Functional Dissection of the Pol V Largest Subunit CTD in RNA-Directed DNA Methylation

Publication date: 27 June 2017
Source:Cell Reports, Volume 19, Issue 13
Author(s): Jered M. Wendte, Jeremy R. Haag, Jasleen Singh, Anastasia McKinlay, Olga M. Pontes, Craig S. Pikaard
Plant multisubunit RNA polymerase V (Pol V) transcription recruits Argonaute-small interfering RNA (siRNA) complexes that specify sites of RNA-directed DNA methylation (RdDM) for gene silencing. Pol V's largest subunit, NRPE1, evolved from the largest subunit of Pol II but has a distinctive C-terminal domain (CTD). We show that the Pol V CTD is dispensable for catalytic activity in vitro yet essential in vivo. One CTD subdomain (DeCL) is required for Pol V function at virtually all loci. Other CTD subdomains have locus-specific effects. In a yeast two-hybrid screen, the 3′→ 5′ exoribonuclease RRP6L1 was identified as an interactor with the DeCL and glutamine-serine (QS)-rich subdomains located downstream of an Argonaute-binding subdomain. Experimental evidence indicates that RRP6L1 trims the 3′ ends of Pol V transcripts sliced by Argonaute 4 (AGO4), suggesting a model whereby the CTD enables the spatial and temporal coordination of AGO4 and RRP6L1 RNA processing activities.

Graphical abstract

Teaser

In plants, transcription by RNA polymerase V (Pol V) specifies sites of RNA-directed DNA methylation (RdDM) and gene silencing. Wendte et al. define locus-specific functions for subdomains of the Pol V largest subunit's C-terminal domain (CTD). The exonuclease RRP6L1 is shown to interact with the CTD and trim the 3′ ends of Pol V transcripts.


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