Regulating Plant Development Research Articles

Nitro-fatty acids modulate germination onset through S-nitrosothiol metabolism.

-Nitro-fatty acids (NO2-FAs) have emerged as key components of nitric oxide (NO) signalling in eukaryotes. We previously described how nitro-linolenic acid (NO2-Ln), the major NO2-FA detected in plants, regulates S-nitrosoglutathione (GSNO) levels in Arabidopsis (Arabidopsis thaliana). However, the underlying molecular mechanisms remain undefined. Here, we used a combination of physiological, biochemical, and molecular approaches to provide evidence that NO2-Ln modulates S-nitrosothiol (SNO) content through S-nitrosylation of S-nitrosoglutathione reductase1 (GSNOR1) and its impact on germination onset. The aer mutant (a knockout mutant of the alkenal reductase enzyme; AER) exhibits higher NO2-Ln content and lower GSNOR1 transcript levels, reflected by higher SNO content and S-nitrosylated proteins. Given its capacity to release NO, NO2-Ln mediates the S-nitrosylation of GSNOR1, demonstrating that NO2-FAs can indirectly modulate total SNO content in plants. Moreover, the ectopic application of NO2-Ln to dormant seeds enhances germination success similarly to the aer germination rate, which is mediated by the degradation of master regulator ABSCISIC ACID INSENSITIVE 5 (ABI5). Our results establish that NO2-FAs regulate plant development through NO and SNO metabolism and reveal a role of NO2-FAs in plant physiology.

26S Proteasome Subunit SlPBB2 Regulates Fruit Development and Ripening in Tomato.

Proteasomes are protein complexes responsible for degrading unneeded or damaged proteins through proteolysis and play critical roles in regulating plant development and response to environmental stresses. However, it is still unclear whether proteasomes regulate fruit development and ripening. In this study, we investigated the function of a core proteasome subunit, SlPBB2, in tomato fruit. We found that silencing the SlPPB2 gene through virus-induced gene silencing resulted in tomato seedling death, while SlPPB2 RNA interference (RNAi-SlPBB2) plants driven by a fruit-specific promoter PP2C present normal development. Further observation reveals that silencing SlPBB2 in fruits impaired the chloroplast development and chlorophyll metabolism of fruits, increased the fruit size, and delayed fruit ripening. These findings provide novel insights into the role of proteasomes in regulating fruit development and ripening.

Transcriptome Reveals the Differential Regulation of Sugar Metabolism to Saline-Alkali Stress in Different Resistant Oats.

Saline-alkali stress is a major factor limiting the growth of oats. Sugar is the primary carbon and energy source in plants which regulates plant development and growth by regulating enzyme activity and gene expression. Sucrose, glucose, and fructose are ubiquitous plant-soluble sugars that act as signalling molecules in the transcriptional regulation of various metabolic and defence-related genes. In this study, soluble sugars, fructose, sucrose, and starch contents were measured, and transcriptomics was used to determine the differentially expressed genes (DEGs) in saline-sensitive and saline-tolerant oats after 6, 12, 24, and 48 h. DEGs annotated to carbohydrates were selected using the Kyoto Encyclopedia of Genes and Genomes. DEGs involved in carbohydrate metabolism were mainly enriched in the glycolysis/gluconeogenesis and pentose phosphate pathways, fructose and mannose metabolism, and starch and sucrose metabolism. GAPDH, SUPI, SUS2, ATP-PEK, HXK6, FBA4, TBA4, TKT, ISA3, PPDK1, and BAM2 were significantly expressed, and a quantitative reverse transcription polymerase chain reaction verified the transcriptome sequencing results. In this study, oats with different salinity tolerances were used to determine sugar contents under four salinity stress durations, and transcriptome sequencing was used to explore the regulatory mechanism of sugars and provide a reference for elucidating the sugar signalling regulatory mechanism under abiotic stress.

S-Sulfenylation-mediated inhibition of the GSNOR1 activity regulates ovule development in Arabidopsis.

