Regulation of gene expression at the post-transcriptional level is mainly achieved by proteins containing well-defined sequence motifs involved in RNA binding. The most widely spread motifs are the RNA recognition motif (RRM) and the K homology (KH) domain. In this article, we survey the complete Arabidopsis thaliana genome for proteins containing RRM and KH RNA-binding domains. The Arabidopsis genome encodes 196 RRM-containing proteins, a more complex set than found in Caenorhabditis elegans and Drosophila melanogaster. In addition, the Arabidopsis genome contains 26 KH domain proteins. Most of the Arabidopsis RRM-containing proteins can be classified into structural and/or functional groups, based on similarity with either known metazoan or Arabidopsis proteins. Approximately 50% of Arabidopsis RRM-containing proteins do not have obvious homologues in metazoa, and for most of those that are predicted to be orthologues of metazoan proteins, no experimental data exist to confirm this. Additionally, the function of most Arabidopsis RRM proteins and of all KH proteins is unknown. Based on the data presented here, it is evident that among all eukaryotes, only those RNA-binding proteins that are involved in the most essential processes of post-transcriptional gene regulation are preserved in structure and, most probably, in function. However, the higher complexity of RNA-binding proteins in Arabidopsis, as evident in groups of SR splicing factors and poly(A)-binding proteins, may account for the observed differences in mRNA maturation between plants and metazoa. This survey provides a first systematic analysis of plant RNA-binding proteins, which may serve as a basis for functional characterisation of this important protein group in plants.

DNA methylation is an important epigenetic mark in many eukaryotes1-5. In plants, 24-nt small interfering RNAs (siRNAs) bound to the effector protein, Argonaute 4 (AGO4) can direct de novo DNA methylation by the methyltransferase DRM22,4-6. Here we report a new regulator of RNA-directed DNA methylation (RdDM) in Arabidopsis: RDM1. Loss-of-function mutations in the RDM1 gene impair the accumulation of 24-nt siRNAs, reduce DNA methylation, and release transcriptional gene silencing at RdDM target loci. RDM1 encodes a small protein that appears to bind single-stranded methyl DNA, and associates and co-localizes with RNA polymerase II, AGO4 and DRM2 in the nucleus. Our results suggest that RDM1 is a component of the RdDM effector complex and may play a role in linking siRNA production with pre-existing or de novo cytosine methylation. Our results also suggest that although RDM1 and Pol V may function together at some RdDM target sites in the peri-nucleolar siRNA processing center, Pol II rather than Pol V is associated with the RdDM effector complex at target sites in the nucleoplasm.

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