SlERF.F12 modulates the transition to ripening in tomato fruit by recruiting the co-repressor TOPLESS and histone deacetylases to repress key ripening genes.

Ethylene response factors (ERFs) are downstream components of ethylene-signaling pathways known to play critical roles in ethylene-controlled climacteric fruit ripening, yet little is known about the molecular mechanism underlying their mode of action. Here, we demonstrate that SlERF.F12, a member of the ERF.F subfamily containing Ethylene-responsive element-binding factor-associated Amphiphilic Repression (EAR) motifs, negatively regulates the onset of tomato (Solanum lycopersicum) fruit ripening by recruiting the co-repressor TOPLESS 2 (TPL2) and the histone deacetylases (HDAs) HDA1/HDA3 to repress the transcription of ripening-related genes. The SlERF.F12-mediated transcriptional repression of key ripening-related genes 1-AMINO-CYCLOPROPANE-1-CARBOXYLATE SYNTHASE 2 (ACS2), ACS4, POLYGALACTURONASE 2a, and PECTATE LYASE is dependent on the presence of its C-terminal EAR motif. We show that SlERF.F12 interacts with the co-repressor TPL2 via the C-terminal EAR motif and recruits HDAs SlHDA1 and SlHDA3 to form a tripartite complex in vivo that actively represses transcription of ripening genes by decreasing the level of the permissive histone acetylation marks H3K9Ac and H3K27Ac at their promoter regions. These findings provide new insights into the ripening regulatory network and uncover a direct link between repressor ERFs and histone modifiers in modulating the transition to ripening of climacteric fruit.

MicroRNA482/2118 is lineage-specifically involved in gibberellin signalling via the regulation of GID1 expression by targeting noncoding PHAS genes and subsequently instigated phasiRNAs.

MicroRNA482/2118 (miR482/2118) is a 22-nt miRNA superfamily, with conserved functions in disease resistance and plant development. It usually instigates the production of phased small interfering RNAs (phasiRNAs) from its targets to expand or reinforce its silencing effect. Using a new high-quality reference genome sequence and comprehensive small RNA profiling, we characterized a newly evolved regulatory pathway of miR482/2118 in litchi. In this pathway, miR482/2118 cleaved a novel noncoding trans-acting gene (LcTASL1) and triggered phasiRNAs to regulate the expression of gibberellin (GA) receptor gene GIBBERELLIN INSENSITIVE DWARF1 (GID1) in trans; another trans-acting gene LcTASL2, targeted by LcTASL1-derived phasiRNAs, produced phasiRNAs as well to target LcGID1 to reinforce the silencing effect of LcTASL1. We found this miR482/2118-TASL-GID1 pathway was likely involved in fruit development, especially the seed development in litchi. In vivo construction of the miR482a-TASL-GID1 pathway in Arabidopsis could lead to defects in flower and silique development, analogous to the phenotype of gid1 mutants. Finally, we found that a GA-responsive transcription factor, LcGAMYB33, could regulate LcMIR482/2118 as a feedback mechanism of the sRNA-silencing pathway. Our results deciphered a lineage-specifically evolved regulatory module of miR482/2118, demonstrating the high dynamics of miR482/2118 function in plants.

Photosynthesis Response and Transcriptional Analysis: Dissecting the Role of SlHB8 in Regulating Drought Resistance in Tomato Plants.

Deciphering drought resistance in crops is crucial for enhancing water productivity. Previous studies have highlighted the significant role of the transcription factor SlHB8 in regulating developmental processes in tomato plants but its involvement in drought resistance remains unclear. Here, gene overexpression (SlHB8-OE) and gene knockout (slhb8) tomato plants were utilized to study the role of SlHB8 in regulating drought resistance. Our findings showed that slhb8 plants exhibited a robust resistant phenotype under drought stress conditions. The stomata of slhb8 tomato leaves displayed significant closure, effectively mitigating the adverse effects of drought stress on photosynthetic efficiency. The slhb8 plants exhibited a decrease in oxidative damage and a substantial increase in antioxidant enzyme activity. Moreover, slhb8 effectively alleviated the degree of photoinhibition and chloroplast damage caused by drought stress. SlHB8 regulates the expression of numerous genes related to photosynthesis (such as SlPSAN, SlPSAL, SlPSBP, and SlTIC62) and stress signal transduction (such as SlCIPK25, SlABA4, and SlJA2) in response to drought stress. Additionally, slhb8 plants exhibited enhanced water absorption capacity and upregulated expression of several aquaporin genes including SlPIP1;3, SlPIP2;6, SlTIP3;1, SlNIP1;2, and SlXIP1;1. Collectively, our findings suggest that SlHB8 plays a negative regulatory role in the drought resistance of tomato plants.

