Phosphatidylethanolamine-binding Protein Research Articles

The pepper phosphatidyl ethanolamine-binding proteins CaMFT02 and CaMFT03 have distinct roles in responses to drought and salt stresses

Plants have established defense systems to overcome diverse environmental threats. Abscisic acid (ABA) is one of the representative phytohormones that involves in stress response. Here, we isolated and verified the function of pepper CaMFT (Capsicum annuum Mother of FT and TFL1) genes, which negatively modulate ABA-mediated drought and salt tolerance. MFT is part of the phosphatidyl ethanolamine-binding protein (PEBP) family, which also contains the flowering locus T (FT) and terminal flower 1 (TFL1) subfamilies. While FT and TFL1 have been identified as regulators of flowering time in Arabidopsis, the biological function of MFT is largely unknown. To elucidate how CaMFT02 acts in ABA signaling, along with drought and salt stress responses, we conducted loss-of-function and gain-of-function assays with pepper and Arabidopsis, respectively. CaMFT02-silenced pepper exhibited increased ABA sensitivity and drought tolerance, whereas CaMFT02-overexpressing Arabidopsis showed ABA hyposensitivity and drought hypersensitivity. Silencing of CaMFT03, a CaMFT02 homolog, improved salt stress tolerance in pepper, and this negative regulatory role of CaMFT03 in salt stress tolerance was also observed in CaMFT03-overexpressing Arabidopsis. Taken together, our findings indicate that CaMFT02 negatively regulates drought stress tolerance via ABA signaling in pepper by modulating ABA signaling and suggest that CaMFT03 plays a similar negative regulatory role in salt stress tolerance.

Mutation of the PEBP-like domain of the mitoribosomal MrpL35/mL38 protein results in production of nascent chains with impaired capacity to assemble into OXPHOS complexes.

Located in the central protuberance region of the mitoribosome and mitospecific mL38 proteins display homology to PEBP (Phosphatidylethanolamine Binding Protein) proteins, a diverse family of proteins reported to bind anionic substrates/ligands and implicated in cellular signaling and differentiation pathways. In this study, we have performed a mutational analysis of the yeast mitoribosomal protein MrpL35/mL38 and demonstrate that mutation of the PEBP-invariant ligand binding residues Asp(D)232 and Arg(R)288 impacted MrpL35/mL38's ability to support OXPHOS-based growth of the cell. Furthermore, our data indicate these residues exist in a functionally important charged microenvironment, which also includes Asp(D)167 of MrpL35/mL38 and Arg(R)127 of the neighboring Mrp7/bL27m protein. We report that mutation of each of these charged residues resulted in a strong reduction in OXPHOS complex levels that was not attributed to a corresponding inhibition of the mitochondrial translation process. Rather, our findings indicate that a disconnect exists in these mutants between the processes of mitochondrial protein translation and the events required to ensure the competency and/or availability of the newly synthesized proteins to assemble into OXPHOS enzymes. Based on our findings, we postulate that the PEBP-homology domain of MrpL35/mL38, together with its partner Mrp7/bL27m, form a key regulatory region of the mitoribosome.

Genome-Wide Characterization of PEBP Gene Family and Functional Analysis of TERMINAL FLOWER 1 Homologs in Macadamia integrifolia.

Edible Macadamia is one of the most important commercial nut trees cultivated in many countries, but its large tree size and long juvenile period pose barriers to commercial cultivation. The short domestication period and well-annotated genome of Macadamia integrifolia create great opportunities to breed commercial varieties with superior traits. Recent studies have shown that members of the phosphatidylethanolamine binding protein (PEBP) family play pivotal roles in regulating plant architecture and flowering time in various plants. In this study, thirteen members of MiPEBP were identified in the genome of M. integrifolia, and they are highly similarity in both motif and gene structure. A phylogenetic analysis divided the MiPEBP genes into three subfamilies: MFT-like, FT-like and TFL1-like. We subsequently identified two TERMINAL FLOWER 1 homologues from the TFL1-like subfamily, MiTFL1 and MiTFL1-like, both of which were highly expressed in stems and vegetative shoots, while MiTFL1-like was highly expressed in young leaves and early flowers. A subcellular location analysis revealed that both MiTFL1 and MiTFL1-like are localized in the cytoplasm and nucleus. The ectopic expression of MiTFL1 can rescue the early-flowering and terminal-flower phenotypes in the tfl1-14 mutant of Arabidopsis thaliana, and it indicates the conserved functions in controlling the inflorescence architecture and flowering time. This study will provide insight into the isolation of PEBP family members and the key targets for breeding M. integrifolia with improved traits in plant architecture and flowering time.

