Enrichment of Cytoplasmic RNA Granules from Arabidopsis Seedlings

Zhen Lei; Eun Yu Kim; Jungnam Cho

doi:10.21769/BioProtoc.4212

Enrichment of Cytoplasmic RNA Granules from Arabidopsis Seedlings

Bio Protoc. 2021 Nov 5;11(21):e4212. doi: 10.21769/BioProtoc.4212.

Authors

Zhen Lei^{1

2}, Eun Yu Kim¹, Jungnam Cho^{1

2

3}

Affiliations

¹ National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences (CEMPS), Shanghai Institute of Plant Physiology and Ecology, Chinese Academy of Sciences, Shanghai, China.
² University of Chinese Academy of Sciences, Beijing, China.
³ CAS-JIC Centre of Excellence for Plant and Microbial Science (CEPAMS), Chinese Academy of Sciences, Shanghai, China.

Abstract

RNA granules (RGs) are membraneless intracellular compartments that play important roles in the post-transcriptional control of gene expression. Stress granules (SGs) are a type of RGs that form under environmental challenges and/or internal cellular stresses. Stress treatments lead to strong mRNAs translational inhibition and storage in SGs until the normal growth conditions are restored. Intriguingly, we recently showed that plant stress granules are associated with siRNA bodies, where the RDR6-mediated and transposon-derived siRNA biogenesis occurs ( Kim et al., 2021 ). This protocol provides a technical workflow for the enrichment of cytoplasmic RGs from Arabidopsis seedlings. We used the DNA methylation-deficient ddm1 mutant in our study, but the method can be applied to any other plant samples with strong RG formation. The resulting RG fractions can be further tested for either RNAs or proteins using RNA-seq and mass spectrometry-based proteomics.

Keywords: DDM1; RNA granule; Stress granule; Transposon; siRNA body.