Histone methylation readers MRG1/2 interact with PIF4 to promote thermomorphogenesis in Arabidopsis.

Warm ambient conditions induce thermomorphogenesis and affect plant growth and development. However, the chromatin regulatory mechanisms involved in thermomorphogenesis remain largely obscure. In this study, we show that the histone methylation readers MORF-related gene 1 and 2 (MRG1/2) are required to promote hypocotyl elongation in response to warm ambient conditions. A transcriptome sequencing analysis indicates that MRG1/2 and phytochrome interacting factor 4 (PIF4) coactivate a number of thermoresponsive genes, including YUCCA8, which encodes a rate-limiting enzyme in the auxin biosynthesis pathway. Additionally, MRG2 physically interacts with PIF4 to bind to thermoresponsive genes and enhances the H4K5 acetylation of the chromatin of target genes in a PIF4-dependent manner. Furthermore, MRG2 competes with phyB for binding to PIF4 and stabilizes PIF4 in planta. Our study indicates that MRG1/2 activate thermoresponsive genes by inducing histone acetylation and stabilizing PIF4 in Arabidopsis.

Trifluoperazine activates AMPK / mTOR / ULK1 signaling pathway to induce mitophagy in osteosarcoma cells.

Osteosarcoma is a prevalent kind of primary bone malignancy. Trifluoperazine, as an antipsychotic drug, has anti-tumor activity against a variety of cancers. Nevertheless, the impact of trifluoperazine on osteosarcoma is unclear. Our investigation aimed to explore the mechanism of trifluoperazine's effect on osteosarcoma. We found that trifluoperazine inhibited 143B and U2-OS osteosarcoma cell proliferation in a method based on the dose. Furthermore, it was shown that trifluoperazine induced the accumulation of reactive oxygen species (ROS) to cause mitochondrial damage and induced mitophagy in osteosarcoma cells. Finally, combined with RNA-seq results, we first demonstrated the AMPK/mTOR/ULK1 signaling pathway as a potential mechanism of trifluoperazine-mediated mitophagy in osteosarcoma cells and can be suppressed by AMPK inhibitor Compound C.

Cannabinoids improve mitochondrial function in skeletal muscle of exhaustive exercise training rats by inhibiting mitophagy through the PINK1/PARKIN and BNIP3 pathways.

Cannabidiol (CBD) is a pure natural phytocannabinoid derived from cannabis that has anti-inflammatory, antiapoptotic and antioxidative stress abilities. In recent years, an increasing number of studies have reported the regulatory effect of CBD on skeletal muscle injury induced by exercise, but its mechanism is still unclear. Mitochondria are the main organelles responsible for the energy supply within eukaryotic cells, and their function has been closely linked to cellular health. Moderate exercise improves mitochondrial function, but the excessive exercise has a negative impact on mitochondria. Therefore, we speculate that CBD may promote exercise induced skeletal muscle cell damage by improving mitochondrial function. In this study, by establishing an animal model of exhaustive exercise training in rats, the protective effect of CBD on skeletal muscle mitochondrial structure and function was elaborated, and the possible molecular mechanism was discussed based on transcriptomics. Our results indicate that skeletal muscle mitochondrial structure and function were improved after CBD intervention. GO and KEGG pathway enrichment analysis showed that exhaustive exercise training induced mitochondrial dysfunction in skeletal muscle is associated with excessive autophagy/mitophagy, the signaling pathways involved in FOXO3 and GABARAPL1 may play important roles. After CBD intervention, the protein expression of PINK1, PARKIN and BNIP3 was down-regulated, indicating that CBD may improve the mitochondrial function by inhibiting mitophagy through the PINK1/PARKIN and BNIP3 pathway.

Discovery of a novel dual-targeting D-peptide to block CD24/Siglec-10 and PD-1/PD-L1 interaction and synergize with radiotherapy for cancer immunotherapy.