Overexpression of the Capebp2 Gene Encoding the PEBP-like Protein Promotes the Cap Redifferentiation in Cyclocybe aegerita.

Phosphatidylethanolamine-binding protein (PEBP) is widely involved in various physiological behaviors, such as the transition from vegetative growth to reproductive growth in plants, tumorigenesis in the human, etc. However, few functional studies have examined pebp genes affecting the development of fungi. In this study, Capebp2 was cloned from Cyclocybe aegerita AC0007 strains based on the genome sequence and gene prediction, and the sequence alignment of CaPEBP2 with other PEBP proteins from other biological sources including plant, animal, fungi, and bacteria indicated that PEBP had low sequence similarity in fungi, whereas all protein sequences had some conserved motifs such as DPDAP and HRY. Expression analysis showed the transcription level of Capebp2 increased approximately 20-fold in fruiting bodies compared with mycelia. To uncover the function of Capebp2 in C. aegetita development, Capebp2 was cloned into a pATH vector driven by the actin promoter for obtaining overexpression transformant lines. Fruiting experiments showed the transformed strains overexpressing Capebp2 exhibited redifferentiation of the cap on their surface, including intact fruiting bodies or partial lamella during fruiting development stage, and the longitudinal section indicated that all regenerated bodies or lamella sprouted from the flesh and shared the epidermis with the mother fruiting bodies. In summary, the sequence characterization of Capebp2, expression level during different development stages, and function on fruiting body development were documented in this study, and these findings provided a reference to study the role of pebp in the development process of basidiomycetes. Importantly, gene mining of pebp, function characterization, and the regulating pathways involved need to be uncovered in further studies.

Multiple sevoflurane exposures during mid-trimester induce neurotoxicity in the developing brain initiated by 15LO2-Mediated ferroptosis.

Mid-gestational sevoflurane exposure may induce notable long-term neurocognitive impairment in offspring. This study was designed to investigate the role and potential mechanism of ferroptosis in developmental neurotoxicity induced by sevoflurane in the second trimester. Pregnant rats on day 13 of gestation (G13) were treated with or without 3.0% sevoflurane, Ferrostatin-1 (Fer-1), PD146176, or Ku55933 on three consecutive days. Mitochondrial morphology, ferroptosis-relative proteins, malondialdehyde (MDA) levels, total iron content, and glutathione peroxidase 4 (GPX4) activities were measured. Hippocampal neuronal development in offspring was also examined. Subsequently, 15-lipoxygenase 2 (15LO2)-phosphatidylethanolamine binding protein 1 (PEBP1) interaction and expression of Ataxia telangiectasia mutated (ATM) and its downstream proteins were also detected. Furthermore, Morris water maze (MWM) and Nissl's staining were applied to estimate the long-term neurotoxic effects of sevoflurane. Ferroptosis mitochondria were observed after maternal sevoflurane exposures. Sevoflurane elevated MDA and iron levels while inhibiting GPX4 activity, and resultant long-term learning and memory dysfunction, which were alleviated by Fer-1, PD146176, and Ku55933. Sevoflurane could enhance 15LO2-PEBP1 interaction and activate ATM and its downstream P53/SAT1 pathway, which might be attributed to excessive p-ATM nuclear translocation. This study proposes that 15LO2-mediated ferroptosis might contribute to neurotoxicity induced by maternal sevoflurane anesthesia during the mid-trimester in the offspring and its mechanism may be ascribed to hyperactivation of ATM and enhancement of 15LO2-PEBP1 interaction, indicating a potential therapeutic target for ameliorating sevoflurane-induced neurotoxicity.

