Search Results (199)

5F-ADB, MDMB-4en-PINACA and ADB-4en-PINACA are three potent indazole-carboxamide synthetic cannabinoids (SCs) that have been widely abused in recent years. However, the pharmacological research on these compounds remains limited, especially in vivo research data. The purpose of the present study was to investigate the pharmacological effects of 5F-ADB, MDMB-4en-PINACA and ADB-4en-PINACA in mice, comparing their in vivo effects with those caused by Δ⁹-tetrahydrocannabinol (Δ⁹-THC), the main psychoactive substance in cannabis. We evaluated the cannabinoid-specific pharmacological effects of 5F-ADB, MDMB-4en-PINACA and ADB-4en-PINACA using the tetrad assay (locomotion inhibition, hypothermia, analgesia and catalepsy). Then we conducted conditioned place preference (CPP) and precipitated withdrawal assay to assess the rewarding effect and physical dependence, with Δ⁹-THC as a positive control. The results showed that all of the three SCs exhibited potential tetrad effects in a dose-dependent manner, with median effective dose (ED₅₀) values ranging from 0.03 to 0.77 mg/kg. In the CPP tests, they all exhibited a significant biphasic effect of conditioned place preference (CPP) and conditioned place aversion (CPA). A significant increase in paw tremors and head twitches was observed in the rimonabant-precipitated withdrawal assay, indicating that the repeated administration of these SCs can lead to potential physical dependence. All effective doses were lower than Δ⁹-THC. These findings strongly suggested that the three SCs exhibited similar but stronger cannabinoid-specific tetrad effects, rewarding effect and physical dependence compared with Δ⁹-THC, indicating their high abuse potential and possible threats to human health. The rank order of abuse potential for these drugs was 5F-ADB > MDMB-4en-PINACA > ADB-4en-PINACA > Δ⁹-THC. Full article

Background: Anxiety disorders affect millions of people worldwide, but current treatments often have limited effectiveness and produce unpredictable responses in patients. This underscores the need for novel anxiolytics. While behavior-based screens are valuable for discovering new small molecules, high-throughput anxiety-related assays in vertebrates are lacking. The larval zebrafish dark avoidance, which corresponds to aversive responses to a predator’s shadow, can be reduced by known anxiolytics and involves neural pathways known to regulate human anxiety. Methods: Larval zebrafish exhibiting strong dark avoidance (SDA) have been characterized, reflecting pathological anxiety. We developed a high-throughput behavioral assay using a 96-well plate and showed that SDA larvae displayed significant dark avoidance in this setup. We tested known anxiolytics, Chlordiazepoxide and Buspirone, and found that they significantly reduced dark avoidance in this 96-well assay. We then tested a new candidate compound, Analgesic Screen 1 (AS1), previously shown to reverse avoidance of noxious stimuli such as high temperature in larval zebrafish. Results: The optimized 96-well plate assay reliably detected the anxiolytic activity of Chlordiazepoxide and Buspirone and revealed the effect of AS1 in reducing dark avoidance, thereby establishing the platform’s sensitivity and validity. Conclusions: This study demonstrates that the dark avoidance assay is scalable to a 96-well plate format in a small arena. This finding provides an effective platform for discovering novel anxiolytic compounds. Full article

The Chinese jujube (Ziziphus jujuba Mill.), an economically significant fruit tree native to China, is valued for both fresh and dried uses. In plants, 2n gametes serve as the fundamental basis for creating a sexual polyploid germplasm. This study investigated the 2n gametogenesis frequency in triploid hybrid jujubes through meiotic analysis of the hybrid strain Q161 and a two-year pollen analysis on hybrid progeny, assessing the natural 2n pollen frequencies to identify a high-2n-pollen germplasm and revealing the occurrence of 2n pollen. Meiotic analysis of the triploid hybrid Q161 (2n = 36) revealed cytological anomalies, including binucleate cells (22.80% abnormal tetrads), with natural 2n pollen production rates reaching 4.00% and 4.67% over two consecutive years. Scanning electron microscopy (SEM) revealed that the 2n pollen exhibited pronounced exine ornamentation with cerebroid sculpturing and tubercle-like structures at the apertures. Analysis of the triploid progeny for two consecutive years demonstrated a pollen viability of 30.45% and 23.83% (CV: 19. 39–29.69%), with the mean 2n pollen frequencies of 22.52% and 7.64%, peaking at 52.16% and 28.95% in elite individuals. Six triploid germplasm accessions with naturally elevated 2n pollen frequencies were identified. Under natural conditions, a triploid hybrid germplasm in Chinese jujube produces 2n pollen grains due to abnormal meiotic behavior, and a natural triploid germplasm with high pollen productivity was identified. This research provides a critical theoretical foundation for sexual polyploid breeding strategies. Full article