BACKGROUND: Aside from immune checkpoint inhibitors targeting programmed cell death protein 1 (PD-1) and programmed death ligand 1 (PD-L1), intervention of CD47/Sirpα mediated 'don't eat me' signal between macrophage and tumor cell is considered as a promising therapeutic approach for cancer immunotherapy. Compared with CD47, the novel immune checkpoint CD24/Siglec-10 can also deliver 'don't eat me' signal and CD24 shows much lower expression level in normal tissue which might avoid unwanted side effects. METHODS: Cell-based phage display biopanning and D-amino acid modification strategy were used to identify the CD24/Siglec-10 blocking peptide. Cell-based blocking assay and microscale thermophoresis assay were used to validate the blocking and binding activities of the peptide. Phagocytosis and co-culture assays were used to explore the in vitro function of the peptide. Flow cytometry was performed to assess the immune microenvironment after the peptide treatment in vivo. RESULTS: A CD24/Siglec-10 blocking peptide (CSBP) with hydrolysis-resistant property was identified. Surprisingly, we found that CSBP could not only block the interaction of CD24/Siglec-10 but also PD-1/PD-L1. CSBP could induce the phagocytosis of tumor cell by both the macrophages and monocytic myeloid-derived suppressor cells (M-MDSCs), which can further activate CD8+ T cells. Besides, combination of radiotherapy and CSBP synergistically reduced tumor growth and altered the tumor microenvironment in both anti-PD-1-responsive MC38 and anti-PD-1-resistant 4T1 tumor models. CONCLUSIONS: In summary, this is the first CD24/Siglec-10 blocking peptide which blocked PD-1/PD-L1 interaction as well, functioned via enhancing the phagocytosis of tumor cells by macrophages and M-MDSCs, and elevating the activity of CD8+ T cells for cancer immunotherapy.

Histone chaperones AtChz1A and AtChz1B are required for H2A.Z deposition and interact with the SWR1 chromatin-remodeling complex in Arabidopsis thaliana.

The histone variant H2A.Z plays key functions in transcription and genome stability in all eukaryotes ranging from yeast to human, but the molecular mechanisms by which H2A.Z is incorporated into chromatin remain largely obscure. Here, we characterized the two homologs of yeast Chaperone for H2A.Z-H2B (Chz1) in Arabidopsis thaliana, AtChz1A and AtChz1B. AtChz1A/AtChz1B were verified to bind to H2A.Z-H2B and facilitate nucleosome assembly in vitro. Simultaneous knockdown of AtChz1A and AtChz1B, which exhibit redundant functions, led to a genome-wide reduction in H2A.Z and phenotypes similar to those of the H2A.Z-deficient mutant hta9-1hta11-2, including early flowering and abnormal flower morphologies. Interestingly, AtChz1A was found to physically interact with ACTIN-RELATED PROTEIN 6 (ARP6), an evolutionarily conserved subunit of the SWR1 chromatin-remodeling complex. Genetic interaction analyses showed that atchz1a-1atchz1b-1 was hypostatic to arp6-1. Consistently, genome-wide profiling analyses revealed partially overlapping genes and fewer misregulated genes and H2A.Z-reduced chromatin regions in atchz1a-1atchz1b-1 compared with arp6-1. Together, our results demonstrate that AtChz1A and AtChz1B act as histone chaperones to assist the deposition of H2A.Z into chromatin via interacting with SWR1, thereby playing critical roles in the transcription of genes involved in flowering and many other processes.

PIF7-mediated epigenetic reprogramming promotes the transcriptional response to shade in Arabidopsis.

For shade-intolerant plants, changes in light quality through competition from neighbors trigger shade avoidance syndrome (SAS): a series of morphological and physiological adaptations that are ultimately detrimental to plant health and crop yield. Phytochrome-interacting factor 7 (PIF7) is a major transcriptional regulator of SAS in Arabidopsis; however, how it regulates gene expression is not fully understood. Here, we show that PIF7 directly interacts with the histone chaperone anti-silencing factor 1 (ASF1). The ASF1-deprived asf1ab mutant showed defective shade-induced hypocotyl elongation. Histone regulator homolog A (HIRA), which mediates deposition of the H3.3 variant into chromatin, is also involved in SAS. RNA/ChIP-sequencing analyses identified the role of ASF1 in the direct regulation of a subset of PIF7 target genes. Furthermore, shade-elicited gene activation is accompanied by H3.3 enrichment, which is mediated by the PIF7-ASF1-HIRA regulatory module. Collectively, our data reveal that PIF7 recruits ASF1-HIRA to increase H3.3 incorporation into chromatin to promote gene transcription, thus enabling plants to effectively respond to environmental shade.