BcSOC1 Promotes Bolting and Stem Elongation in Flowering Chinese Cabbage.

Flowering Chinese cabbage is one of the most economically important stalk vegetables. However, the molecular mechanisms underlying bolting, which is directly related to stalk quality and yield, in this species remain unknown. Previously, we examined five key stem development stages in flowering Chinese cabbage. Here, we identified a gene, BcSOC1 (SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1), in flowering Chinese cabbage using transcriptome analysis, whose expression was positively correlated with bolting. Exogenous gibberellin (GA3) and low-temperature treatments significantly upregulated BcSOC1 and promoted early bolting and flowering. Additionally, BcSOC1 overexpression accelerated early flowering and stem elongation in both Arabidopsis and flowering Chinese cabbage, whereas its knockdown dramatically delayed bolting and flowering and inhibited stem elongation in the latter; the inhibition of stem elongation was more notable than delayed flowering. BcSOC1 overexpression also induced cell expansion by upregulating genes encoding cell wall structural proteins, such as BcEXPA11 (cell wall structural proteins and enzymes) and BcXTH3 (xyloglucan endotransglycosidase/hydrolase), upon exogenous GA3 and low-temperature treatments. Moreover, the length of pith cells was correlated with stem height, and BcSOC1 interacted with BcAGL6 (AGAMOUS-LIKE 6) and BcAGL24 (AGAMOUS-LIKE 24). Thus, BcSOC1 plays a vital role in bolting and stem elongation of flowering Chinese cabbage and may play a novel role in regulating stalk development, apart from the conserved function of Arabidopsis SOC1 in flowering alone.

Identification of BcARR Genes and CTK Effects on Stalk Development of Flowering Chinese Cabbage.

Flowering Chinese cabbage (Brassica campestris L. ssp. Chinensis var. utilis Tsen et Lee) is an important and extensively cultivated vegetable in south China, whose major food product is the stalk. In the process of stalk formation, its initiation and development are regulated by a series of hormonal signals, such as cytokinin and gibberellin. In this study, we analyzed the effects of zeatin (ZT) and gibberellin A3 (GA3), and their interaction, on the bolting of flowering Chinese cabbage. The results indicated that the three-true-leaf spraying of ZT and GA synthesis inhibitor (PAC) inhibited plant height but increased stem diameter. Cytokinin (CTK) synthesis inhibitor (YZJ) and GA3 treatment increased plant height and decreased stem diameter. In addition, ZT and GA3 co-treated plants displayed antagonistic effect. Further, 19 type-B authentic response regulators (ARR-Bs), the positive regulators of cytokinin signal transduction were identified from flowering Chinese cabbage. Comprehensive analysis of phylogeny showed BcARR-Bs clustered into three subfamilies with 10 conserved motifs. Analysis of their expression patterns in different tissues and at various growth stage, and their response to hormone treatment suggest that ARR1-b localized in the nucleus displayed unique highest expression patterns in stem tips, are responsive both to ZT and GA, suggesting a significant role in mediating the crosstalk of ZT and GA in the bolting of flowering Chinese cabbage.

Nitric Oxide Is Essential for Melatonin to Enhance Nitrate Tolerance of Cucumber Seedlings.