Reactive oxygen species (ROS) and nitric oxide (NO) are two critical classes of signaling molecules that regulate plant development and stress responses. The intracellular level of S-nitrosoglutathione (GSNO), a major bioactive NO species, is regulated by the highly conserved GSNO reductase (GSNOR). However, the molecular mechanisms underlying ROS-mediated regulation of GSNOR remain largely unclear. Here, we show that H2O2 negatively regulates the activity of GSNOR1 during ovule development in Arabidopsis. S-sulfenylation of GSNOR1 at Cys-284 inhibits its enzymatic activity. A GSNOR1C284S mutation causes a reduction of the total SNO level in pistils, thereby disrupting NO homeostasis and eventually leading to defective ovule development. These findings illustrate a unique mechanism by which ROS regulates ovule development through S-sulfenylation-mediated inhibition of the GSNOR activity, thereby establishing a molecular link between ROS and NO signaling pathways in reproductive development.

Insights of cellular and molecular changes in sugarcane response to oxidative signaling

Main conclusionSignificant changes in the proteome highlight essential metabolic adaptations for development and oxidative signaling induced by the treatment of young sugarcane plants with hydrogen peroxide. These adaptations suggest that hydrogen peroxide acts not only as a stressor but primarily as a signaling molecule, triggering specific metabolic pathways that regulate growth and plant resilience.Sugarcane is a crucial crop for sugar and ethanol production, often influenced by environmental signals. Hydrogen peroxide (H2O2) is increasingly recognized as an important signaling molecule that regulates plant development and adaptation. In this study, two-month-old sugarcane plants were treated with varying concentrations of H2O2 to investigate how this molecule acts as a signal at the cellular, biochemical, and proteomic levels. Antioxidant enzyme activity exhibited fluctuations, suggesting a dynamic response to oxidative signaling. Lipid peroxidation, observed through TBARs and scanning electron microscopy, highlighted early membrane modifications. Proteomic analysis (ProteomeXchange PXD048142) identified 2,699 proteins, with 155 showing significant expression changes in response to H2O2 signaling. Bioinformatics, including Principal Component Analysis, revealed distinct proteomic profiles in roots and leaves, indicating tissue-specific metabolic reprogramming. Functional annotation through Gene Ontology and KEGG pathway enrichment showed that oxidative signaling led to the repression of photosynthesis-related pathways in leaves, while promoting pathways related to protein processing, glycolysis, and carbon metabolism in roots. Additionally, bioinformatic tools identified proteins involved in amino acid metabolism, the TCA cycle, and carbohydrate metabolism as critical components of sugarcane's adaptive signaling response. The data suggest that sugarcane plants responded to oxidative signals by adjusting their metabolic networks, promoting sustained development and potential pathways for targeted plant breeding.

Redox modification of m6A demethylase SlALKBH2 in tomato regulates fruit ripening.

Hydrogen peroxide (H2O2) functions as a critical signalling molecule in controlling multiple biological processes. How H2O2 signalling integrates with other regulatory pathways such as epigenetic modification to coordinately regulate plant development remains elusive. Here we report that SlALKBH2, an m6A demethylase required for normal ripening of tomato fruit, is sensitive to oxidative modification by H2O2, which leads to the formation of homodimers mediated by intermolecular disulfide bonds, and Cys39 serves as a key site in this process. The oxidation of SlALKBH2 promotes protein stability and facilitates its function towards the target transcripts including the pivotal ripening gene SlDML2 encoding a DNA demethylase. Furthermore, we demonstrate that the thioredoxin reductase SlNTRC interacts with SlALKBH2 and catalyses its reduction, thereby modulating m6A levels and fruit ripening. Our study establishes a molecular link between H2O2 and m6A methylation and highlights the importance of redox regulation of m6A modifiers in controlling fruit ripening.

β-Aminobutyric acid activates SA-signalling systemic acquired resistance in peach fruit by suppressing the circadian clock associated protein1.

Circadian clock regulates plant development and physiology by anticipating daily environmental changes. Here we studied the core clock protein involved in β-aminobutyric acid (BABA)-inducible SAR resistance to Rhizopus stolonifer in peach fruit. BABA elicitation barely primed the accumulation of jasmonate or ethylene, whose regulation was associated with morning-loop gene expression. Notably, BABA-induced resistance depended on the upregulation of SA signalling, accompanied by increased transcription of specific evening-loop genes. Through Y2H screening, pull-down and co-IP analyses, CIRCADIAN CLOCK ASSOCIATED 1 (CCA1), a morning-expressed clock protein repressed by BABA, was identified as an interacting partner of NPR1 in regulating SA-dependent SAR. A CUT&Tag analysis indicated that the association of CCA1 with its target genes, which are enriched in EE or CBS motifs, was involved in SA pathway. Furthermore, EMSA, DLR, Y3H and Co-ip assays suggested that CCA1 did not directly affect the expression of SA-inducible genes but instead hindered the interaction between NPR1 and TGA1. Overexpression of PpCCA1 attenuated the transcription of SA-responsive PR genes, while mutation of PpCCA1 elevated these expressions. Collectively, PpCCA1 functions as a negative regulator of NPR1-dependent SA signalling through antagonistic crosstalk with the NPR1-TGA1 system, but BABA activates SAR by suppressing PpCCA1 in peach fruit.

