Os10g0577600

The Rice JMJ706 is a jmjC domain-containing gene, encoding H3K9 demethylase which is required for floral organ development.

Annotated Information

Background

Histone lysine methylation is an important epigenetic modification with both activating and repressive roles in gene expression . There are six lysine residues in histone N-termini that are predominantly methylated, with the methylation of histone H3 lysine 4 (H3K4) and lysine 36 (H3K36) primarily having an activating function, whereas the methylation of histone H3 lysine 9 (H3K9) and lysine 27 (H3K27) and histone H4lysine 20 (H4K20) is essentially associated with repressed chromatin . Histone lysine residues can be mono-, di- or trimethylated. Each distinct methyl state is associated with different biological functions .

Jumonji C (jmjC) domain-containing proteins have been suggested to function as histone demethylases . Using biochemical approaches, the first jmjC domain-containing histone demethylase, JHDM1 (jmjC domain-containing histone demethylase 1), was identified and shown to reverse H3K36 mono- and dimethylation (H3K36me1 and H3K36me2), and the jmjC domain was demonstrated to be the catalytic domain . Jumonji C domain-containing histone demethylases catalyze lysine demethylation through an oxidative reaction that requires iron Fe(II) and α-ketoglutarate as cofactors.

Function

"Fig.1 In vitro assays of JMJ706 histone demethylation activity (from reference )."

JMJ706 encodes a heterochromatin-associated H3K9 demethylase involved in the regulation of flower development in rice. Histone lysine ethylation is an important epigenetic modification with both activating and repressive roles in gene expression. Jumonji C (jmjC) domain-containing proteins have been shown to reverse histone methylation in non plant model systems. Here, we show that plant Jumonji C proteins have both conserved and specific features compared with mammalian homologues. In particular, the rice JMJD2 family jmjC gene JMJ706 is shown to encode a heterochromatin-enriched protein. The JMJ706 protein specifically reverses di- and trimethylations of lysine 9 of histone H3 (H3K9) in vitro. Loss-of-function mutations of the gene lead to increased di- and trimethylations of H3K9 and affect the spikelet development, including altered floral morphology and organ number. Gene expression and histone modification analysis indicates that JMJ706 regulates a ubset of flower development regulatory genes.

Histone demethylation activity has been shown to be jmjC domain dependent . To study JMJ706 function, a truncated JMJ706 protein containing the jmjN and jmjC domains and an additional GST tag was purified from Escherichia coli cells and tested for in vitro histone demethylase activity. The incubated histones were analyzed by Western blot tests with antibodies specific to histone H3 modification modules. As shown in Fig.1, the GST-JMJ706 fusion protein reduced the level of H3K9me2 and H3K9me3 but slightly increased the level of H3K9me1. The methylation of other lysine residues was not significantly altered, except a slight decrease observed for H3K36me2. Glutathione S-transferase alone had no effect on histone modification. This suggests that JMJ706 may be mainly involved in the demethylation of H3K9me2 and H3K9me3.

Evolution

"Fig.2 Phylogenetic relationship of jmjC domain-containing proteins from O. sativa (Os), A. thaliana (At), and H. sapiens (Hs). (from reference )."

A genomic survey revealed 20 and 21 jmjC domain-containing genes in rice and Arabidopsis, respectively (Fig. 2). The sequence alignment resulted in the discrimination of five groups containing the plant proteins. Groups I, II, and IV were found to be related to the JARID, JMJD2, and JHDM2 subfamilies, respectively. Group V could be divided into several subgroups, each of which was found to have related members of the human 'JmjC domain-only' subfamily .

Localization

"Fig.3 Subcellular localization of the protein (from reference )."

JMJ706 is enriched in heterochromatin domains. To study JMJ706 localization in rice nuclei, a construct expressing a JMJ706 fusion with the GFP was used to transform WT rice plants. Nuclei of the transgenic leaf mesophyll cells showed an enrichment of the fusion protein in spots that overlapped with 4,6-diamidino-2-phenylindole (DAPI)–stained regions (Fig. 3B). To confirm this result, a construct expressing FLAG-tagged JMJ706 protein was used to complement JMJ706 T-DNA insertion mutant lines . The FLAG-tagged protein was also observed in enriched DAPI-stained domains (Fig. 3C).

Mutation

"Fig.4 JMJ706 loss-of-function mutation affects spikelet development.(from reference )."

JMJ706 Loss-of-Function Mutation Affects Floral Organogenesis. A search of a database of a rice T-DNA insertion library identified two insertion lines of the JMJ706 gene (Fig. 4A).Characterization of the insertion mutants revealed similar spikelet morphology defects (Fig. 4B).The phenotype cosegregated with the homozygous state of the insertions and the absence of JMJ706 mRNA accumulation (Fig. 4C).The mutant spikelets showed a variety of defects, mainly on floral organ number (Fig. 4 D–H). Some spikelets were depleted of lemma and/or palea (Fig. 4D), whereas others had an additional piece of palea (Fig. 4E). Increased numbers of stamens and pistils were also observed (Fig. 4F). In addition, vitrified tissues were seen in some spikelets (Fig. 4G). Scanning microscopy revealed irregular cell size and arrangement on the mutant palea surface with increased bristle numbers (Fig. 4H). Some spikelets could still produce seeds. These mutant seeds had a deformed shape, but they germinated and produced normal seedlings, suggesting that embryogenesis was not affected.

Labs working on this gene

Dao-Xiu Zhou, Institut de Biotechnologie des Plantes, Universite´ Paris Sud 11, 91405 Orsay, France