Melatonin (MT) and nitric oxide (NO) in plants can function cooperatively to alleviate salt stress, sodic alkaline stress and immune response, as well as adventitious root formation. The interaction of MT and NO on the nitrate stress tolerance of cucumber seedlings are not well understood. We investigated the effects of exogenous MT, NO donor (SNP) and NO scavenger (cPTIO) on the growth; photosynthesis; characteristics of root morphological; accumulation of mineral elements, endogenous NO, MT, IAA and ABA; and related genes expression in cucumber (Cucumis sativus L. "Jin You No. 1") seedlings grown under high nitrate condition (HN). The results showed that MT and NO independently alleviated the inhibition of growth and photosynthesis capacity of cucumber seedlings under nitrate stress. NO was required for MT to enhance the root activity, root length, lateral root number and the accumulation of calcium, magnesium and iron in the roots of cucumber seedlings grown under nitrate stress. Consistently, the expression of adventitious rootless 1 gene (CsARL1) was modulated. Furthermore, exogenous MT induced accumulation of endogenous MT, NO, indole-3-acetic acid (IAA) and abscisic acid (ABA), mainly within 24 h after treatment, in which MT and NO were further increased at 48 h and 96 h, IAA and ABA were further increased at 16 h in the presence of SNP. In contrast, the accumulation of endogenous IAA, MT and ABA slightly decreased within 24 h, NO significantly decreased at 192 h in the presence of cPTIO. Correspondingly, the expression levels of genes involved in nitrogen metabolism (CsNR1 and CsNR2), MT metabolism (CsT5H, CsSNAT2 and Cs2-ODD33), auxin carriers and response factors (CsAUX1, CsGH3.5, CsARF17), ABA synthesis and catabolism (CsNCED1, CsNCED3 and CsCYP707A1) were upregulated by MT, in which CsNR1, CsNR2, CsAUX1, CsNCED3 and CsT5H were further induced in the presence of SNP in roots of cucumber seedlings. These observations indicated that NO act as a crucial factor in MT, alleviating nitrate stress through regulating the mechanism of root growth in cucumber seedlings.

The SlTPL3-SlWUS module regulates multi-locule formation in tomato by modulating auxin and gibberellin levels in the shoot apical meristem.

Malformed fruits depreciate a plant's market value. In tomato (Solanum lycopersicum), fruit malformation is associated with the multi-locule trait, which involves genes regulating shoot apical meristem (SAM) development. The expression pattern of TOPLESS3 (SlTPL3) throughout SAM development prompted us to investigate its functional significance via RNA interference (RNAi) and clustered regularly interspaced short palindromic repeats/CRISPR-associated nuclease 9 (Cas9)-mediated gene editing. Lower SlTPL3 transcript levels resulted in larger fruits with more locules and larger SAMs at the 5 d after germination (DAG5) stage. Differentially expressed genes in the SAM of wild-type (WT) and SlTPL3-RNAi plants, identified by transcriptome deep sequencing (RNA-seq), were enriched in the gibberellin (GA) biosynthesis and plant hormone signaling pathways. Moreover, exogenous auxin and paclobutrazol treatments rescued the multi-locule phenotype, indicating that SlTPL3 affects SAM size by mediating auxin and GA levels in the SAM. Furthermore, SlTPL3 interacted with WUSCHEL (SlWUS), which plays an important role in SAM size maintenance. We conducted RNA-seq and DNA affinity purification followed by sequencing (DAP-seq) analyses to identify the genes regulated by SlTPL3 and SlWUS in the SAM and to determine how they regulate SAM size. We detected 24 overlapping genes regulated by SlTPL3 and SlWUS and harboring an SlWUS-binding motif in their promoters. Furthermore, functional annotation revealed a notable enrichment for functions in auxin transport, auxin signal transduction, and GA biosynthesis. Dual-luciferase assays also revealed that SlTPL3 enhances SlWUS-mediated regulation (repression and activation) of SlPIN3 and SlGA2ox4 transcription, indicating that the SlTPL3-SlWUS module regulates SAM size by mediating auxin distribution and GA levels, and perturbations of this module result in enlarged SAM. These results provide novel insights into the molecular mechanism of SAM maintenance and locule formation in tomato and highlight the SlTPL3-SlWUS module as a key regulator.

Metabolomic and Transcriptomic Profiling Provide Novel Insights into Fruit Ripening and Ripening Disorder Caused by 1-MCP Treatments in Papaya.