GmMFT: a potential step forward in soybean breeding for high oil and yield.

This article is a Commentary on Cai et al. (2023), 239: 905–919.

Functional characterization of three TERMINAL FLOWER 1-like genes from Platanus acerifolia.

TFL1-like genes of the basal eudicot Platanus acerifolia have conserved roles in maintaining vegetative growth and inhibiting flowering, but may act through distinct regulatory mechanism. Three TERMINAL FLOWER 1 (TFL1)-like genes were isolated and characterized from London plane tree (Platanus acerifolia). All genes have conserved genomic organization and characteristic of the phosphatidylethanolamine-binding protein (PEBP) family. Sequence alignment and phylogenetic analysis indicated that two genes belong to the TFL1 clade, designated as PlacTFL1a and PlacTFL1b, while another one was grouped in the BFT clade, named as PlacBFT. qRT-PCR analysis showed that all three genes primarily expressed in vegetative phase, but the expression of PlacTFL1a was much higher and wider than that of PlacTFL1b, with the latter only detected at relatively low expression levels in apical and lateral buds in April. PlacBFT was mainly expressed in young stems of adult trees followed by juvenile tissues. Ectopic expression of any TFL1-like gene in Arabidopsis showed phenotypes of delayed or repressed flowering. Furthermore, overexpression of PlacTFL1a gene in petunia also resulted in extremely delayed flowering. In non-flowering 35:PlacTFL1a transgenic petunia plants, the FT-like gene (PhFT) gene was significantly upregulated and AP1 homologues PFG, FBP26 and FBP29 were significantly down-regulated in leaves. Yeast two-hybrid analysis indicated that only weak interactions were detected between PlacTFL1a and PlacFDL, and PlacTFL1a showed no interaction with PhFDL1/2. These results indicated that the TFL1-like genes of Platanus have conserved roles in repressing flowering, but probably via a distinct regulatory mechanism.

OsMFT1 Inhibits Seed Germination by Modulating Abscisic Acid Signaling and Gibberellin Biosynthesis under Salt Stress in Rice.

Seed dormancy and germination are regulated by endogenous gene expression as well as hormonal and environmental conditions, such as salinity, which greatly inhibits seed germination. MOTHER OF FT AND TFL1 (MFT), which encodes a phosphatidylethanolamine-binding protein, is a key regulator of seed germination in Arabidopsis thaliana. There are two orthologous genes of AtMFT in rice (Oryza sativa), namely, OsMFT1 and OsMFT2. However, the functions of these two genes in regulating rice seed germination under salt stress remain unknown. In this study, we found that seeds of loss-of-function osmft1 mutants germinated faster than wild-type (WT) seeds under salt stress, but this was not the case for loss-of-function osmft2 mutants. Overexpression of OsMFT1 (OsMFT1OE) or OsMFT2 increased the sensitivity to salt stress during seed germination. Transcriptome comparisons of osmft1 vs WT in the absence and presence of salt stress yielded several differentially expressed genes, which were associated with salt stress, plant hormone metabolism and signaling pathways, such as B-BOX ZINC FINGER 6, O. sativa bZIP PROTEIN 8 and GIBBERELLIN (GA) 20-oxidase 1. In addition, the sensitivity of OsMFT1OE seeds to GA and osmft1 seeds to abscisic acid (ABA) during seed germination increased under salt stress. Overall, our results indicate that ABA and GA metabolism and their signaling pathways are regulated by OsMFT1, modulating seed germination in rice under salt stress.

Genome-edited TaTFL1-5 mutation decreases tiller and spikelet numbers in common wheat.

