Inositol Pyrophosphate lnsP8 Acts as an Intracellular Phosphate Signal in Arabidopsis

The maintenance of cellular phosphate(Pi)homeostasis is of great importance in living organisms.The SPX domain-containing protein 1(SPX1)proteins from both Arabidopsis and rice have been proposed to act as sensors of Pi status.The molecular signal indicating the cellular Pi status and regulating Pi homeostasis in plants,however,remains to be identified,as Pi itself does not bind to the SPX domain.Here,we report the identification of the inositol pyrophosphate lnsP8 as a signaling molecule that regulates Pi homeostasis in Arabidopsis.Polyacrylamide gel electrophoresis profiling of InsPs revealed that lnsP8 level positively cor­relates with cellular Pi concentration.We demonstrated that the homologs of diphosphoinositol pentaki-sphosphate kinase(PPIP5K),VIH1 and VIH2,function redundantly to synthesize lnsP8,and that the vih1 vih2 double mutant overaccumulates Pi.SPX1 directly interacts with PHR1,the central regulator of Pi star­vation responses,to inhibit its function under Pi-replete conditions.However,this interaction is compro­mised in the vih1 vih2 double mutant,resulting in the constitutive induction of Pi starvation-induced genes,indicating that plant cells cannot sense cellular Pi status without lnsP8.Furthermore,we showed that lnsP8 could directly bind to the SPX domain of SPX1 and is essential for the interaction between SPX1 and PHR1.Collectively,our study suggests that lnsP8 is the intracellular Pi signaling molecule serving as the ligand of SPX1 for controlling Pi homeostasis in plants.

The H3K9me2-binding protein AGDP3 limits DNA methylation and transcriptional gene silencing in Arabidopsis

DNA methylation,a conserved epigenetic mark,is critical for tuning temporal and spatial gene expression.The Arabidopsis thaliana DNA glycosylase/lyase REPRESSOR OF SILENCING 1(ROS1)initiates active DNA demethylation and is required to prevent DNA hypermethylation at thousands of genomic loci.However,how ROS1 is recruited to specific loci is not well understood.Here,we report the discovery of Arabidopsis AGENET Domain Containing Protein 3(AGDP3)as a cellular factor that is required to prevent gene silencing and DNA hypermethylation.AGDP3 binds to H3K9me2 marks in its target DNA via its AGD12 cassette.Analysis of the crystal structure of the AGD12 cassette of AGDP3 in complex with an H3K9me2 peptide revealed that dimethylated H3 K9 and unmodified H3 K4 are specifically anchored into two different surface pockets.A histidine residue located in the methyllysine binding aromatic cage provides AGDP3 with pH-dependent H3K9me2 binding capacity.Our results uncover a molecular mechanism for the regulation of DNA demethylation by the gene silencing mark H3K9me2.