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Analyzing the pattern of altitudinal variation in the leaf traits and their networks of a particular tree species of similar age and its influencing factors could contribute to understanding the impacts of environmental factors on leaf traits and excluding the interference of genetic factors. We investigated the stomatal, structural, chemical, and vein traits of Daphniphyllum macropodum leaves in middle-aged forests, following the altitudinal gradient (1100, 1500, and 1900 m) on Mao'er Mountain. The objectives of this study were to reveal patterns in leaf trait and leaf trait networks variation, the life strategy of the tree species, and the major environmental factors affecting the altitudinal variations. The results showed that leaf area, specific leaf area, leaf thickness, leaf dry matter content, chlorophyll content, nitrogen content, phosphorus content, C:N, C:P, vein density, and vein diameter varied significantly across altitudes. Mean annual temperature and total radiation explained 42.1% and 16.2% of leaf-trait variation, respectively. They served as key environmental factors driving the altitudinal variation in leaf traits. Mean annual temperature exhibited the greatest influence on leaf area (R2=0.73), and total radiation exerted the most prominent effect on leaf thickness (R2=0.72). Both relationships were significantly positive. D. macropodum exhibited low leaf nitrogen and phosphorus at the low altitude of 1100 m, and the overall and local trait networks were loose, adopting a conservative resource strategy. At the medium altitude of 1500 m, leaf nutrient contents were relatively high. The overall network of leaf traits was tightly connected and local network was loose. By enhancing the dependency among leaf traits, and improving phosphorus utilization efficiency, D. macropodum could cope with competition in deciduous forests and adopt resource acquisition strategies. Further, at the highest altitude of 1900 m, D. macropodum had relatively large leaf thickness, chlorophyll content, and leaf dry matter content, but relatively small leaf area. The local network connections were tight while the overall network looseness, indicating a resource conserving strategy. The trade-off relationship between C:P and leaf phosphorus content was closely related to phosphorus use efficiency, and its variation was an important indicator for identifying life strategies of D. macropodum in different altitudes.

Identification of natural product 3, 5-diiodotyrosine as APOBEC3B inhibitor to prevent somatic mutation accumulation and cancer progression.

BACKGROUND: The development of cancer is largely dependent on the accumulation of somatic mutations, indicating the potential to develop cancer chemoprevention agents targeting mutation drivers. However, ideal cancer chemoprevention agents that can effectively inhibit the mutation drivers have not been identified yet. METHODS: The somatic mutation signatures and expression analyses of APOBEC3B were performed in patient with pan-cancer. The computer-aided screening and skeleton-based searching were performed to identify natural products that can inhibit the activity of APOBEC3B. 4-nitroquinoline-1-oxide (4-NQO)-induced spontaneous esophageal squamous cell carcinoma (ESCC) and azoxymethane/dextran sulfate sodium (AOM/DSS)-induced spontaneous colon cancer mouse models were conducted to investigate the influences of APOBEC3B inhibitor on the prevention of somatic mutation accumulation and cancer progression. RESULTS: Here, we discovered that the cytidine deaminase APOBEC3B correlated somatic mutations were widely observed in a variety of cancers, and its overexpression indicated poor survival. SMC247 (3, 5-diiodotyrosine), as a source of kelp iodine without side effects, could strongly bind APOBEC3B (KD=65 nM) and effectively inhibit its deaminase activity (IC50=1.69 µM). Interestingly, 3, 5-diiodotyrosine could significantly reduce the clusters of mutations, prevent the precancerous lesion progression, and prolong the survival in 4-NQO-induced spontaneous ESCC and AOM/DSS-induced spontaneous colon cancer mouse models. Furthermore, 3, 5-diiodotyrosine could reduce colitis, increase the proportion and function of T lymphocytes via IL-15 in tumor microenvironment. The synergistic cancer prevention effects were observed when 3, 5-diiodotyrosine combined with PD-1/PD-L1 blockade. CONCLUSIONS: This is the first prove-of-concept study to elucidate that the natural product 3, 5-diiodotyrosine could prevent somatic mutation accumulation and cancer progression through inhibiting the enzymatic activity of APOBEC3B. In addition, 3, 5-diiodotyrosine could reduce the colitis and increase the infiltration and function of T lymphocytes via IL-15 in tumor microenvironment. 3, 5-diiodotyrosine combined with PD-1/PD-L1 blockade could elicit synergistic cancer prevention effects, indicating a novel strategy for both prevent the somatic mutation accumulation and the immune-suppressive microenvironment exacerbation.