T40214 (STAT) and its recently investigated analogue STATB are G-quadruplex (G4) forming aptamers characterized by an unusually high percentage of C. The therapeutic potential of T40214 relies on its ability to inhibit the signalling pathway of STAT3, a protein frequently overexpressed in tumor cells. STAT adopts a dimeric 5′-5′ end-stacked quadruplex structure, characterized by parallel strands, three G-tetrads and three propeller-shaped loops formed by a cytidine residue. STATB folds in a very similar structure, apart from an additional cytidine bulge loop. Many studies suggest that thermal stability and topology of G4 can be significantly affected by C methylation, thus resulting in altered interaction of G4-binding proteins with these structures. Considering this, two series of STAT and STATB analogues containing a single 5-methyl-2′-deoxycytidine (mC) residue instead of canonical C nucleotide in the loop have been prepared and investigated by a combination of spectroscopic and electrophoretic techniques. CD, NMR and PAGE data clearly indicate that all derivatives adopt dimeric G4 strictly similar to that assumed by parent aptamers, but with higher stabilities. Furthermore, the resistance to nucleases and the antiproliferative activity of these mC-containing derivatives against HCT116 (human colorectal carcinoma) and T24 (human bladder carcinoma) cell lines have been evaluated. In most of the cases, STAT and STATB derivatives inhibit cell proliferation to different extents, although to a lesser degree than the unmodified parent sequences. All the data highlight the key role of the loops and indicate mC as a useful tool to contribute favorably to the stability of G4-forming aptamers without alteration of their topology, required for the biological activity. Full article

Background/Objectives: The oil grape (Idesia polycarpa), often called the “golden tree”, is an essential woody plant valued for its edible oil. Although its economic significance is recognized, the specifics of its chloroplast genome and evolutionary connections remain unclear. This study sequenced the chloroplast genome of I. polycarpa and performed a comparative analysis of its genome structure, genetic diversity, and phylogenetics using chloroplast data from related species. Methods: In this study, we sequenced and annotated the whole chloroplast genome of I. polycarpa via GISEQ-500 sequencing and de novo assembly. Results: The chloroplast genome of I. polycarpa exhibits a typical tetrad structure, with a length of 155,899 bp and a GC content of 36.78%. It comprises 130 unique genes, including 85 coding genes, 37 tRNAs, and eight rRNAs, showing notable conservation in gene composition and arrangement compared to closely related species. However, the inverted repeat region boundaries are narrower. Phylogenetic analysis showed strong relationships among I. polycarpa, Bennettiodendron brevipes, Poliothyrsis sinensis, Itoa orientalis, and Carrierea calycina within the Salicaceae family. Additionally, positive selection analysis revealed that rpl16, ycf1, rps18, and rpl22 are under significant selective pressure in related species, likely linked to adaptations for photosynthesis and environmental responses. Conclusions: This research provides vital molecular foundations for the conservation, classification, and enhancement of I. polycarpa germplasm resources, advancing the study of adaptive evolutionary mechanisms and broadening the genomic database for I. polycarpa. Full article

The current work records for the first time the rare-metal pegmatites with mixed NYF-LCT located at Wadi Sikait, south Eastern Desert of the Egyptian Nubian Shield. Most of the Sikait pegmatites are associated with sheared granite and are surrounded by an alteration zone cross-cutting through greisen bodies. Sikait pegmatites show zoned and complex types, where the outer wall zones are highly mineralized (Nb, Ta, Y, Th, Hf, REE, U) than the barren cores. They consist essentially of K-feldspar, quartz, micas (muscovite, lepidolite, and zinnwaldite), and less albite. They contain a wide range of accessory minerals, including garnet, columbite, fergusonite-(Y), cassiterite, allanite, monazite, bastnaesite (Y, Ce, Nd), thorite, zircon, beryl, topaz, apatite, and Fe-Ti oxides. In the present work, the discovery of Li-bearing minerals for the first time in the Wadi Sikait pegmatite is highly significant. Sikait pegmatites are highly mineralized and yield higher maximum concentrations of several metals than the associated sheared granite. They are strongly enriched in Li (900–1791 ppm), Nb (1181–1771 ppm), Ta (138–191 ppm), Y (626–998 ppm), Hf (201–303 ppm), Th (413–685 ppm), Zr (2592–4429 ppm), U (224–699 ppm), and ∑REE (830–1711 ppm). The pegmatites and associated sheared granite represent highly differentiated peraluminous rocks that are typical of post-collisional rare-metal bearing granites. They show parallel chondrite-normalized REE patterns, enriched in HREE relative to LREE [(La/Lu)_n = 0.04–0.12] and strongly negative Eu anomalies [(Eu/Eu*) = 0.03–0.10]. The REE patterns show an M-type tetrad effect, usually observed in granites that are strongly differentiated and ascribed to hydrothermal fluid exchange. The pegmatite has mineralogical and geochemical characteristics of the mixed NYF-LCT family and shows non-CHARAC behavior due to a hydrothermal effect. Late-stage metasomatism processes caused redistribution, concentrated on the primary rare metals, and drove the development of greisen and quartz veins along the fracture systems. The genetic relationship between the Sikait pegmatite and the surrounding sheared granite was demonstrated by the similarities in their geochemical properties. The source magmas were mostly derived from the juvenile continental crust of the Nubian Shield through partial melting and subsequently subjected to a high fractional crystallization degree. During the late hydrothermal stage, the exsolution of F-rich fluids transported some elements and locally increased their concentrations to the economic grades. The investigated pegmatite and sheared granite should be considered as a potential resource to warrant exploration for REEs and other rare metals. Full article