Comparative analysis of the JRL gene family in the whole-genome of five gramineous plants.

The Jacalin-related lectins (JRLs) gene family play a crucial role in regulating plant development and responding to environmental stress. However, a systematic bioinformatics analysis of the JRL gene family in Gramineae plants has been lacking. In this study, we identified 101 JRL proteins from five Gramineae species and classified them into eight distinct clades. Most of the AtJRL proteins clustered in the same group and were differentiated from the Gramineae JRL proteins. The analysis of protein motifs, gene structures and protein domain revealed that the JRL genes play diverse functions in Gramineae plants. Duplication events indicated that tandem duplication significantly contributed to the expansion of the JRL family, with most JRL members underwent purifying selection. Tissue expression profile analysis showed that most OsJRL genes were highly expressed in the roots, while ZmJRL genes exhibited high expression in inflorescences. Furthermore, the expression level of OsJRL and ZmJRL genes were influenced by drought, cold, heat and salt stresses, respectively, implying that these genes play important roles in response to various abiotic stresses. RT-qPCR results demonstrated that OsJRL4 was up-regulated under PEG6000 and NaCl stresses, while OsJRL12 and OsJRL26 were down-regulated under PEG6000. These findings provide comprehensive insights into the JRL gene family in Gramineae plants and will facilitate further functional characterization of JRLs.

MicroRNA analysis reveals 2 modules that antagonistically regulate xylem tracheary element development in Arabidopsis.

Tracheary elements (TEs) are vital in the transport of various substances and contribute to plant growth. The differentiation of TEs is complex and regulated by a variety of microRNAs (miRNAs). However, the dynamic changes in miRNAs during each stage of TE differentiation remain unclear, and the miRNA regulatory network is not yet complete. This study employed Vascular cell Induction culture System Using Arabidopsis Leaves to profile the miRNome during TE differentiation in Arabidopsis (Arabidopsis thaliana) and established comprehensive miRNA co-expression networks functioning at the different stages of TE differentiation. Two negatively correlated modules exist in the miRNA networks, each exhibiting strong intra-module positive correlation and strong inter-module negative correlation. Thus, the 2 modules may play opposite roles in TE differentiation and vascular development. Indeed, we found that miR408 promotes cambium formation and TE differentiation, consistent with miR408 as a key node in the networks of fate determination and the initiation of TE differentiation. Additionally, we found that miR163 inhibits secondary cell wall formation and TE differentiation, corresponding to miR163 as a key node in the TE maturation network. Moreover, we discovered that the miRNA co-expression network in poplar (Populus tomentosa) xylem development is also composed of 2 negatively correlated modules that contain miRNAs orthologous to those in Arabidopsis. Therefore, the 2 negatively correlated modules of the miRNA co-expression network are likely conserved and fundamental to xylem TE differentiation. These results provide insights into microRNA regulation in plant development.

Amide conjugates of the jasmonate precursor cis-(+)-12-oxo-phytodienoic acid regulate its homeostasis during plant stress responses.

Jasmonates are a family of oxylipin phytohormones regulating plant development and growth and mediating "defense versus growth" responses. The upstream JA biosynthetic precursor cis-(+)-12-oxo-phytodienoic acid (cis-OPDA) acts independently of CORONATIVE INSENSITIVE 1-mediated JA signaling in several stress-induced and developmental processes. However, its perception and metabolism are only partially understood. An isoleucine analog of the biologically active JA-Ile, OPDA-Ile, was detected years ago in wounded leaves of flowering plants, opening up the possibility that conjugation of cis-OPDA to amino acids might be a relevant mechanism for cis-OPDA regulation. Here, we extended the analysis of amino acid conjugates of cis-OPDA and identified naturally occurring OPDA-Val, OPDA-Phe, OPDA-Ala, OPDA-Glu, and OPDA-Asp accumulating in response to biotic and abiotic stress in Arabidopsis (Arabidopsis thaliana). The OPDA amino acid conjugates displayed cis-OPDA-related plant responses in a JA-Ile-dependent manner. We also showed that the synthesis and hydrolysis of cis-OPDA amino acid conjugates are mediated by members of the amidosynthetase GRETCHEN HAGEN 3 and the amidohydrolase INDOLE-3-ACETYL-LEUCINE RESISTANT 1/ILR1-like families. Thus, OPDA amino acid conjugates function in the catabolism or temporary storage of cis-OPDA in stress responses instead of acting as chemical signals per se.

