Six-Rowed Spike3 (VRS3) Is a Histone Demethylase That Controls Lateral Spikelet Development in Barley

Abstract

The complex nature of crop genomes has long prohibited the efficient isolation of agronomically relevant genes. However, recent advances in next-generation sequencing technologies provide new ways to accelerate fine-mapping and gene isolation in crops. We used RNA sequencing of allelic six-rowed spike3 (vrs3) mutants with altered spikelet development for gene identification and functional analysis in barley (Hordeum vulgare). Variant calling in two allelic vrs3 mutants revealed that VRS3 encodes a putative histone Lys demethylase with a conserved zinc finger and Jumonji C and N domain. Sanger sequencing of this candidate gene in independent allelic vrs3 mutants revealed a series of mutations in conserved domains, thus confirming our candidate as the VRS3 gene and suggesting that the row type in barley is determined epigenetically. Global transcriptional profiling in developing shoot apical meristems of vrs3 suggested that VRS3 acts as a transcriptional activator of the row-type genes VRS1 (Hv.HOMEOBOX1) and INTERMEDIUM-C (INT-C; Hv.TEOSINTE BRANCHED1). Comparative transcriptome analysis of the row-type mutants vrs3, vrs4 (Hv.RAMOSA2), and int-c confirmed that all three genes act as transcriptional activators of VRS1 and quantitative variation in the expression levels of VRS1 in these mutants correlated with differences in the number of developed lateral spikelets. The identification of genes and pathways affecting seed number in small grain cereals will enable to further unravel the transcriptional networks controlling this important yield component.

Genetic pathways controlling inflorescence architecture and development in wheat and barley

“…This conclusion is supported by a previous study demonstrating that reduced levels of GA extend the duration of inflorescence development and delay flowering in barley (Boden et al ). These studies implicate hormones as important contributors to spikelet row‐type architecture and inflorescence development in barley, which is supported by genome‐wide expression analysis in vrs3 mutants that showed differential expression of genes involved in cytokinin and jasmonic acid metabolism, relative to wild‐type plants (Bull et al ; van Esse et al ; Youssef et al ). Taken together, these results suggest that adjustments to the levels and distribution of hormones, during inflorescence development, can be used to generate diverse inflorescence architectures in barley (Boden ).…”

“…This conclusion is supported by the increased lateral spikelet fertility observed in lines that contain mutations preventing production of a functional VRS5/INT‐C protein (Ramsay et al ). Less is known about the specific spatial and temporal expression of Vrs5 , relative to Vrs1 , Vrs3 and Vrs4 ; however, Vrs5 is expressed in the developing inflorescence and is reduced in vrs3 mutants, which is consistent with a role in suppressing lateral spikelet development (Ramsay et al ; van Esse et al ; Bull et al ). Interestingly, Vrs1 expression is not significantly different in vrs5 / int‐c.5 loss of function mutants, relative to wild‐type, suggesting that Vrs5 acts independently of Vrs1 to suppress lateral spikelet fertility (Sakuma et al ).…”

“…Expression of Vrs4 is detected earlier than Vrs1 , and is localized predominantly within the lateral spikelet primordia during early developmental stages, suggesting that VRS4 protein regulates lateral spikelet fertility by acting upstream of Vrs1 (Koppolu et al ). Similarly, Vrs1 expression is reduced in vrs3 mutants that contain mutations within a gene that encodes a histone demethylase, which is predicted to facilitate activation of Vrs1 by removing repressive methyl marks and acting as a positive regulator of Vrs4 (van Esse et al ; Bull et al ). Taken together, these findings indicate that Vrs3 and Vrs4 regulate row‐type architecture of the barley inflorescence by converging to positively regulate transcription of Vrs1 .…”

Developmental pathways for shaping spike inflorescence architecture in barley and wheat

“…Genetics and gene expression analyses suggest that HvRA2 regulates the barley row‐type pathway by promoting VRS1 transcription (Koppolu et al ). Unlike other row‐type genes, which encode transcription factors, VRS3 codes for an enzyme, a putative Jumonji C‐type (JMJC) H3K9me2/3 histone demethylase (HvJMJC) (Bull et al ; van Esse et al ). Interestingly, VRS3 appears to promote the transcription of all known row‐type genes supposedly by clearing repressive chromatin methylation marks (Bull et al ; van Esse et al ) (Figure G).…”

Interaction between row‐type genes in barley controls meristem determinacy and reveals novel routes to improved grain

“…VRS2 encodes a homologue of the Arabidopsis SHORT INTERNODES gene, whose loss of function is associated with hormonal imbalances between auxin and cytokinin along the spike (Youssef et al, 2017). VRS3 encodes a putative Jumonji C-type (JMJC) H3K9me2/3 histone demethylase (Bull et al, 2017;van Esse et al, 2017), orthologous to the rice gene OsJMJ706 that regulates rice spikelet morphology (Sun & Zhou, 2008). Comparative transcriptomics suggests that VRS3 promotes the expression of other VRS genes (Bull et al, 2017;van Esse et al, 2017).…”