Tillering is a critical agronomic trait of wheat (Triticum aestivum L.) that determines the shoot architecture and affects grain yield. TERMINAL FLOWER 1 (TFL1), encoding a phosphatidylethanolamine-binding protein, is implicated in the transition to flowering and shoot architecture in plant development. However, the roles of TFL1 homologs is little known in wheat development. CRISPR/Cas9-mediated targeted mutagenesis was used in this study to generate a set of wheat (Fielder) mutants with single, double or triple-null tatfl1-5 alleles. The wheat tatfl1-5 mutations decreased the tiller number per plant in the vegetative growth stage and the effective tiller number per plant and spikelet number per spike at maturity in the field. RNA-seq analysis showed that the expression of the auxin signaling-related and cytokinin signaling-related genes was significantly changed in the axillary buds of tatfl1-5 mutant seedlings. The results suggested that wheat TaTFL1-5s were implicated in tiller regulation by auxin and cytokinin signaling.

Identification and Analysis of Phosphatidylethanolamine-Binding Protein Family Genes in the Hangzhou White Chrysanthemum (Chrysanthemum morifolium Ramat)

The Hangzhou White Chrysanthemum (Chrysanthemum morifolium Ramat) is one of the “Zhejiang eight flavors” in traditional Chinese medicine. The phosphatidylethanolamine-binding protein (PEBP) plays an important role in flowering and floral organ development. Even so, the biological role of PEBPs in the Hangzhou White Chrysanthemum has not been studied, which attracted us. Here, nine CmPEBP genes that contain the PF01161 domain were identified in the Hangzhou White Chrysanthemum for the first time, and their biological role in flowering was preliminarily studied. A phylogenetic analysis classified the CmPEBP genes into three subfamilies: MFT-like, TFL-like, and FT-like genes. The differential expression analysis was performed under different tissues and different stressors using qRT-PCR. It showed that each CmPEBP displayed tissue-specific expression patterns. Expression patterns in response to different temperatures and hormone stressors were investigated. They were finally demonstrated to be differentially expressed. TFL-like gene expression, which delayed reproductive growth, was upregulated under heat stress. Conversely, FT-like gene expression was upregulated under low temperatures. CmFT1 expression could be inhibited by GA (gibberellin), 6-BA (benzylaminopurine), ET (ethylene), and MeSA (methyl salicylate) but could be activated by IAA (indole-3-aceticacid), ABA (abscisic acid), and SA (salicylic acid) in the dark, whereas CmFT2 and CmFT3 expression levels were upregulated by ET, MeJA (methyl jasmonate), and ABA but were downregulated by 6-BA, SA, and MeSA. GA, IAA, SA, and MeSA inhibited CmTFL gene expression under light and dark treatments. Further research on CmPEBP genes in the Hangzhou White Chrysanthemum could better determine their roles in flowering and floral organ development, especially in response to the prolonged spraying of exogenous hormones.

Vemurafenib and Dabrafenib Downregulates RIPK4 Level.

Vemurafenib and dabrafenib are BRAF kinase inhibitors (BRAFi) used for the treatment of patients with melanoma carrying the V600E BRAF mutation. However, melanoma cells develop resistance to both drugs when used as monotherapy. Therefore, mechanisms of drug resistance are investigated, and new molecular targets are sought that could completely inhibit melanoma progression. Since receptor-interacting protein kinase (RIPK4) probably functions as an oncogene in melanoma and its structure is similar to the BRAF protein, we analyzed the impact of vemurafenib and dabrafenib on RIPK4 in melanomas. The in silico study confirmed the high similarity of BRAF kinase domains to the RIPK4 protein at both the sequence and structural levels and suggests that BRAFi could directly bind to RIPK4 even more strongly than to ATP. Furthermore, BRAFi inhibited ERK1/2 activity and lowered RIPK4 protein levels in BRAF-mutated melanoma cells (A375 and WM266.4), while in wild-type BRAF cells (BLM and LoVo), both inhibitors decreased the level of RIPK4 and enhanced ERK1/2 activity. The phosphorylation of phosphatidylethanolamine binding protein 1 (PEBP1)-a suppressor of the BRAF/MEK/ERK pathway-via RIPK4 observed in pancreatic cancer did not occur in melanoma. Neither downregulation nor upregulation of RIPK4 in BRAF- mutated cells affected PEBP1 levels or the BRAF/MEK/ERK pathway. The downregulation of RIPK4 inhibited cell proliferation and the FAK/AKT pathway, and increased BRAFi efficiency in WM266.4 cells. However, the silencing of RIPK4 did not induce apoptosis or necroptosis. Our study suggests that RIPK4 may be an off-target for BRAF inhibitors.