Cytosine-rich DNA sequences form i-motif structures associated with various cellular functions including gene regulation. DNA sequences containing consecutive C residues are widely deemed essential for i-motif formation; however, some sequences lacking C-tracts have been reported to form minimal i-motif structures. We systematically investigated the variability in the minimal i-motif-forming DNA sequence comprising two TCGTTCCGT sequence units, which forms two C:C⁺ pairs and two G:C:G:T base tetrads. A comprehensive analysis of structural stability by DNA thermal melting temperature measurements revealed that oligonucleotides disrupting the formation of the base tetrad or its stacking interactions with a C:C⁺ pair prevent stable i-motif formation, and modifications to the sequence context and length of the lateral loops are difficult. This study further demonstrated that spermine effectively restores the stability reduction caused by creating a bulge, long loop, or dangling end within the minimal i-motif structure, which is less pronounced in the C-rich i-motif. The results suggest that the formation of minimal i-motifs with various sequences is facilitated in polyamine-rich environments, such as the nucleus of mammalian cells. These findings are valuable for identifying potential i-motif-forming sites lacking C-tracts in genomes and provide insights into the electrostatic interactions between i-motif structures and biological polyamines. Full article

Lilies are one of the most popular ornamental flowers in the world. However, the abundant pollen produced in their anthers causes significant inconvenience for producers and consumers. Pollen abortion induced by molecular breeding techniques is one of the effective ways to solve this problem. In this study, the LoTDF1 gene, which is involved in regulating lily anther development, was identified and cloned from lily anthers based on transcriptome data. The open reading frame of LoTDF1 is 936 bp and encodes a protein with 311 amino acids. Multiple sequence alignment and phylogenetic tree analysis revealed that the LoTDF1 protein contained a conserved R2R3 domain, belonging to the MYB transcription factor family. Subcellular localization and transcriptional activation assays demonstrated that LoTDF1 localized to the nucleus and functioned as a transcription activator. The transcriptional activation domain was located within the last 195 amino acids (117–311a) of the C-terminus, and there may be more than one transcriptional activation domain in the region. The expression level of the LoTDF1 gene was highest during the pollen mother cell (PMC) stage of lily anther development (2 cm anther), followed by the tetrad stage (4 cm anther). In situ hybridization experiments further confirmed that LoTDF1 transcripts were predominantly localized in PMCs, tapetal cells, middle layer cells, dyads, and tetrads. The experiment data suggest that LoTDF1 plays a critical role in regulating early anther development in lily. LoTDF1 could be a promising candidate gene for molecular breeding strategies aimed at developing pollen-free lily cultivars to enhance commercial and consumer appeal. Full article

Background: Tomatoes are self-pollinating plants, and successful fruit set depends on the production of functional pollen within the same flower. Our previous studies have shown that the ‘Black Vernissage’ tomato variety exhibits greater resilience to heat stress in terms of pollen productivity compared to the ‘Micro-Tom’ variety. Pollen productivity is determined by meiotic activity during microsporogenesis and the development of free microspores during gametogenesis. This study focused on identifying heat stress (HS)-induced proteomes in pollen mother cells (PMCs) and microspores. Methods: Tomato plants were grown under two temperature conditions: 26 °C (non-heat-treated control) and 37 °C (heat-treated). Homogeneous cell samples of meiotic PMCs (prior to the tetrad stage) and free microspores were collected using laser capture microdissection (LCM). The heat-induced proteomes were identified using tandem mass tag (TMT)–quantitative proteomics analysis. Results: The enrichment of the meiotic cell cycle in PMCs and the pre-mitotic process in free microspores confirmed the correlation between proteome expression and developmental stage. Under HS, PMCs in both tomato varieties were enriched with heat shock proteins (HSPs). However, the ‘Black Vernissage’ variety exhibited a greater diversity of HSP species and a higher level of enrichment compared to the ‘Micro-Tom’ variety. Additionally, several proteins involved in gene expression and protein translation were downregulated in PMCs and microspores of both varieties. In the PMC proteomes, the relative abundance of proteins showed no significant differences between the two varieties under normal conditions, with very few exceptions. However, HS induced significant differential expression both within and between the varieties. More importantly, these heat-induced differentially abundant proteins (DAPs) in PMCs are directly involved in meiotic cell division, including the meiosis-specific protein ASY3 (Solyc01g079080), the cell division protein kinase 2 (Solyc11g070140), COP9 signalosome complex subunit 1 (Solyc01g091650), the kinetochore protein ndc80 (Solyc01g104570), MORC family CW-type zinc finger 3 (Solyc02g084700), and several HSPs that function in protecting the fidelity of the meiotic processes, including the DNAJ chaperone (Solyc04g009770, Solyc05g055160), chaperone protein htpG (Solyc04g081570), and class I and class II HSPs. In the microspores, most of the HS-induced DAPs were consistently observed across both varieties, with only a few proteins showing significant differences between them under heat stress. These HS-induced DAPs include proteases, antioxidant proteins, and proteins related to cell wall remodeling and the generation of pollen exine. Conclusions: HS induced more dynamic proteomic changes in meiotic PMCs compared to microspores, and the inter-varietal differences in the PMC proteomes align with the effects of HS on pollen productivity observed in the two varieties. This research highlights the importance of the cell-type-specific proteomics approach in identifying the molecular mechanisms that are critical for the pollen developmental process under elevated temperature conditions. Full article

