title: De novo methyltransferase, OsDRM2, interacts with the ATP-dependent RNA helicase, OseIF4A, in rice.
abstract: Domains rearranged methyltransferases (DRMs) are the de novo methyltransferases that regulate cytosine methylation in plants in a manner similar to the animal de novo methyltransferases, DNMT3a and DNMT3b. These enzymes catalyze the establishment of new methylation patterns and are guided to target sites by small RNAs through the process of RNA-directed DNA methylation (RdDM). In the current accepted view for RdDM, intricate interactions among transcription factors/chromatin modifying proteins and the large subunits of plant-specific polymerases, Pol IV and Pol V, regulate the 24-nt small interfering RNA guided de novo methylation of cytosines. The RNA-induced silencing complex assembled on Pol-V-transcribed non-coding RNA finally facilitates the recruitment of DRM2 by unknown mechanism/protein interactions to chromatin sites. In an attempt to determine the cellular proteins that specifically interact with DRM2, a yeast two-hybrid screen was performed using young rice panicles. We report that rice DRM2 interacts with the ATP-dependent RNA helicase, eIF4A. Direct interaction between the two proteins is demonstrated in vivo by bimolecular fluorescence complementation method and in vitro by histidine-pull-down assays. Deletion analysis reveals that interaction between OsDRM2 and OseIF4A is specifically mediated through ubiquitin-associated domain of OsDRM2 while, both domains 1 and 2 of OseIF4A are critical for mediating strong interaction with OsDRM2 in vivo. Interaction between Arabidopsis eIF4AI and eIF4AII with OsDRM2 and nuclear localization of these complexes suggests possible conservation of functional interaction between de novo methyltransferases and the translation initiation factor, eIF4A, in RdDM across plant species.
authors.affiliation: University School of Biotechnology, Guru Gobind Singh Indraprastha University, Sector 16C, Dwarka, New Delhi 110078, India.
authors.forename: Meenakshi
authors.initials: M
authors.lastname: Dangwal
authors.forename: Garima
authors.initials: G
authors.lastname: Malik
authors.forename: Sanjay
authors.initials: S
authors.lastname: Kapoor
authors.forename: Meenu
authors.initials: M
authors.lastname: Kapoor
mesh_terms: D000818:Animals; D002498:Centrifugation; D039601:Eukaryotic Initiation Factor-4A / Q000378:metabolism*; D005453:Fluorescence; D008780:Methyltransferases / Q000378:metabolism*; D012275:Oryza / Q000201:enzymology*; D010940:Plant Proteins / Q000378:metabolism; D011485:Protein Binding; D025941:Protein Interaction Mapping; D017384:Sequence Deletion; D020798:Two-Hybrid System Techniques
chemical_list: D010940:Plant Proteins; D008780:Methyltransferases; D039601:Eukaryotic Initiation Factor-4A
keywords: BiFC; C-terminal domain; CTD; DEAD box; DNA methyltransferase; DRM; RISC; RNA-directed DNA methylation; RNA-induced silencing complex; RdDM; bimolecular fluorescence complementation; domains rearranged methyltransferase; protein interaction; siRNA; small interfering RNA; translation initiation; ubiquitin-associated domain