Genome-wide identification and expression analysis disclose the pivotal PHOSPHATIDYLETHANOLAMINE BINDING PROTEIN members that may be utilized for yield improvement of Chenopodium quinoa.

Quinoa (Chenopodium quinoa) is a prospective orphan crop that needs yield improvement. Previous studies indicate PHOSPHATIDYLETHANOLAMINE BINDING PROTEIN (PEBP) family genes are highly associated with the key agronomic traits of crops. Characterizing the pivotal PEBP genes will speed up the domestication and yield improvement of quinoa. Previous investigations on PEBP genes of Chenopodium species indicated that, the PEBP genes, despite in the same subclade, may have experienced functional diversification. Especially, the allotetraploidy (AABB) and numerous segmental duplications and chromosomal rearrangements in quinoa make it more difficult to understand the functions of PEBP genes. More recently, 6 quinoa FT subfamily genes were predicted to be related to flowering of quinoa. However, investigation on the whole PEBP family members is still lacking. In this study, we obtained 23 PEBP genes, including 5 MFT, 11 FTL and 7 TFL genes. We found 7 orthologous gene pairs, from sub-genome A and sub-genome B, respectively, showing collinearities with sugar beet. Evolution analysis on PEBP genes of two quinoa sub-genomes, sugar beet and relatives of diploid ancestors indicated that, the reasons for gene duplication events varied and 4 tandem duplications are the major reason for PEBP family expansion. Tissue-specific expression analysis suggested that expression patterns are mostly differing between orthologous gene pairs. Analysis on gene expressions at 6 stages suggested the possible positive roles of CqFTL1/CqFTL2, CqFTL5, CqFTL8, CqFTL6/CqFTL9 and CqTFL6/CqTFL7, and negative roles of CqTFL1/CqTFL2/CqTFL3, CqTFL4/CqTFL5 in inflorescence branching. Expression analysis in ABA-treated seed, in combination with the cis-acting element distribution analysis, indicated that CqMFT2, CqMFT3 and CqMFT4 may regulate seed germination via ABA signaling. Observations on responses to night break and photoperiod changes highlighted the roles of CqFTL5 and CqFTL8 under short day, and CqFTL6 under long day for quinoa flowering. Further, co-expression network analysis indicated that 64 transcription factors may act upstream of CqFTL5 and CqFTL8 to regulate flowering. Together, this study will help us identify the pivotal PEBP genes that may be utilized for quinoa breeding in future.

PEBP Signaling Network in Tubers and Tuberous Root Crops.

Tubers and tuberous root crops are essential carbohydrate sources and staple foods for humans, second only to cereals. The developmental phase transition, including floral initiation and underground storage organ formation, is controlled by complex signaling processes involving the integration of environmental and endogenous cues. FLOWERING LOCUS T (FT) and TERMINAL FLOWER 1/CENTRORADIALIS (TFL1/CEN), members of the phosphatidylethanolamine-binding protein (PEBP) gene family, play a central role in this developmental phase transition process. FT and FT-like proteins have a function to promote developmental phase transition, while TFL1/CEN act oppositely. The balance between FT and TFL1/CEN is critical to ensure a successful plant life cycle. Here, we present a summarized review of the role and signaling network of PEBP in floral initiation and underground storage organ formation, specifically in tubers and tuberous root crops. Lastly, we point out several questions that need to be answered in order to have a more complete understanding of the PEBP signaling network, which is crucial for the agronomical improvement of tubers and tuberous crops.