Treatment with 1-methylcyclopropylene (1-MCP) is an effective technique to preserve fruits, but inappropriate treatment with 1-MCP causes a ripening disorder (rubbery texture) in papaya fruit. In this study, a combined metabolomic and transcriptomic analysis was conducted to reveal the possible mechanism of the ripening disorder caused by unsuitable 1-MCP in papaya. A total of 203 differential accumulated metabolites (DAMs) were identified in the metabolome analysis. Only 24 DAMs were identified in the control (CK) vs. the 1-MCP 2 h group, and they were primarily flavonoids. Ninety and 89 DAMs were identified in the CK vs. 1-MCP 16 h and 1-MCP 2 h vs. 1-MCP 16 h groups, respectively, indicating that long-term 1-MCP treatment severely altered the metabolites during fruit ripening. 1-MCP 16 h treatment severely reduced the number of metabolites, which primarily consisted of flavonoids, lipids, phenolic acids, alkaloids, and organic acids. An integrated analysis of RNA-Seq and metabolomics showed that various energy metabolites for the tricarboxylic acid cycle were reduced by long-term treatment with 1-MCP, and the glycolic acid cycle was the most significantly affected, as well as the phenylpropane pathway. These results provide valuable information for fruit quality control and new insight into the ripening disorder caused by unsuitable treatment with 1-MCP in papaya.

Identification of DELLA Genes and Key Stage for GA Sensitivity in Bolting and Flowering of Flowering Chinese Cabbage.

Flowering Chinese cabbage (Brassica campestris L. ssp. chinensis var. utilis Tsen et Lee) is an important and extensively cultivated vegetable in south China, and its stalk development is mainly regulated by gibberellin (GA). DELLA proteins negatively regulate GA signal transduction and may play an important role in determining bolting and flowering. Nevertheless, no systematic study of the DELLA gene family has been undertaken in flowering Chinese cabbage. In the present study, we found that the two-true-leaf spraying of gibberellin A3 (GA3) did not promote bolting but did promote flowering, whereas the three-true-leaf spraying of GA3 promoted both bolting and flowering. In addition, we identified five DELLA genes in flowering Chinese cabbage. All five proteins contained DELLA, VHYNP, VHIID, and SAW conserved domains. Protein-protein interaction results showed that in the presence of GA3, all five DELLA proteins interacted with BcGID1b (GA-INSENSITIVE DWARF 1b) but not with BcGID1a (GA-INSENSITIVE DWARF 1a) or BcGID1c (GA-INSENSITIVE DWARF 1c). Their expression analysis showed that the DELLA genes exhibited tissue-specific expression, and their reversible expression profiles responded to exogenous GA3 depending on the treatment stage. We also found that the DELLA genes showed distinct expression patterns in the two varieties of flowering Chinese cabbage. BcRGL1 may play a major role in the early bud differentiation process of different varieties, affecting bolting and flowering. Taken together, these results provide a theoretical basis for further dissecting the DELLA regulatory mechanism in the bolting and flowering of flowering Chinese cabbage.

Knockout of Auxin Response Factor SlARF4 Improves Tomato Resistance to Water Deficit.

Auxin response factors (ARFs) play important roles in various plant physiological processes; however, knowledge of the exact role of ARFs in plant responses to water deficit is limited. In this study, SlARF4, a member of the ARF family, was functionally characterized under water deficit. Real-time fluorescence quantitative polymerase chain reaction (PCR) and ß-glucuronidase (GUS) staining showed that water deficit and abscisic acid (ABA) treatment reduced the expression of SlARF4. SlARF4 was expressed in the vascular bundles and guard cells of tomato stomata. Loss of function of SlARF4 (arf4) by using Clustered Regularly Interspaced Short Palindromic Repeats/Cas 9 (CRISPR/Cas 9) technology enhanced plant resistance to water stress and rehydration ability. The arf4 mutant plants exhibited curly leaves and a thick stem. Malondialdehyde content was significantly lower in arf4 mutants than in wildtype plants under water stress; furthermore, arf4 mutants showed higher content of antioxidant substances, superoxide dismutase, actual photochemical efficiency of photosystem II (PSII), and catalase activities. Stomatal and vascular bundle morphology was changed in arf4 mutants. We identified 628 differentially expressed genes specifically expressed under water deficit in arf4 mutants; six of these genes, including ABA signaling pathway-related genes, were differentially expressed between the wildtype and arf4 mutants under water deficit and unlimited water supply. Auxin responsive element (AuxRE) elements were found in these genes' promoters indicating that SlARF4 participates in ABA signaling pathways by regulating the expression of SlABI5/ABF and SCL3, thereby influencing stomatal morphology and vascular bundle development and ultimately improving plant resistance to water deficit.

