Plant RNA-binding proteins:Phase separation dynamics and functional mechanisms underlying plant development and stress responses

RNA-binding proteins(RBPs)accompany RNA from synthesis to decay,mediating every aspect of RNA metabolism and impacting diverse cellular and developmental processes in eukaryotes.Many RBPs undergo phase separation along with their bound RNA to form and function in dynamic membraneless biomolecular condensates for spatiotemporal coordination or regulation of RNA metabolism.Increasing evidence suggests that phase-separating RBPs with RNA-binding domains and intrinsically disordered regions play important roles in plant development and stress adaptation.Here,we summarize the current knowledge about how dynamic partitioning of RBPs into condensates controls plant development and enables sensing of experimental changes to confer growth plasticity under stress conditions,with a focus on the dynamics and functional mechanisms of RBP-rich nuclear condensates and cytoplasmic granules in mediating RNA metabolism.We also discuss roles of multiple factors,such as environmental signals,protein modifications,and N6-methyladenosine RNA methylation,in modulating the phase separation behaviors of RBPs,and highlight the prospects and challenges for future research on phase-separating RBPs in crops.

A dephosphorylation-dependentmolecular switch for FT repression mediates flowering in Arabidopsis

The reproductive success offlowering plants relies greatly on precise timing of thefloral transition,which isnely modulated by a complex network offloral regulators.As a mainfloral integrator,FLOWERING LOCUS T(FT)is also an essential constituent of theflorigen that is transported from leaves to shoot apices to induceflowering.FT is specically transcribed in leaf vascular tissues,where its production is suppressed by manyflowering repressors,including the MYB transcription factor EARLY FLOWERING MYB PROTEIN(EFM).Here,we show that a plant CTD phosphatase,C-TERMINAL DOMAIN PHOSPHATASE-LIKE 2(CPL2),suppresses FT expression in leaf vascular tissues by modulating the binding activity of EFM.CPL2 interacts with and dephosphorylates EFM to facilitate the binding of dephosphorylated EFM to FT chromatin,thereby inhibitingflowering.Our results suggest that CPL2-mediated dephosphorylation of thefloral repressor EFM serves as a molecular switch,adding another layer of regulation tone-tune FT transcription and ensure thatflowering occurs at an appropriate time.

Nuclear translocation of OsMADS25 facilitated by OsNAR2.1 in reponse to nitrate signals promotes rice root growth by targeting OsMADS27 and OsARF7

Nitrate is an important nitrogen source and signaling molecule that regulates plant growth and development.Although several components of the nitrate signaling pathway have been identified,the detailed mechanisms are still unclear.Our previous results showed that OsMADS25 can regulate root development in response to nitrate signals,but the mechanism is still unknown.Here,we try to answer two key questions:how does OsMADS25 move from the cytoplasm to the nucleus,and what are the direct target genes activated by OsMADS25 to regulate root growth after it moves to the nucleus in response to nitrate?Our results demonstrated that OsMADS25 moves from the cytoplasm to the nucleus in the presence of nitrate in an OsNAR2.1-dependentmanner.Chromatin immunoprecipitation sequencing,chromatin immunoprecipitation qPCR,yeast one-hybrid,and luciferase experiments showed that OsMADS25 directly activates the expression of OsMADS27 and OsARF7,which are reported to be associated with root growth.Finally,OsMADS25-RNAi lines,the Osnar2.1 mutant,and OsMADS25-RNAi Osnar2.1 lines exhibited significantly reduced root growth compared with the wild type in response to nitrate supply,and expression of OsMADS27 and OsARF7 was significantly suppressed in these lines.Collectively,these results reveal a new mechanismby which OsMADS25 interacts with OsNAR2.1.This interaction is required for nuclear accumulation of OsMADS25,which promotes OsMADS27 and OsARF7 expression and root growth in a nitratedependent manner.

Whole-Genome Resequencing of a Worldwide Collection of Rapeseed Accessions Reveals the Genetic Basis of Ecotype Divergence

Rapeseed (Brassica napus),an important oilseed crop,has adapted to diverse climate zones and latitudes by forming three main ecotype groups,namely winter,semiwinter,and spring types. However,genetic variations underlying the divergence of these ecotypes are largely unknown. Here,we report the global pattern of genetic polymorphisms in rapeseed determined by resequencing a worldwide collection of 991 germplasm accessions.A total of 5.56 and 5.53 million singlenucleotide polymorphisms (SNPs)as Well as 1.86 and 1.92 million InDels were identified by mapping reads to the reference genomes of "Darmor-bzh"and "Tapidor,"respectively.We generated a map of allelic drift paths that shows splits and mixtures of the main populations,and revealed an asymmetric evolution of the two subgenomes of B.napus by calculating the genetic diversity and linkage disequilibrium parameters.Selective-sweep analysis revealed genetic changes in genes orthologous to those regulating various aspects of plant development and response to stresses.A genome-wide association study identified SNPs in the promoter regions of FLOWERING LOCUS T and FLOWERING LOCUS C orthologs that corresponded to the different rapeseed ecotype groups. Our study provides important insights into the genomic footprints of rapeseed evolution and flowering-time divergence among three ecotype groups,and will facilitate screening of molecular markers for accelerating rapeseed breeding.