BIL7 enhances plant growth by regulating the transcription factor BIL1/BZR1 during brassinosteroid signaling‡.

Brassinosteroids (BRs) are plant steroid hormones that regulate plant development and environmental responses. BIL1/BZR1, a master transcription factor that regulates approximately 3000 genes in the BR signaling pathway, is transported to the nucleus from the cytosol in response to BR signaling; however, the molecular mechanism underlying this process is unknown. Here, we identify a novel BR signaling factor, BIL7, that enhances plant growth and positively regulates the nuclear accumulation of BIL1/BZR1 in Arabidopsis thaliana. BIL7-overexpressing plants were resistant to the BR biosynthesis inhibitor Brz and taller than wild-type (WT) plants were due to increased cell division. BIL7 is mainly localized to the plasma membrane, but during the early stages of cell growth, it was also localized to the nucleus. BIL7 was directly phosphorylated by the kinase BIN2, and nuclear localization of BIL7 was enhanced by the BIN2 inhibitor bikinin. BIL7 was found to bind to BIL1/BZR1, and nuclear accumulation of BIL1/BZR1 was strongly enhanced by BIL7 overexpression. Finally, double overexpression of BIL1/BZR1 and BIL7 led to greatly elongated hypocotyls in the presence of Brz. These findings suggest that BIL7 mediates nuclear accumulation of BIL1/BZR1, which activates inflorescence elongation in plants via BR signaling.

Identification of GiOMT gene family in Glycyrrhiza inflata bat and expression analysis under UV-B stresses

BackgroundO-Methyltransferase (OMTs) is a class of conserved multifunctional enzymes that play important roles in plant developmental regulation, hormone signaling, secondary metabolite synthesis and abiotic stress response. The GiOMT gene family has been identified and analyzed in species such as citrus, alfalfa, Populus and grape, but has not been reported in Glycyrrhiza inflata Bat.ResultsIn this study, we systematically identified and analyzed the GiOMT gene family of G. inflata by bioinformatics, and analyzed their physicochemical properties, conserved motifs, conserved structural domains, gene structures, phylogenetic relationships, chromosomal localization and fragment duplications, and the expression patterns of GiOMT genes in combination with transcriptomic data and qRT-PCR. The results showed that a total of 41 GiOMTs were identified in G. inflata, which were named GiOMT1 ~ GiOMT41 based on their chromosomal locations. Protein characterization showed that 29 GiOMT proteins were hydrophilic and 12 GiOMT proteins were hydrophobic. Subcellular predicted localization revealed that most GiOMT proteins localized in the cytoplasm and chloroplasts. Phylogenetic relationships showed that the OMT genes of three species, G. inflata, Arabidopsis and alfalfa, were distributed in three taxa, while the GiOMT genes were distributed in taxa I and II. Promoters of GiOMT genes contained light responsive element and many hormone responsive elements. The expression levels of GiOMT genes under UV-B stress were varied, indicating that GiOMT gene was in response to abiotic stresses in G. inflata.ConclusionIn this study, we investigated the genome-wide identification, structure, evolution and expression analysis of the GiOMT gene in G. inflata. The basal sequence of GiOMT genes was highly conserved throughout the evolutionary history of G. inflata. Most of the GiOMT genes were highly expressed in roots and were involved in the response to UV-B stress. The GiOMT genes may lead to the accumulation of flavonoids and enhancement of G. inflata quality and drug activity in G. inflata under UV-B radiation.

MYB proteins: Versatile regulators of plant development, stress responses, and secondary metabolite biosynthetic pathways.