Histone modification and chromatin remodeling in plant response to pathogens.

As sessile organisms, plants are constantly exposed to changing environments frequently under diverse stresses. Invasion by pathogens, including virus, bacterial and fungal infections, can severely impede plant growth and development, causing important yield loss and thus challenging food/feed security worldwide. During evolution, plants have adapted complex systems, including coordinated global gene expression networks, to defend against pathogen attacks. In recent years, growing evidences indicate that pathogen infections can trigger local and global epigenetic changes that reprogram the transcription of plant defense genes, which in turn helps plants to fight against pathogens. Here, we summarize up plant defense pathways and epigenetic mechanisms and we review in depth current knowledge's about histone modifications and chromatin-remodeling factors found in the epigenetic regulation of plant response to biotic stresses. It is anticipated that epigenetic mechanisms may be explorable in the design of tools to generate stress-resistant plant varieties.

Structural insights into molecular mechanism for N6-adenosine methylation by MT-A70 family methyltransferase METTL4.

METTL4 belongs to a subclade of MT-A70 family members of methyltransferase (MTase) proteins shown to mediate N6-adenosine methylation for both RNA and DNA in diverse eukaryotes. Here, we report that Arabidopsis METTL4 functions as U2 snRNA MTase for N6-2'-O-dimethyladenosine (m6Am) in vivo that regulates flowering time, and specifically catalyzes N6-methylation of 2'-O-methyladenosine (Am) within a single-stranded RNA in vitro. The apo structures of full-length Arabidopsis METTL4 bound to S-adenosyl-L-methionine (SAM) and the complex structure with an Am-containing RNA substrate, combined with mutagenesis and in vitro enzymatic assays, uncover a preformed L-shaped, positively-charged cavity surrounded by four loops for substrate binding and a catalytic center composed of conserved residues for specific Am nucleotide recognition and N6-methylation activity. Structural comparison of METTL4 with the mRNA m6A enzyme METTL3/METTL14 heterodimer and modeling analysis suggest a catalytic mechanism for N6-adenosine methylation by METTL4, which may be shared among MT-A70 family members.

Structural insights into partner selection for MYB and bHLH transcription factor complexes.

MYB and basic helix-loop-helix (bHLH) transcription factors form complexes to regulate diverse metabolic and developmental processes in plants. However, the molecular mechanisms responsible for MYB-bHLH interaction and partner selection remain unclear. Here, we report the crystal structures of three MYB-bHLH complexes (WER-EGL3, CPC-EGL3 and MYB29-MYC3), uncovering two MYB-bHLH interaction modes. WER and CPC are R2R3- and R3-type MYBs, respectively, but interact with EGL3 through their N-terminal R3 domain in a similar mode. A single amino acid of CPC, Met49, is crucial for competition with WER to interact with EGL3. MYB29, a R2R3-type MYB transcription factor, interacts with MYC3 by its C-terminal MYC-interaction motif. The WER-EGL3 and MYB29-MYC3 binding modes are widely applied among MYB-bHLH complexes in Arabidopsis and evolve independently in plants.

Arabidopsis CHROMATIN REMODELING 19 acts as a transcriptional repressor and contributes to plant pathogen resistance.