5-Methylcytosine RNA Methylation in Arabidopsis Thaliana

5-Methylcytosine （m^5C） is a well-characterized DNA modification, and is also predominantly reported in abundant non-coding RNAs in both prokaryotes and eukaryotes. However, the distribution and biological functions of m^5C in plant mRNAs remain largely unknown. Here, we report transcriptome-wide profiling of RNA m^5C in Arabidopsis thaliana by applying m^5C RNA immunoprecipitation followed by a deep- sequencing approach （m^5C-RIP-seq）. LC-MS/MS and dot blot analyses reveal a dynamic pattern of m^5C mRNA modification in various tissues and at different developmental stages, m^5C-RIP-seq analysis identified 6045 m^5C peaks in 4465 expressed genes in young seedlings. We found that m^5C is enriched in coding sequences with two peaks located immediately after start codons and before stop codons, and is associated with mRNAs with low translation activity. We further demonstrated that an RNA （cytosine-5）-methyl- transferase, tRNA-specific methyltransferase 4B （TRM4B）, exhibits m^5C RNA methyltransferase activity. Mutations in TRM4B display defects in root development and decreased m^5C peaks. TRM4B affects the transcript levels of the genes involved in root development, which is positively correlated with their mRNA stability and m^5C levels. Our results suggest that m^5C in mRNA is a new epitranscriptome marker inArabidopsis, and that regulation of this modification is an integral part of gene regulatory networks underlying plant development.

New insights into gibberellin signaling in regulating flowering in Arabidopsis

In angiosperms,floral transition is a key developmental transition from the vegetative to reproductive growth,and requires precise regulation to maximize the reproductive success.A complex regulatory network governs this transition through integrating flowering pathways in response to multiple exogenous and endogenous cues.Phytohormones are essential for proper plant developmental regulation and have been extensively studied for their involvement in the floral transition.Among various phytohormones,gibberellin(GA)plays a major role in affecting flowering in the model plant Arabidopsis thaliana.The GA pathway interact with other flowering genetic pathways and phytohormone signaling pathways through either DELLA proteins or mediating GA homeostasis.In this review,we summarize the recent advances in understanding the mechanisms of DELLA-mediated GA pathway in flowering time control in Arabidopsis,and discuss its possible link with other phytohormone pathways during the floral transition.

Resistant starch formation in rice:Genetic regulation and beyond

Resistant starch(RS),a healthy dietary fiber,is a particular type of starch that has attracted much research attention in recent years.RS has important roles in reducing glycemic index,postprandial blood glucose levels,and serum cholesterol levels,thereby improving and preventing many diseases,such as diabetes,obesity,and cardiovascular disease.The formation of RS is influenced by intrinsic properties of starch(e.g.,starch granule structure,starch crystal structure,and amylose-to-amylopectin ratio)and non-starch components(e.g.,proteins,lipids,and sugars),aswell as storage and processing conditions.Recent studies have revealed that several starch-synthesis-related genes(SSRGs)are crucial for the formation of RS during seed development.Several transcription factors and mRNA splicing factors have been shown to affect the expression or splicing of SSRGs that regulate RS content,suggesting their potential roles in RS formation.This review focuses mainly on recent research progress on the genetic regulation of RS content and discusses the emerging genetic and molecular mechanisms of RS formation in rice.

Nucleoporin 160 Regulates Flowering through Anchoring HOS1 for Destabilizing CO in Arabidopsis

Nuclear pore complexes(NPCs),which comprise multiple copies of nucleoporins(Nups),are large protein assemblies embedded in the nuclear envelope connecting the nucleus and cytoplasm.Although it has been known that Nups affect flowering in Arabidopsis,the underlying mechanisms are poorly understood.Here,we show that loss of function of Nucleoporin 160(Nup160)leads to increased abundance of CONSTANS(CO)protein and the resulting upregulation of FLOWERING LOCUS T(FT)specifically in the morning.We demonstrate that Nup160 regulates CO protein stability through affecting NPC localization of an E3-ubiquitin ligase,HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES1(HOS1),which destabilizes CO protein in the morning period.Taken together,these results provide a mechanistic understanding of Nup function in the transition from vegetative to reproductive growth,suggesting that deposition of HOS1 at NPCs by Nup160 is essential for preventing precocious flowering in response to photoperiod in Arabidopsis.

Epitranscriptome engineering in crop improvement

Biochemical modifications of RNAs,collectively designated as the epitranscriptome,have emerged as fundamental mechanisms derlying gene regulation.Recent studies have shown that N6-methyladenosine(m6A)and 5-methycytosine(m5C)modifications govern multifaceted developmental processes and environmental resporises in plants.The potential of engineering the epitranscrip-tome in crop improvement was nicely demonstrated in a recent study(Yu et al.,2021),where the authors took advantage of epitranscriptome engi neering to vigorously in crease crop yield.