Ancient Origin and Recent Innovations of RNA Polymerase IV and V

Abstract

Small RNA-mediated chromatin modification is a conserved feature of eukaryotes. In flowering plants, the short interfering (si)RNAs that direct transcriptional silencing are abundant and subfunctionalization has led to specialized machinery responsible for synthesis and action of these small RNAs. In particular, plants possess polymerase (Pol) IV and Pol V, multi-subunit homologs of the canonical DNA-dependent RNA Pol II, as well as specialized members of the RNA-dependent RNA Polymerase (RDR), Dicer-like (DCL), and Argonaute (AGO) families. Together these enzymes are required for production and activity of Pol IV-dependent (p4-)siRNAs, which trigger RNA-directed DNA methylation (RdDM) at homologous sequences. p4-siRNAs accumulate highly in developing endosperm, a specialized tissue found only in flowering plants, and are rare in nonflowering plants, suggesting that the evolution of flowers might coincide with the emergence of specialized RdDM machinery. Through comprehensive identification of RdDM genes from species representing the breadth of the land plant phylogeny, we describe the ancient origin of Pol IV and Pol V, suggesting that a nearly complete and functional RdDM pathway could have existed in the earliest land plants. We also uncover innovations in these enzymes that are coincident with the emergence of seed plants and flowering plants, and recent duplications that might indicate additional subfunctionalization. Phylogenetic analysis reveals rapid evolution of Pol IV and Pol V subunits relative to their Pol II counterparts and suggests that duplicates were retained and subfunctionalized through Escape from Adaptive Conflict. Evolution within the carboxy-terminal domain of the Pol V largest subunit is particularly striking, where illegitimate recombination facilitated extreme sequence divergence.

Trans-Homolog Interactions Facilitating Paramutation in Maize

“…Arabidopsis has only one functional second largest subunit shared by both Pol IV and Pol V, but maize and other grasses have three (Haag et al, 2014;Huang et al, 2015). Consistent with genetic data showing diverse functions of more than one Pol IV holoenzyme , recent proteomic profiles support the presence of at least two Pol IV (Pol IVa and Pol IVb) and three Pol V (Pol Va, Pol Vb, and Pol Vc) subtypes in maize callus ( Fig.…”

Comprehensive Annotation of Physcomitrella patens Small RNA Loci Reveals That the Heterochromatic Short Interfering RNA Pathway Is Largely Conserved in Land Plants

“…The Selaginella genome contains DCL3, RDR2, AGO4, and Pol IV/V largest subunit homologs (Banks et al, 2011), suggesting that the absence of 24-nucleotide RNAs in initial small RNA-seq libraries may be due to tissue-restricted expression. Phylogenetic analysis clearly identifies DCL3, AGO4, and Pol V-related homologs in basal plants (Huang et al, 2015). Finally, our previous analysis demonstrated that the P. patens DCL3 homolog is required for the accumulation of 22-, 23-, and 24-nucleotide RNAs from a handful of siRNA hot spots (Cho et al, 2008).…”

“…Dicers emerged early in the eukaryotic lineage and independently diverged in plants and animals (Mukherjee et al, 2013). Plants contain four ancient clades of DCL genes, with members in each clade being sub-functionalized for different types of small RNAs (Margis et al, 2006;Huang et al, 2015). P. patens has been reported to have no members of the DCL2 clade, single members of both the DCL3 and DCL4 clades, and two members of the DCL1 clade (Pp DCL1a and Pp DCL1b) (Axtell et al, 2007).…”

“…Previous analyses collectively indicated that P. patens has one homolog of the Arabidopsis Pol IV largest subunit (Pp NRPD1) and two homologs of the Pol V largest subunit (Pp NPRE1a and Pp NRPE1b) (Arif et al, 2013;Huang et al, 2015). A high density of multiple GW/WG/GWG motifs in the C-terminal domain is a characteristic of the Pol V, but not the Pol IV largest subunit (Haag and Pikaard, 2011).…”

Revisiting Criteria for Plant MicroRNA Annotation in the Era of Big Data

“…These observations led to the suggestion that, unlike phasiRNAs and miRNAs, hc-siRNAs were not universal features found in all land plants (Dolgosheina et al, 2008). However, homologs of key genes known to be responsible for hcsiRNA biogenesis and function in angiosperms clearly exist in diverse plant lineages (Zong et al, 2009;Banks et al, 2011;Huang et al, 2015;Wang and Ma, 2015;You et al, 2017). Reverse genetic analyses of these homologs, coupled with sRNA-seq analyses of the mutants, has shown that hc-siRNAs exist in the moss P. patens (Cho et al, 2008;Coruh et al, 2015), which implies that the pathway was most likely present in the last common ancestor of all land plants.…”