted in partial reversal of splicing defects. We propose that AtNTR1 is part of a transcription elongation checkpoint at alternative exons in Arabidopsis . Synopsis AtNTR1, the Arabidopsis homologue of spliceosome disassembly factor NTR1, is required for pausing of RNA polymerase II at strong alternative splicing sites. AtNTR1 is required for proper splicing of seed dormancy regulator DOG1. AtNTR1‐deficient plants display enhanced RNA PolII elongation rate across strong alternative splice sites. AtNTR1 co‐localises with RNA PolII and associates to alternatively spliced target genes. Plants deficient in AtNTR1 and TFIIS show opposite directionality in splice site selection. Graphical Abstract AtNTR1, the Arabidopsis homologue of spliceosome disassembly factor NTR1, is required for pausing of RNA polymerase II at strong alternative splicing sites.