The SlHB8 Acts as a Negative Regulator in Stem Development and Lignin Biosynthesis.

The stem is an important organ in supporting plant body, transporting nutrients and communicating signals for plant growing. However, studies on the regulation of stem development in tomato are rather limited. In our study, we demonstrated that SlHB8 negatively regulated tomato stem development. SlHB8 belongs to homeo domain-leucine zipper Class III gene family transcription factors and expressed in all the organs examined including root, stem, leaves, flower, and fruit. Among these tissues, SlHB8 showed stable high expression level during tomato stem development. Overexpression of SlHB8 gene decreased stem diameter with inhibited xylem width and xylem cell layers, while loss of function of SlHB8gene increased the stem diameter and xylem width. The contents of lignin were decreased both in leaves and stems of SlHB8 overexpression plants. RNA-seq analysis on the stems of wild type and SlHB8 transgenic plants showed that the 116 DEGs (differential expressed genes) with reversible expression profiles in SlHB8-ox and SlHB8-cr plants were significantly enriched in the phenylpropanoid biosynthesis pathway and plant-pathogen pathway which were related to lignin biosynthesis and disease resistance. Meanwhile, the key genes involved in the lignin biosynthesis pathway such as SlCCR (cinnamoyl-CoA reductase), SlCYP73A14/C4H (cinnamate 4-hydroxylase), SlC3H (coumarate 3-hydroxylase) and SlCAD (cinnamoyl alcohol dehydrogenase) were down-regulated in both stem and leaves of SlHB8 overexpression plants, indicating a negative regulatory role of SlHB8 in the lignin biosynthesis and stem development.

Application of Scikit and Keras Libraries for the Classification of Iron Ore Data Acquired by Laser-Induced Breakdown Spectroscopy (LIBS).

Due to the complexity of, and low accuracy in, iron ore classification, a method of Laser-Induced Breakdown Spectroscopy (LIBS) combined with machine learning is proposed. In the research, we collected LIBS spectra of 10 iron ore samples. At the beginning, principal component analysis algorithm was employed to reduce the dimensionality of spectral data, then we applied k-nearest neighbor model, neural network model, and support vector machine model to the classification. The results showed that the accuracy of three models were 82.96%, 93.33%, and 94.07% respectively. The results also demonstrated that LIBS with machine learning model exhibits an excellent classification performance. Therefore, LIBS technique combined with machine learning can achieve a rapid, precise classification of iron ores, and can provide a completely new method for iron ores' selection in the metallurgical industry.

AUXIN RESPONSE FACTOR 2 Intersects Hormonal Signals in the Regulation of Tomato Fruit Ripening.

The involvement of ethylene in fruit ripening is well documented, though knowledge regarding the crosstalk between ethylene and other hormones in ripening is lacking. We discovered that AUXIN RESPONSE FACTOR 2A (ARF2A), a recognized auxin signaling component, functions in the control of ripening. ARF2A expression is ripening regulated and reduced in the rin, nor and nr ripening mutants. It is also responsive to exogenous application of ethylene, auxin and abscisic acid (ABA). Over-expressing ARF2A in tomato resulted in blotchy ripening in which certain fruit regions turn red and possess accelerated ripening. ARF2A over-expressing fruit displayed early ethylene emission and ethylene signaling inhibition delayed their ripening phenotype, suggesting ethylene dependency. Both green and red fruit regions showed the induction of ethylene signaling components and master regulators of ripening. Comprehensive hormone profiling revealed that altered ARF2A expression in fruit significantly modified abscisates, cytokinins and salicylic acid while gibberellic acid and auxin metabolites were unaffected. Silencing of ARF2A further validated these observations as reducing ARF2A expression let to retarded fruit ripening, parthenocarpy and a disturbed hormonal profile. Finally, we show that ARF2A both homodimerizes and interacts with the ABA STRESS RIPENING (ASR1) protein, suggesting that ASR1 might be linking ABA and ethylene-dependent ripening. These results revealed that ARF2A interconnects signals of ethylene and additional hormones to co-ordinate the capacity of fruit tissue to initiate the complex ripening process.

