Epigenetic Mutation of RAV6 Affects Leaf Angle and Seed Size in Rice  

Abstract

Heritable epigenetic variants of genes, termed epialleles, can broaden genetic and phenotypic diversity in eukaryotes. Epialleles may also provide a new source of beneficial traits for crop breeding, but very few epialleles related to agricultural traits have been identified in crops. Here, we identified Epi-rav6, a gain-of-function epiallele of rice (Oryza sativa) RELATED TO ABSCISIC ACID INSENSITIVE3 (ABI3)/VIVIPAROUS1 (VP1) 6 (RAV6), which encodes a B3 DNA-binding domain-containing protein. The Epi-rav6 plants show larger lamina inclination and smaller grain size; these agronomically important phenotypes are inherited in a semidominant manner. We did not find nucleotide sequence variation of RAV6. Instead, we found hypomethylation in the promoter region of RAV6, which caused ectopic expression of RAV6 in Epi-rav6 plants. Bisulfite analysis revealed that cytosine methylation of four CG and two CNG loci within a continuous 96-bp region plays essential roles in regulating RAV6 expression; this region contains a conserved miniature inverted repeat transposable element transposon insertion in cultivated rice genomes. Overexpression of RAV6 in the wild type phenocopied the Epi-rav6 phenotype. The brassinosteroid (BR) receptor BR INSENSITIVE1 and BR biosynthetic genes EBISU DWARF, DWARF11, and BR-DEFICIENT DWARF1 were ectopically expressed in Epi-rav6 plants. Also, treatment with a BR biosynthesis inhibitor restored the leaf angle defects of Epi-rav6 plants. This indicates that RAV6 affects rice leaf angle by modulating BR homeostasis and demonstrates an essential regulatory role of epigenetic modification on a key gene controlling important agricultural traits. Thus, our work identifies a unique rice epiallele, which may represent a common phenomenon in complex crop genomes.Epigenetic gene variants (epialleles) carry heritable changes in gene expression that do not result from alterations in the underlying DNA sequence (Kakutani, 2002). In eukaryotes, cytosine DNA methylation, a conserved epigenetic mark, plays essential roles in the silencing of transposable elements (TEs) and genes (Law and Jacobsen, 2010). In higher plants, the few known epialleles involve alterations in DNA methylation, indicating that this epigenetic marker makes a large contribution to epigenetic diversity. The Arabidopsis (Arabidopsis thaliana) clark kent epiallele, which has hypermethylated cytosines at the SUPERMAN locus, causes increased numbers of stamens and carpels (Jacobsen and Meyerowitz, 1997). Also, DNA hypomethylation at two direct repeats in the promoter region of FLOWERING WAGENINGEN (FWA; Soppe et al., 2000) causes the late-flowering phenotype in Arabidopsis plants carrying the fwa epiallele. Plants carrying the natural epiallele hypermethylated at Linaria cycloidea-like show altered floral symmetry, from bilateral to radial, in Linaria vulgaris (Cubas et al., 1999). In tomato (Solanum lycopersicum), the Colorless nonripening phenotype results from hypermethylation at the promoter of SQUAMOSA promoter-binding protein like (Mann...

An Efficient Strategy Combining SSR Markers- and Advanced QTL-seq-driven QTL Mapping Unravels Candidate Genes Regulating Grain Weight in Rice

“…Interestingly, the strong grain weight-associated SNP allelic variants-containing RAV1AAT and CARGCW8GAT known cis -regulatory elements/TF-binding sites of two genes identified are known to be involved in growth and development by transcriptional regulation/interaction and preferential/tissue-specific expression of many important TF genes in seeds of rice (Zheng et al, 2009; Bajaj et al, 2015b). The direct interactions of the RAV1 (related to ABI3/VP1 ) and CArG TF DNA-binding protein with other TF genes including MADS-domain protein-encoding genes, SEPALLATA3 and AGAMOUS-like 15 ( AGL15 ), respectively, have been established to be the key regulators of seed development in Arabidopsis (Tang and Perry, 2003; Le et al, 2010; Agarwal et al, 2011; Zhang et al, 2015). Therefore, it would be interesting to study transcriptional regulation and binding/interactions of two SCP and GH genes at known cis -regulatory element regions/TF-binding sites with MADS TF genes in controlling grain weight/size of rice.…”

Metabolomics for Plant Improvement: Status and Prospects

“…In Arabidopsis, disruption of MSH1 caused altered plant growth phenotypes due to hypermethylation of chromosome segments ( Virdi et al, 2015 ). Hypomethylation of RAV6 promoter in rice Epi-rav6 mutant resulted altered leaf size and grain size, via modulating brassinosteroid (BR) homeostasis ( Xianwei et al, 2015 ). In maize, the IPA mutation affects the biosynthesis and accumulation of phytic acid in the seed, which influences germination along with affecting plant growth and responses to various environmental conditions ( Pilu et al, 2009 ).…”

Regulation of Brassinosteroid Homeostasis in Higher Plants

“…Loss of CYP85A2/BR6ox2 farnesylation results in reduced BL accumulation, similar to the mutation of CYP85A2/BR6ox2 ( Northey et al, 2016 ; Jamshed et al, 2017 ). In rice, RAVL1 and RAV6, two homologous B3 transcription factors, mediate activation of both OsBRI1 and the BR biosynthetic genes that have antagonistic actions on BR levels to ensure the basal activity of the BR signaling and biosynthetic pathways ( Il Je et al, 2010 ; Zhang et al, 2015 ). Rice microRNA osa-miR1848 mediates OsCYP51G3 mRNA cleavage to regulate phytosterol and BR biosynthesis ( Xia et al, 2015 ).…”