Chromatin remodelers act in an ATP-dependent manner to modulate chromatin structure and thus genome function. Here, we report that the Arabidopsis (Arabidopsis thaliana) remodeler CHROMATIN REMODELING19 (CHR19) is enriched in gene body regions, and its depletion causes massive changes in nucleosome position and occupancy in the genome. Consistent with these changes, an in vitro assay verified that CHR19 can utilize ATP to slide nucleosomes. A variety of inducible genes, including several important genes in the salicylic acid (SA) and jasmonic acid (JA) pathways, were transcriptionally upregulated in the chr19 mutant under normal growth conditions, indicative of a role of CHR19 in transcriptional repression. In addition, the chr19 mutation triggered higher susceptibility to the JA pathway-defended necrotrophic fungal pathogen Botrytis cinerea, but did not affect the growth of the SA pathway-defended hemibiotrophic bacterial pathogen Pseudomonas syringae pv. tomato DC3000. Expression of CHR19 was tissue-specific and inhibited specifically by SA treatment. Such inhibition significantly decreased the local chromatin enrichment of CHR19 at the associated SA pathway genes, which resulted in their full activation upon SA treatment. Overall, our findings clarify CHR19 to be a novel regulator acting at the chromatin level to impact the transcription of genes underlying plant resistance to different pathogens.

The patterns of distant metastasis and prognostic factors in patients with primary metastatic Ewing sarcoma of the bone.

BACKGROUND: Ewing sarcoma (ES) of bone is accounting for the second most common type of primary bone cancer in children and adolescents. However, the patterns of distant metastasis (DM) and the effect of the sites of DM on survival outcomes were not investigated. AIMS: This study aimed to investigate the patterns of DM and the prognostic factors related to outcomes in primary metastatic ES of the bone. METHODS: Patients who were diagnosed with primary metastatic ES between 2010 and 2018 were identified from the Surveillance, Epidemiology, and End Results database. Kaplan-Meier analysis, log-rank tests, and Cox proportional-hazards regression models were used for statistical analyses. RESULTS: We identified 277 patients in this study and 95.3% of them (n = 264) receiving chemotherapy. A total of 371 sites of DM were observed. Lung was the most common distant metastatic site (n = 182, 49.1%), followed by bone (n = 139, 37.5%), distant lymph node (n = 26, 7.0%), liver (n = 14, 3.8%), and brain (n = 10, 2.7%). Three-year cause-specific survival (CSS) was 56.1% in the entire cohort. Older age (hazard ratio [HR] 2.210, P < 0.001) and bone metastasis (HR 1.903, P = 0.002) were the independent prognostic factors associated with inferior CSS. Similar results were found in those with bone-only metastasis (n = 80) or lung-only metastasis (n = 117), which showed that patients with bone-only metastasis had an inferior CSS compared to those with metastases only to the lung (HR 1.926, P = 0.005). CONCLUSIONS: Lung and bone are the most frequently distant metastatic sites in patients with primary metastatic ES of bone. Bone metastasis is an independent risk factor for inferior survival.

H3K36 methyltransferase SDG708 enhances drought tolerance by promoting abscisic acid biosynthesis in rice.

Chromatin modifications play important roles in plant adaptation to abiotic stresses, but the precise function of histone H3 lysine 36 (H3K36) methylation in drought tolerance remains poorly evaluated. Here, we report that SDG708, a specific H3K36 methyltransferase, functions as a positive regulator of drought tolerance in rice. SDG708 promoted abscisic acid (ABA) biosynthesis by directly targeting and activating the crucial ABA biosynthesis genes NINE-CIS-EPOXYCAROTENOID DIOXYGENASE 3 (OsNCED3) and NINE-CIS-EPOXYCAROTENOID DIOXYGENASE 5 (OsNCED5). Additionally, SDG708 induced hydrogen peroxide accumulation in the guard cells and promoted stomatal closure to reduce water loss. Overexpression of SDG708 concomitantly enhanced rice drought tolerance and increased grain yield under normal and drought stress conditions. Thus, SDG708 is potentially useful as an epigenetic regulator in breeding for grain yield improvement.

AtINO80 represses photomorphogenesis by modulating nucleosome density and H2A.Z incorporation in light-related genes.