Auxin Response Factor SlARF2 Is an Essential Component of the Regulatory Mechanism Controlling Fruit Ripening in Tomato.

Ethylene is the main regulator of climacteric fruit ripening, by contrast the putative role of other phytohormones in this process remains poorly understood. The present study brings auxin signaling components into the mechanism regulating tomato fruit ripening through the functional characterization of Auxin Response Factor2 (SlARF2) which encodes a downstream component of auxin signaling. Two paralogs, SlARF2A and SlARF2B, are found in the tomato genome, both displaying a marked ripening-associated expression but distinct responsiveness to ethylene and auxin. Down-regulation of either SlARF2A or SlARF2B resulted in ripening defects while simultaneous silencing of both genes led to severe ripening inhibition suggesting a functional redundancy among the two ARFs. Tomato fruits under-expressing SlARF2 produced less climacteric ethylene and exhibited a dramatic down-regulation of the key ripening regulators RIN, CNR, NOR and TAGL1. Ethylene treatment failed to reverse the non-ripening phenotype and the expression of ethylene signaling and biosynthesis genes was strongly altered in SlARF2 down-regulated fruits. Although both SlARF proteins are transcriptional repressors the data indicate they work as positive regulators of tomato fruit ripening. Altogether, the study defines SlARF2 as a new component of the regulatory network controlling the ripening process in tomato.

Isomerization of Secondary Phosphirane into Terminal Phosphinidene Complexes: An Analogy between Monovalent Phosphorus and Transition Metals.

Secondary phosphirane complexes isomerize above 100 °C to give the corresponding terminal phosphinidene complexes, which can be trapped by alkenes and alkynes. This reaction is a rare instance of the isomerization of a P(III) derivative into a P(I) derivative. It appears to mimic the reductive elimination of alkanes from transition-alkylmetal hydrides.

SlARF4, an auxin response factor involved in the control of sugar metabolism during tomato fruit development.

Successful completion of fruit developmental programs depends on the interplay between multiple phytohormones. However, besides ethylene, the impact of other hormones on fruit quality traits remains elusive. A previous study has shown that down-regulation of SlARF4, a member of the tomato (Solanum lycopersicum) auxin response factor (ARF) gene family, results in a dark-green fruit phenotype with increased chloroplasts (Jones et al., 2002). This study further examines the role of this auxin transcriptional regulator during tomato fruit development at the level of transcripts, enzyme activities, and metabolites. It is noteworthy that the dark-green phenotype of antisense SlARF4-suppressed lines is restricted to fruit, suggesting that SlARF4 controls chlorophyll accumulation specifically in this organ. The SlARF4 underexpressing lines accumulate more starch at early stages of fruit development and display enhanced chlorophyll content and photochemical efficiency, which is consistent with the idea that fruit photosynthetic activity accounts for the elevated starch levels. SlARF4 expression is high in pericarp tissues of immature fruit and then undergoes a dramatic decline at the onset of ripening concomitant with the increase in sugar content. The higher starch content in developing fruits of SlARF4 down-regulated lines correlates with the up-regulation of genes and enzyme activities involved in starch biosynthesis, suggesting their negative regulation by SlARF4. Altogether, the data uncover the involvement of ARFs in the control of sugar content, an essential feature of fruit quality, and provide insight into the link between auxin signaling, chloroplastic activity, and sugar metabolism in developing fruit.

Genome-wide identification, phylogenetic analysis, expression profiling, and protein-protein interaction properties of TOPLESS gene family members in tomato.