Overexpression of mango MiMFT inhibits seed germination and enhances abiotic stress tolerance in transgenic Arabidopsis

Mother of FT and TFL1 (MFT) is a member of the phosphatidylethanolamine-binding protein (PEBP) family, which plays important roles in seed development, the stress response, and flowering time regulation. In the present study, the MFT homologous gene MiMFT was identified in mango cultivar ‘SiJiMi’. The promoter of MiMFT contained phytohormone-, plant growth-, plant development-, and abiotic stress-responsive elements. MiMFT was significantly highly expressed in the seeds, and its expression strongly increased in response to polyethylene glycol (PEG) and NaCl treatment. Overexpression of MiMFT in Arabidopsis adversely affected seed germination and upregulated seed germination-related gene expression, but it did not affect flowering. Additionally, the salt and drought tolerance of transgenic Arabidopsis lines overexpressing MiMFT considerably increased, and these plants presented decreased sensitivity to abscisic acid (ABA), with significantly increased expression of stress-related and ABA signaling pathway genes. Subcellular localization analysis showed that MiMFT strongly localizes in the nucleus, and bimolecular fluorescence complementation (BiFC) assays showed that MiMFT proteins interact with Mi14–3–3D2. In summary, MiMFT was determined to be involved in seed germination as well as in the response to abiotic stress.

Effects of Raf kinase inhibitor protein on biological characteristics of hepatocellular carcinoma cells and its potential therapeutic effects

Raf kinase inhibitor protein (RKIP) is an important member of the phosphatidylethanolamine-binding protein family. This study investigated the inhibitory effect of RKIP on the malignant biological behavior of liver cancer cells in vivo and in vitro. An RKIP gene expression vector was constructed using the pcDNA3.1 vector and transfected into human hepatoma cell line HepG2 to overexpress the RKIP gene. Growth, proliferation, the cell cycle, apoptosis, migration, and invasion of the cell line were analyzed. For in vivo experiments, the vector was transfected into tumor tissue of hepatoma-bearing animals and then tumor growth was analyzed and compared with the control to assess its anti-tumor effect. Compared with the control group, cell growth in the RKIP gene transfection group (HEP-RKIP) was significantly slower. The average colony formation rate of the HEP-RKIP group was significantly lower than that of the other two groups (p < 0.05). The proportion of HEP-RKIP cells in G0/G1 phase was significantly higher than that of the control group (p < 0.05), while the proportion of cells in G2/M phase was significantly lower than that in the control group (p < 0.05). The apoptosis rate of HEP-RKIP cells was approximately 6.4%, which was significantly higher than that of the control group (p < 0.05). Migration of HEP-RKIP cells was significantly slower than that of the other two groups (p < 0.05). In an inhibition experiment of the liver tumor animal model, the tumor inhibition rate of the RKIP in vivo transfection group was significantly higher than that of each control group (p < 0.05). RKIP inhibits malignant biological behavior of liver cancer cells in vitro and has a tumor inhibitory effect in vivo, which may be a potential target for gene therapy of liver cancer.

Citrus VASCULAR PLANT ONE-ZINC FINGER1 (VOZ1) interacts with CuFT1 and CuFT3, affecting flowering in transgenic Arabidopsis

Understanding the flowering process is essential for developing methods to shorten the breeding period and regulate the yield of tree fruits, such as Satsuma mandarin (Citrus unshiu Marc.). In the flowering network system in Arabidopsis, FLOWERING LOCUS T (FT) acts as a transmissible floral inducer. In this study, a citrus ortholog of the transcription factor (TF) VASCULAR PLANT ONE-ZINC FINGER1 (VOZ1)-like gene, CuVOZ1, and two FT-like genes, CuFT1 and CuFT3, were isolated from the Satsuma mandarin ‘Aoshima’. Protein-protein interaction between CuVOZ1 and CuFTs was confirmed in vitro with a yeast two-hybrid (Y2H) system. N-terminal 400 amino acids of CuVOZ1, consisting of three motifs, namely, domain of unknown function 4749 (DUF4749), no apical meristem (NAM), and a zinc coordination motif, were assumed to be involved in the CuVOZ1–CuFT1 and CuVOZ1–CuFT3 complexes. Docking simulation suggested that only the zinc coordination motif of CuVOZ1 was possibly involved in the CuVOZ1–CuFT3 interaction and that the phosphatidylethanolamine-binding protein (PBP) motif in exon 4 of CuFTs was predicted to be crucial for the interaction between CuVOZ1 and CuFTs. The distance between the amino acid residues involved in docking was predicted to be from 2.69 to 3.37 Å, suggesting that the forces between CuVOZ1 and CuFTs in the CuVOZ1–CuFT complexes were weak Van der Waals forces. Constitutive expression of CuVOZ1 in Arabidopsis affected the flowering time, plant size, morphology of inflorescence, number of flowers and siliques, and formation of flower buds on the elongated stem. It was suggested that CuVOZ1 might act as a trigger for early flowering and might be involved in the elongation and branching of the inflorescence. The CuVOZ1–CuFT complexes might regulate cellular proliferation and the formation of new tissues and affect both vegetative and reproductive development.