MYB proteins are ubiquitous in nature, regulating key aspects of plant growth and development. Although MYB proteins are known for regulating genes involved in secondary metabolite biosynthesis, particularly phenylpropanoids, their roles in terpenoid, glucosinolate, and alkaloid biosynthesis remain less understood. This review explores the structural and functional differences between activator and repressor MYB proteins along with their roles in plant growth, development, stress responses, and secondary metabolite production. MYB proteins serve as central hubs in protein-protein interaction networks that regulate expression of numerous genes involved in the adaptation of plants to varying environmental conditions. Thus, we also highlight key interacting partners of MYB proteins and their roles in these adaptation mechanisms. We further discuss the mechanisms regulating MYB proteins, including autoregulation, epigenetics, and post-transcriptional and post-translational modifications. Overall, we propose MYB proteins as versatile regulators for improving plant traits, stress responses, and secondary metabolite production.

The PpPep2-Triggered PTI-like Response in Peach Trees Is Mediated by miRNAs.

Plant diseases diminish crop yields and put the world's food supply at risk. Plant elicitor peptides (Peps) are innate danger signals inducing defense responses both naturally and after external application onto plants. Pep-triggered defense networks are compatible with pattern-triggered immunity (PTI). Nevertheless, in complex regulatory pathways, there is crosstalk among different signaling pathways, involving noncoding RNAs in the natural response to pathogen attack. Here, we used Prunus persica, PpPep2 and a miRNA-Seq approach to show for the first time that Peps regulate, in parallel with a set of protein-coding genes, a set of plant miRNAs (~15%). Some PpPep2-regulated miRNAs have been described to participate in the response to pathogens in various plant-pathogen systems. In addition, numerous predicted target mRNAs of PpPep2-regulated miRNAs are themselves regulated by PpPep2 in peach trees. As an example, peach miRNA156 and miRNA390 probably have a role in plant development regulation under stress conditions, while others, such as miRNA482 and miRNA395, would be involved in the regulation of resistance (R) genes and sulfate-mediated protection against oxygen free radicals, respectively. This adds to the established role of Peps in triggering plant defense systems by incorporating the miRNA regulatory network and to the possible use of Peps as sustainable phytosanitary products.

Unlocking the small RNAs: local and systemic modulators for advancing agronomic enhancement.

Small regulatory RNAs (sRNAs) are essential regulators of gene expression across a wide range of organisms to precisely modulate gene activity based on sequence-specific recognition. In model plants like Arabidopsis thaliana, extensive research has primarily concentrated on 21 to 24-nucleotide (nt) sRNAs, particularly microRNAs (miRNAs). Recent advancements in cell and tissue isolation techniques, coupled with advanced sequencing technologies, are revealing a diverse array of preciously uncharacterized sRNA species. These include previously novel structural RNA fragments as well as numerous cell- and tissue-specific sRNAs that are active during distinct developmental stages, thereby enhancing our understanding of the precise and dynamic regulatory roles of sRNAs in plant development regulation. Additionally, a notable feature of sRNAs is their capacity for amplification and movement between cells and tissues, which facilitates long-distance communication-an adaptation critical to plants due to their sessile nature. In this review, we will discuss the classification and mechanisms of action of sRNAs, using legumes as a primary example due to their essential engagement for the unique organ establishment of root nodules and long-distance signaling, and further illustrating the potential applications of sRNAs in modern agricultural breeding and environmentally sustainable plant protection strategies.

Ascorbate, as a versatile regulator of plant development: practical implications for enhancing crop productivity, quality, and postharvest storage

Ascorbate, as a versatile regulator of plant development: practical implications for enhancing crop productivity, quality, and postharvest storage

Genome-wide identification of TaeGRASs responsive to biotic stresses and functional analysis of TaeSCL6 in wheat resistance to powdery mildew

