The NRPD1 N-terminus contains a Pol IV-specific motif that is critical for genome surveillance in Arabidopsis

L. Ferrafiat (1) , D. Pflieger (1) , Jasleen Singh (2) , M. Thieme (1) , M. Böhrer (1) , C. Himber (1) , A. Gerbaud (1) , E. Bucher (3) , Craig S. Pikaard (2) , T. Blevins (1)

1 IBMP - Institut de Biologie Moléculaire des Plantes
2 Indiana University [Bloomington]
3 Botanisches Institut

RNA-guided surveillance systems constrain the activity of transposable elements (TEs) in host genomes. In plants, RNA polymerase IV (Pol IV) transcribes TEs into primary transcripts from which RDR2 synthesizes double-stranded RNA precursors for small interfering RNAs (siRNAs) that guide TE methylation and silencing. How the core subunits of Pol IV, homologs of RNA polymerase II subunits, diverged to support siRNA biogenesis in a TE-rich, repressive chromatin context is not well understood. Here we studied the N-terminus of Pol IV’s largest subunit, NRPD1. Arabidopsis lines harboring missense mutations in this N-terminus produce wild-type (WT) levels of NRPD1, which co-purifies with other Pol IV subunits and RDR2. Our in vitro transcription and genomic analyses reveal that the NRPD1 N-terminus is critical for robust Pol IV-dependent transcription, siRNA production and DNA methylation. However, residual RNA-directed DNA methylation observed in one mutant genotype indicates that Pol IV can operate uncoupled from the high siRNA levels typically observed in WT plants. This mutation disrupts a motif uniquely conserved in Pol IV, crippling the enzyme's ability to inhibit retrotransposon mobilization. We propose that the NRPD1 N-terminus motif evolved to regulate Pol IV function in genome surveillance.

Domaines: Botanique; Génomique, Transcriptomique et Protéomique [q-bio.GN]

L. Ferrafiat, D. Pflieger, Jasleen Singh, M. Thieme, M. Böhrer, et al.. The NRPD1 N-terminus contains a Pol IV-specific motif that is critical for genome surveillance in Arabidopsis. Nucleic Acids Research, 2019, 47 (17), pp.9037-9052. ⟨10.1093/nar/gkz618⟩. ⟨hal-02262367⟩