Evolution of the PEBP gene family in Juglandaceae and their regulation of flowering pathway under the synergistic effect of JrCO and JrNF-Y proteins

Phosphatidyl ethanolamine-binding protein (PEBP) has a conserved PEBP domain and plays an important role in regulating the flowering time and growth of angiosperms. To understand the evolution of PEBP family genes in walnut family and the mechanism of regulating flowering in photoperiod pathway, 53 genes with PEBP domain were identified from 5 Juglandaceae plants. The PEBP gene family of Juglandaceae can be divided into four subgroups, FT-like, TFL-like, MFT-like and PEBP-like subgroups. These genes all show very high homology for motifs and gene structure in Juglandaceae. In addition, the results of gene replication and collinearity analysis showed that the evolution of PEBP genes was mainly purified and selected, and segmental repetition was the main driving force for the evolution of PEBP gene family in walnut family. We found that PEBP gene family played an important role in female flower bud differentiation, and most JrPEBP genes were highly expressed in leaf bud and female flower bud by qRT-PCR. In Arabidopsis, AtCO can not only directly bind to CORE2, but also interact with NF-Y complex to positively regulate the expression of AtFT gene. In this study, we proved that JrCO (the lineal homologue of AtCO) could not directly regulate the expression of JrFT gene, but could enhance the binding of JrNF-YB4/6 protein to the promoter of JrFT gene by forming a heteropolymer with NF-YB4/NF-YB6. We also confirmed that JrNF-YC1/3/7, JrNF-YB4/6 and JrCO can form a trimer structure similar to AtNF-YB-YC-CO of Arabidopsis, and then bind to the promoter of JrFT gene to promote the transcription of JrFT gene. In a word, through identification and analysis of PEBP gene family in Juglandaceae and study on the mechanism of photoperiod pathway regulating flowering in walnut, we have found that nuclear transcription factor NF-YB/YC plays a more important role in the trimer structure of NF-YB-YC-CO in walnut species. Our study has further perfected the flowering regulatory network of walnut species.

Bull sperm and seminal plasma proteins and their relationship with fertility: a review

The efficiency of artificial insemination (AI) is greatly influenced by the quality of semen. Spermatozoa and seminal plasma are found in semen, which play a role in the reproductive process and its ability to fertilize an egg and maintain the development of an embryo. Various factors will determine the fertility capacity of a sperm, both from the intrinsic factors of the sperm and the plasma component of the semen. Seminal plasma proteins are crucial for maintaining the stability of the membrane, viability, motility of spermatozoa, acrosome reactions, maintaining osmotic pressure and helping the fertilization process. Good quality semen will support the fertilization process. The purpose of this scoping review is to increase our understanding of protein from sperm and seminal plasma of bulls and their relationship with fertility. The sperm proteins that were significantly correlated with fertility were Outer Dense Fiber protein 2 (ODF2), Protamine (PRM), Testis specific histine 2B (TH2B), Phosphatidylethanolamine binding protein (PEBP4), and Ubiquinol-cytochrome-c reductase complex core protein 2 (UQCRSC2). Meanwhile, the seminal plasma proteins positively correlated with fertility were Osteopontin (OPN), Phospholipasea 2 (PLA2), P25b, Acidic seminal fluid proteins (aSFP), Alpha-L-fucosidase (a-L-fucosidase), and Binder of sperm (BSPs).

