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Abstract
Transcription by RNA polymerase V (Pol V) in plants is required for RNA-directed DNA methylation, leading to transcriptional gene silencing. Global chromatin association of Pol V requires components of the DDR complex DRD1, DMS3 and RDM1, but the assembly process of this complex and the underlying mechanism for Pol V recruitment remain unknown. Here we show that all DDR complex components co-localize with Pol V, and we report the cryoEM structures of two complexes associated with Pol V recruitment—DR (DMS3-RDM1) and DDR′ (DMS3-RDM1-DRD1 peptide), at 3.6 Å and 3.5 Å resolution, respectively. RDM1 dimerization at the center frames the assembly of the entire complex and mediates interactions between DMS3 and DRD1 with a stoichiometry of 1 DRD1:4 DMS3:2 RDM1. DRD1 binding to the DR complex induces a drastic movement of a DMS3 coiled-coil helix bundle. We hypothesize that both complexes are functional intermediates that mediate Pol V recruitment.
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1 Department of Molecular, Cellular and Developmental Biology, University of California, Los Angeles (UCLA), Los Angeles, CA, USA; Department of Microbiology, Faculty of Medicine, Chiang Mai University, Chiang Mai, Thailand
2 Department of Microbiology, Immunology and Molecular Genetics, UCLA, Los Angeles, CA, USA; California NanoSystems Institute (CNSI), UCLA, Los Angeles, CA, USA
3 Department of Molecular, Cellular and Developmental Biology, University of California, Los Angeles (UCLA), Los Angeles, CA, USA
4 Department of Biology, Institute of Molecular Systems Biology, ETH Zürich, Zürich, Switzerland
5 Department of Biology, Institute of Molecular Systems Biology, ETH Zürich, Zürich, Switzerland; Faculty of Science, University of Zürich, Zürich, Switzerland
6 Department of Molecular, Cellular and Developmental Biology, University of California, Los Angeles (UCLA), Los Angeles, CA, USA; Zhejiang University-University of Edinburgh Institute, Zhejiang University School of Medicine, Hangzhou, P. R. China
7 Keck School of Medicine, University of Southern California, Los Angeles, CA, USA
8 Department of Biological Chemistry, UCLA, Los Angeles, CA, USA
9 Department of Molecular, Cellular and Developmental Biology, University of California, Los Angeles (UCLA), Los Angeles, CA, USA; Howard Hughes Medical Institute (HHMI), UCLA, Los Angeles, CA, USA