Photomorphogenesis is a critical developmental process bridging light-regulated transcriptional reprogramming with morphological changes in organisms. Strikingly, the chromatin-based transcriptional control of photomorphogenesis remains poorly understood. Here, we show that the Arabidopsis (Arabidopsis thaliana) ortholog of ATP-dependent chromatin-remodeling factor AtINO80 represses plant photomorphogenesis. Loss of AtINO80 inhibited hypocotyl cell elongation and caused anthocyanin accumulation. Both light-induced genes and dark-induced genes were affected in the atino80 mutant. Genome-wide occupancy of the H2A.Z histone variant and levels of histone H3 were reduced in atino80 In particular, AtINO80 bound the gene body of ELONGATED HYPOCOTYL 5 (HY5), resulting in lower chromatin incorporations of H2A.Z and H3 at HY5 in atino80 Genetic analysis revealed that AtINO80 acts in a phytochrome B- and HY5-dependent manner in the regulation of photomorphogenesis. Together, our study elucidates a mechanism wherein AtINO80 modulates nucleosome density and H2A.Z incorporation and represses the transcription of light-related genes, such as HY5, to fine tune plant photomorphogenesis.

OsChz1 acts as a histone chaperone in modulating chromatin organization and genome function in rice.

While the yeast Chz1 acts as a specific histone-chaperone for H2A.Z, functions of CHZ-domain proteins in multicellular eukaryotes remain obscure. Here, we report on the functional characterization of OsChz1, a sole CHZ-domain protein identified in rice. OsChz1 interacts with both the canonical H2A-H2B dimer and the variant H2A.Z-H2B dimer. Within crystal structure the C-terminal region of OsChz1 binds H2A-H2B via an acidic region, pointing to a previously unknown recognition mechanism. Knockout of OsChz1 leads to multiple plant developmental defects. At genome-wide level, loss of OsChz1 causes mis-regulations of thousands of genes and broad alterations of nucleosome occupancy as well as reductions of H2A.Z-enrichment. While OsChz1 associates with chromatin regions enriched of repressive histone marks (H3K27me3 and H3K4me2), its loss does not affect the genome landscape of DNA methylation. Taken together, it is emerging that OsChz1 functions as an important H2A/H2A.Z-H2B chaperone in dynamic regulation of chromatin for higher eukaryote development.

NAP1-Related Protein 1 (NRP1) has multiple interaction modes for chaperoning histones H2A-H2B.

Nucleosome Assembly Protein 1 (NAP1) family proteins are evolutionarily conserved histone chaperones that play important roles in diverse biological processes. In this study, we determined the crystal structure of Arabidopsis NAP1-Related Protein 1 (NRP1) complexed with H2A-H2B and uncovered a previously unknown interaction mechanism in histone chaperoning. Both in vitro binding and in vivo plant rescue assays proved that interaction mediated by the N-terminal α-helix (αN) domain is essential for NRP1 function. In addition, the C-terminal acidic domain (CTAD) of NRP1 binds to H2A-H2B through a conserved mode similar to other histone chaperones. We further extended previous knowledge of the NAP1-conserved earmuff domain by mapping the amino acids of NRP1 involved in association with H2A-H2B. Finally, we showed that H2A-H2B interactions mediated by αN, earmuff, and CTAD domains are all required for the effective chaperone activity of NRP1. Collectively, our results reveal multiple interaction modes of a NAP1 family histone chaperone and shed light on how histone chaperones shield H2A-H2B from nonspecific interaction with DNA.

Effects of vegetation restoration and environmental factors on understory vascular plants in a typical karst ecosystem in southern China.

Understory vegetation is an important component in most forest ecosystems. It is very important for soil and water conservation in karst region, study on understory will provide valuable information for understanding the interaction mechanism between understory flora and karst environment. Thirty-two plots were sampled in three vegetation types along with a restoration gradient (shrubland, forest-shrub transition, and mixed-species broadleaf forest) in typical karst mountains in Southwest Guangxi, China. Overstory trees, understory vascular plants, soil nutrients, and topographic factors were recorded in each 400-m2 plot. Multivariate statistics were used, including the multi-response permutation procedure (MRPP), indicator species analysis, and canonical correlation analysis (CCA). MRPP showed understory species composition significantly differed among the three vegetation types, with the greatest difference between the shrubland and the mixed forest. Twenty-one understory species were identified as significant indicator species, with 13 species being identified as indicators of the shrubland, two of forest-shrub transition, and six of the mixed forest. Light-demanding herbaceous seed plants were common in shrubland, while shade-tolerant calcicole assembled under the mixed forest. Forward selection of CCA ordination revealed that understory plant distribution was most strongly influenced by elevation, followed by soil pH, the concentration of total potassium and exchangeable calcium, slope aspect, slope degree, and the concentration of available potassium. The result reveals that vegetation types affect understory species composition by modifying understory environments. Elevation affects the spatial distribution of vegetation and soil factors, and then the understory plants. Meanwhile, soil Ca content also plays a key role in the understory species distribution. Understory diversity increased with increasing canopy structure complexity from shrubland to mixed-species forest. Thus, it is necessary to take measures to promote natural vegetation restoration and to protect the mixed forests in degraded karst areas.