Phosphatidylethanolamine-binding protein 4 deficiency exacerbates carbon tetrachloride-induced liver fibrosis by regulating the NF-κB signaling pathway.

Liver fibrosis is a pathological process which can progress to hepatocirrhosis, even hepatocellular carcinoma. Phosphatidylethanolamine-binding protein 4 (PEBP4) is a secreted protein involved in regulating many molecular pathways, whereas its roles in diseases including hepatic fibrosis remain undefined. The nuclear factor-κappa B (NF-κB) signaling pathway has been found to be involved in the development of liver fibrosis. In this study, we generated a hepatocyte-conditional knockout (CKO) mouse model of PEBP4, and explored the potential functions of PEBP4 on liver fibrosis and the NF-κB signaling pathway in a mouse model of carbon tetrachloride (CCl4)-induced liver fibrosis. We demonstrated that PEBP4 CKO aggravated CCl4-triggered liver fibrosis, as evidenced by altered histopathology, an increase in the activities of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and hydroxyproline (HYP) levels, and more collagen deposition, as well as by enhanced expression of fibrotic markers including α-smooth muscle actin (α-SMA), collagen I and collagen III. Mechanistically, PEBP4 deficiency activated the NF-κB signaling pathway, as indicated by increased phosphorylation of NF-κB p65 and inhibitor protein κB inhibitor-α (IκB-α), and nuclear NF-κB p65 expression in the fibrotic liver. Notably, the NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC) partially blocked the activation of the NF-κB pathway, and reversed the pro-fibrotic effect of PEBP4 deletion in CCl4-treated mice. Together, these results suggest that PEBP4 deficiency results in aggravation of liver fibrosis and activation of the NF-κB signaling pathway, supporting a novel concept that PEBP4 is a crucial player in hepatic fibrosis, but also might be a negative regulator of the NF-κB signaling in liver fibrosis.

Dihydromyricetin Attenuates Cerebral Ischemia Reperfusion Injury by Inhibiting SPHK1/mTOR Signaling and Targeting Ferroptosis.

BackgroundDihydromyricetin (DHM) exerts protective effects in various brain diseases. The aim of this research was to investigate the biological role of DHM in cerebral ischemia reperfusion (I/R) injury.MethodsWe generated a rat model of cerebral I/R injury by performing middle cerebral artery occlusion/reperfusion (MCAO/R). The neurological score and brain water content of the experimental rats was then evaluated. The infarct volume and extent of apoptosis in brain tissues was then assessed by 2,3,5-triphenyltetrazolium (TTC) and TdT-mediated dUTP nick end labeling (TUNEL) staining. Hippocampal neuronal cells (HT22) were subjected to oxygen-glucose deprivation/reperfusion (OGD/R) and cell counting kit-8 (CCK-8) assays and flow cytometry were performed to detect cell viability and apoptosis. The levels of lipid reactive oxygen species (ROS) and iron were detected and the expression levels of key proteins were assessed by Western blotting.ResultsDHM obviously reduced neurological deficits, brain water content, infarct volume and cell apoptosis in the brain tissues of MCAO/R rats. DHM repressed ferroptosis and inhibited the sphingosine kinase 1 (SPHK1)/mammalian target of rapamycin (mTOR) pathway in MCAO/R rats. In addition, DHM promoted cell viability and repressed apoptosis in OGD/R-treated HT22 cells. DHM also suppressed the levels of lipid ROS and intracellular iron in OGD/R-treated HT22 cells. The expression levels of glutathione peroxidase 4 (GPX4) was enhanced while the levels of acyl-CoA synthetase long-chain family member 4 (ACSL4) and phosphatidylethanolamine binding protein 1 (PEBP1) were reduced in OGD/R-treated HT22 cells in the presence of DHM. Moreover, the influence conferred by DHM was abrogated by the overexpression of SPHK1 or treatment with MHY1485 (an activator of mTOR).ConclusionThis research demonstrated that DHM repressed ferroptosis by inhibiting the SPHK1/mTOR signaling pathway, thereby alleviating cerebral I/R injury. Our findings suggest that DHM may be a candidate drug for cerebral I/R injury treatment.