BackgroundPowdery mildew is a devastating fungal disease that poses a significant threat to wheat yield and quality worldwide. Identifying resistance genes is highly advantageous for the molecular breeding of resistant cultivars. GRAS proteins are important transcription factors that regulate plant development and stress responses. Nonetheless, their roles in wheat-pathogen interactions remain poorly understood.ResultsIn this study, we used bioinformatics tools to identify and analyze wheat GRAS family genes responsive to biotic stresses and elucidated the function of TaeSCL6 within this family. A total of 179 GRAS genes in wheat were unevenly distributed on 7 chromosomes, and classified into 12 subfamilies based on phylogenetic relationship analysis. Gene duplication analysis revealed 13 pairs of tandem repeats and 142 pairs of segmental duplications, which may account for the rapid expansion of the wheat GRAS family. Expression pattern analysis revealed that 75% of the expressed TaeGRAS genes are responsive to biotic stresses. Few studies have focused on the roles of HAM subfamily genes. Consequently, we concentrated our analysis on the members of the HAM subfamily. Fourteen motifs were identified in the HAM family proteins from both Triticeae species and Arabidopsis, indicating that these motifs were highly conserved during evolution. Promoter analysis indicated that the promoters of HAM genes contain several cis-regulatory elements associated with hormone response, stress response, light response, and growth and development. Both qRT-PCR and RNA-seq data analyses demonstrated that TaeSCL6 responds to Blumeria graminis infection. Therefore, we investigated the role of TaeSCL6 in regulating wheat resistance via RNA interference and barley stripe mosaic virus induced gene silencing. Wheat plants with silenced TaeSCL6 exhibited increased susceptibility to powdery mildew.ConclusionsIn summary, this study not only validates the positive role of TaeSCL6 in wheat resistance to powdery mildew, but also provides candidate gene resources for future breeding of disease-resistance wheat cultivars.

Role of CLE peptide signaling in root-knot nematode parasitism of plants.

We summarize recent findings that have provided new insights into the mechanisms underlying CLE signaling systems in the regulation of plant development and phytonematode interactions. CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) peptides are short sequences consisting of 12 or 13 amino acids characterized by hydroxylated proline residues, and their presence has been demonstrated in various plant species and phytonematodes across multiple paralogous genes. Here, we review recently conducted research to understanding the signaling pathway of CLE peptide during plant development and infection caused by phytonematodes. Cell-to-cell communication is important for the coherent functioning of living organisms. CLE peptides combined with their specific transmembrane receptors to induce downstream intracellular signaling pathways shows divergent modes of action in many developmental processes in variable species. Moreover, CLE peptide was also involved in plant disease mechanism caused by various plant parasitic nematodes.

The E3 ubiquitin ligase COP1 and transcription factors HY5 and RHD6 integrate light signaling and root hair development.

Light signaling plays a substantial role in regulating plant development, including the differentiation and elongation of single-celled tissue. However, the identity of the regulatory machine that affects light signaling on root hair cell (RHC) development remains unclear. Here, we investigated how darkness inhibits differentiation and elongation of RHC in Arabidopsis (Arabidopsis thaliana). We found that light promotes the growth and development of RHC. RNA-seq analysis showed that light signaling regulates the differentiation of RHC by promoting the expression of specific genes in the root epidermis associated with cell wall remodeling, JA, auxin, and ethylene signaling pathways. Together, these genes integrate light and phytohormone signals with root hair development. Our investigation also revealed that the core light signal factor ELONGATED HYPOCOTYL 5 (HY5) directly interacts with the key root hair development factor ROOT HAIR DEFECTIVE6 (RHD6), which promotes the transcription of RSL4. However, CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1) repressed RHD6 function through the COP1-HY5 complex. Our genetic studies confirm associations between RHD6, HY5, and COP1, indicating that RHD6 largely depends on HY5 for root hair development. Ultimately, our work suggests a central COP1-HY5-RHD6 regulatory module that integrates light signaling and root hair development with several downstream pathways, offering perspectives to decipher single-celled root hair development.

PWOs repress gene transcription by regulating chromatin structures in Arabidopsis.

PWWP-DOMAIN INTERACTOR OF POLYCOMBS (PWO) family proteins play a vital role in regulating plant development. However, the molecular mechanisms of how PWOs regulate chromatin structure is elusive. Our data show that the PWO1 binding sites are enriched with positive modifications but exclusive with H3K27me3. Moreover, PWO1 binds to the H3K27me3-enriched compartment domain (H3K27me3-CD) boundary regions, and functions to maintain the boundary strength. Meanwhile, we found that PWOs and Polycomb repressive complex 2 (PRC2) function parallelly in maintaining H3K27me3-CDs' structure. Loss of either PWOs or PRC2 leads to H3K27me3-CD strength reduction, B to A compartment switching as well as the H3K27me3-CD relocating away from the nuclear periphery. Additionally, PWOs and lamin-like proteins collaborate to regulate multiple chromatin structures to repress gene transcription within H3K27me3-CDs. We conclude that PWOs maintain H3K27me3-CDs' repressive state and regulate their spatial position in the nucleus.