Members of the TOPLESS gene family emerged recently as key players in gene repression in several mechanisms, especially in auxin perception. The TOPLESS genes constitute, in 'higher-plant' genomes, a small multigenic family comprising four to 11 members. In this study, this family was investigated in tomato, a model plant for Solanaceae species and fleshy fruits. Six open reading frames predicted to encode topless-like proteins (SlTPLs) containing the canonical domains (LisH, CTLH, and two WD40 repeats) were identified in the tomato genome. Nuclear localization was confirmed for all members of the SlTPL family with the exception SlTPL6, which localized at the cytoplasm and was excluded from the nucleus. SlTPL genes displayed distinctive expression patterns in different tomato organs, with SlTPL1 showing the highest levels of transcript accumulation in all tissues tested except in ripening fruit where SlTPL3 and SlTPL4 were the most prominently expressed. To gain insight into the specificity of the different TOPLESS paralogues, a protein-protein interaction map between TOPLESS and auxin/indole-3-acetic acid (Aux/IAA) proteins was built using a yeast two-hybrid approach. The PPI map enabled the distinction of two patterns: TOPLESS isoforms interacting with the majority of Aux/IAA, and isoforms with limited capacity for interaction with these protein partners. Interestingly, evolutionary analyses of the TOPLESS gene family revealed that the highly expressed isoforms (SlTPL1, SlTPL3, and SlTPL4) corresponded to the three TPL-related genes undergoing the strongest purifying selection, while the selection was much weaker for SlTPL6, which was expressed at a low level and encoded a protein lacking the capacity to interact with Aux/IAAs.

Ferroptosis: a new mechanism of traditional Chinese medicine for treating ulcerative colitis.

Ulcerative colitis (UC), a subtype of inflammatory bowel disease, manifests with symptoms such as abdominal pain, diarrhea, and mucopurulent, bloody stools. The pathogenesis of UC is not fully understood. At present, the incidence of UC has increased significantly around the world. Conventional therapeutic arsenals are relatively limited, with often poor efficacy and many adverse effects. In contrast, traditional Chinese medicine (TCM) holds promise due to their notable effectiveness, reduced recurrence rates, and minimal side effects. In recent years, significant progress has been made in the basic research on TCM for UC treatment. It has been found that the inhibition of ferroptosis through the intervention of TCM can significantly promote intestinal mucosal healing and reverse UC. The mechanism of action involves multiple targets and pathways. Aim of the review: This review summarizes the experimental studies on the targeted regulation of ferroptosis by TCM and its impact on UC in recent years, aiming to provide theoretical basis for the prevention, treatment, and further drug development for UC. Results: Ferroptosis disrupts antioxidant mechanisms in intestinal epithelial cells, damages the intestinal mucosa, and participates in the pathological process of UC. TCM acts on various pathways such as Nrf2/HO-1 and GSH/GPX4, blocking the pathological progression of ferroptosis in intestinal epithelial cells, inhibiting pathological damage to the intestinal mucosa, and thereby alleviating UC. Conclusion: The diverse array of TCM single herbs, extracts and herbal formulas facilitates selective and innovative research and development of new TCM methods for targeting UC treatment. Although progress has been made in studying TCM compound formulas, single herbs, and extracts, there are still many issues in clinical and basic experimental designs, necessitating further in-depth scientific exploration and research.

Deciphering the molecular mechanism of Bu Yang Huan Wu Decoction in interference with diabetic pulmonary fibrosis via regulating oxidative stress and lipid metabolism disorder.