The histone methylation readers MRG1/MRG2 and the histone chaperones NRP1/NRP2 associate in fine-tuning Arabidopsis flowering time.

Histones are highly basic proteins involved in packaging DNA into chromatin, and histone modifications are fundamental in epigenetic regulation in eukaryotes. Among the numerous chromatin modifiers identified in Arabidopsis (Arabidopsis thaliana), MORF-RELATED GENE (MRG)1 and MRG2 have redundant functions in reading histone H3 lysine 36 trimethylation (H3K36me3). Here, we show that MRG2 binds histone chaperones belonging to the NUCLEOSOME ASSEMBLY PROTEIN 1 (NAP1) family, including NAP1-RELATED PROTEIN (NRP)1 and NRP2. Characterization of the loss-of-function mutants mrg1 mrg2, nrp1 nrp2 and mrg1 mrg2 nrp1 nrp2 revealed that MRG1/MRG2 and NRP1/NRP2 regulate flowering time through fine-tuning transcription of floral genes by distinct molecular mechanisms. In particular, the physical interaction between NRP1/NRP2 and MRG1/MRG2 inhibited the binding of MRG1/MRG2 to the transcription factor CONSTANS (CO), leading to a transcriptional repression of FLOWERING LOCUS T (FT) through impeded H4K5 acetylation (H4K5ac) within the FT chromatin. By contrast, NRP1/NRP2 and MRG1/MRG2 act together, likely in a multiprotein complex manner, in promoting the transcription of FLOWERING LOCUS C (FLC) via an increase of both H4K5ac and H3K9ac in the FLC chromatin. Because the expression pattern of FLC represents the major category of differentially expressed genes identified by genome-wide RNA-sequencing analysis in the mrg1 mrg2, nrp1 nrp2 and mrg1 mrg2 nrp1 nrp2 mutants, it is reasonable to speculate that the NRP1/NRP2-MRG1/MRG2 complex may be involved in transcriptional activation of genes beyond FLC and flowering time control.

MRG1/2 histone methylation readers and HD2C histone deacetylase associate in repression of the florigen gene FT to set a proper flowering time in response to day-length changes.

Day-length changes represent an important cue for modulating flowering time. In Arabidopsis, the expression of the florigen gene FLOWERING LOCUS T (FT) exhibits a 24-h circadian rhythm under long-day (LD) conditions. Here we focus on the chromatin-based mechanism regarding the control of FT expression. We conducted co-immunoprecipitation assays along with LC-MS/MS analysis and identified HD2C histone deacetylase as the binding protein of the H3K4/H3K36 methylation reader MRG2. HD2C and MRG1/2 regulate flowering time under LD conditions, but not under short-day conditions. Moreover, HD2C functions as an effective deacetylase in planta, mainly targeting H3K9ac, H3K23ac and H3K27ac. At dusk, HD2C is recruited to FT to deacetylate histones and repress transcription in an MRG1/2-dependent manner. More importantly, HD2C competes with CO for the binding of MRG2, and the accumulation of HD2C at the FT locus occurs at the end of the day. Our findings not only reveal a histone deacetylation mechanism contributing to prevent FT overexpression and precocious flowering, but also support the model in which the histone methylation readers MRG1/2 provide a platform on chromatin for connecting regulatory factors involved in activating FT expression in response to daylight and decreasing FT expression around dusk under long days.