BACKGROUND: Diabetes mellitus type 2 and pulmonary fibrosis have been found to be closely related in clinical practice. Diabetic pulmonary fibrosis (DPF) is a complication of diabetes mellitus, but its treatment has yet to be thoroughly investigated. Bu Yang Huan Wu Decoction (BYHWD) is a well-known traditional Chinese prescription that has shown great efficacy in treating pulmonary fibrosis with hypoglycemic and hypolipidemic effects. METHODS: The active ingredients of BYHWD and the corresponding targets were retrieved from the Traditional Chinese Medicine Systematic Pharmacology Database (TCMSP) and SymMap2. Disease-related targets were obtained from the GeneCard, OMIM and CTD databases. GO enrichment and KEGG pathway enrichment were carried out using the DAVID database. AutoDock Vina software was employed to perform molecular docking. Molecular dynamics simulations of proteinligand complexes were conducted by Gromacs. Animal experiments were further performed to validate the effects of BYHWD on the selected core targets, markers of oxidative stress, serum lipids, blood glucose and pulmonary fibrosis. RESULTS: A total of 84 active ingredients and 830 target genes were screened in BYHWD, among which 56 target genes intersected with DPF-related targets. Network pharmacological analysis revealed that the active ingredients can regulate target genes such as IL-6, TNF-α, VEGFA and CASP3, mainly through AGE-RAGE signaling pathway, HIF-1 signaling pathway and TNF signaling pathway. Molecular docking and molecular dynamics simulations suggested that IL6-astragaloside IV, IL6-baicalein, TNFα-astragaloside IV, and TNFα-baicalein docking complexes could bind stably. Animal experiments showed that BYHWD could reduce the expression of core targets such as VEGFA, CASP3, IL-6 and TNF-α. In addition, BYHWD could reduce blood glucose, lipid, and MDA levels in DPF while increasing the activities of SOD, CAT and GSH-Px. BYHWD attenuated the expression of HYP and collagen I, mitigating pathological damage and collagen deposition within lung tissue. CONCLUSIONS: BYHWD modulates lipid metabolism disorders and oxidative stress by targeting the core targets of IL6, TNF-α, VEGFA and CASP3 through the AGE-RAGE signaling pathway, making it a potential therapy for DPF.

Yuanzhi Powder inhibits tau pathology in SAMP8 mice: Mechanism research of a traditional Chinese formula against Alzheimer's disease.

ETHNOPHARMACOLOGICAL RELEVANCE: Yuanzhi Powder (YZP), a classical Chinese medicine formula, is good at tonifying heart-Qi and improving cognitive ability. YZP has been reported to show therapeutic effect on alleviating the symptoms of Alzheimer's disease (AD). AIM OF THE STUDY: This study was conducted to observe the effects of YZP on improving the cognitive abilities of SAMP8 mice, and explore the involved mechanisms on inhibiting the excessive accumulation of phosphorylated tau. MATERIAL AND METHODS: Thirty SAMP8 mice were randomly divided into five groups: AD group, AD + DO group, AD + YZP group, AD + LAC group and AD + LAC + YZP group. Age-matched SAMR1 mice were served as CTL group. AD + LAC group and AD + LAC + YZP group received 1 µg Lactacystin solution via intra-cerebroventricular injection. All mice (except the CTL group and AD + LAC group) were intragastrically administrated for 8 consecutive weeks. Then, the Morris Water Maze (MWM) test was conducted for evaluation of learning and memory abilities. The pathological changes of hippocampal CA1 were observed by Hematoxylin & eosin (H&E) staining. The expression of 26S proteasome in the hippocampus was measured by Western Blot (WB) and immunohistochemistry (IHC). The expressions of total tau (Tau5) and hyperphosphorylated tau (pS199, pT231 and pS396) were detected by WB. The aggregation of hyperphosphorylated tau and the binding ability of tau protein to microtubules were evaluated respectively by immunostaining and Thioflavin-S staining and double-label immunofluorescence. RESULTS: SAMP8 mice showed serious cognitive impairment in behavioral tests. However, treatment of YZP significantly ameliorated the cognitive deficits of SAMP8 mice. The H&E staining suggested that YZP could protect against neuronal loss in SAMP8 mice. The IHC and WB results showed that YZP increases 26S proteasome expression in SAMP8 mice and 26S proteasome expression was effectively inhibited by Lactacystin. Meanwhile, The WB results demonstrated that YZP can inhibit the expression of hyperphosphorylated tau (pT231, pS396 and pS199). Furthermore, the immunostaining and Thioflavin-S staining and double-label immunofluorescence results indicated that YZP attenuates the excessive aggregation of hyperphosphorylated tau and enhances the binding ability of tau to stabilize microtubules in SAMP8 mice. CONCLUSIONS: YZP could enhance cognitive performance and learning of AD, ameliorate tau pathology and significantly improve the binding ability of tau to microtubules, based potentially on inhibiting the excessive aggregation of hyperphosphorylated tau via the 26Sproteasome pathway but not necessarily the only one.