Chinese cabbage (Brassica rapa L. ssp. pekinensis), an important traditional vegetable indigenous to China, is a typical cross-pollinated Brassica crop exhibiting pronounced heterosis. However, its small flower organs make artificial pollination for hybrid seed production highly challenging. The use of male-sterile lines has emerged as a crucial approach in hybrid seed production. Therefore, understanding the genetic and molecular mechanisms underlying male sterility in Chinese cabbage holds profound theoretical and economic importance and is pivotal for advancing Chinese cabbage crossbreeding. Here, cytological comparative analysis of anthers from sterile line 366-2S and fertile line 366-2F revealed abnormalities in 366-2S during the late tetrad stage, including delayed tapetum degradation and the aggregation of tetrad microspores without separation, which prevented pollen production and caused male sterility. Construction of the F₂ segregating population, with 366-2S as the female parent and genetically diverse fertile material Y636-9 as the male parent, indicated that male sterility in 366-2S is controlled by a single recessive gene. Using bulked segregant analysis sequencing and kompetitive allele-specific polymerase chain reaction (KASP) technology, the sterile gene was mapped to 65 kb between the PA11 and PA13 markers, with 11 genes in the candidate region. Functional annotation, expression, and sequence variation analyses identified BraA09g012710.3C, encoding acyl-CoA synthetase 5, as a candidate gene for 366-2S male sterility. Quantitative real-time polymerase chain reaction analysis revealed minimal expression of BraA09g012710.3C in 366-2S but high expression in the flower buds of 366-2F. Further analysis of candidate gene DNA sequences identified a large deletion encompassing BraA09g012710.3C, BraA09g012720.3C, BraA09g012730.3C, and BraA09g012740.3C in sterile line 366-2S (A09: 7452347–7479709). Cloning and verification of the other three deleted genes in the F₂ population via agarose gel electrophoresis confirmed their presence in F₂ sterile individuals, indicating that their deletion was not associated with male sterility, underscoring BraA09g012710.3C as the key gene driving male sterility in 366-2S. Full article

Cytoplasmic male sterility (CMS) and dominant genic male sterility (DGMS) both result in the inability to produce or release functional pollen, making them pivotal systems in the hybridization breeding programs of Brassica crops such as cabbage (B. oleracea var. capitata). However, the underling molecular mechanisms are still largely unexplored. This study integrated transcriptomic and metabolomic analyses of cabbage DGMS line, Ogura CMS line, and the maintainer line to uncover the molecular mechanisms underlying these sterility types. The joint analysis predominantly identified significantly enriched pathways, including carbohydrate metabolism, flavonoid biosynthesis, and phenylpropanoid pathways between the MS lines and the maintainer. Especially, the CMS line exhibited a broader range of metabolic perturbations, with a total of 3556 significantly differentially expressed genes (DEGs) and 439 differentially accumulated metabolites (DAMs) detected, particularly in the vitamin B6 metabolism pathway, which showed significant alterations. Given the differences in the inactivation period of microspores in CMS and DGMS lines, we found that DEGs unique to DGMS and maintainer line, such as BoGRPs and BoLTPs, primarily regulate fertility development before the unicellular stage. The DEGs shared between CMS_vs_maintainer and DGMS_vs_maintainer mainly govern microspore development after release from the tetrad, such as BoHXK1 and BoIDH. Additionally, the DEGs unique to CMS_vs_maintainer may contribute to other damage in floral organs beyond male fertility, potentially leading to severe bud abortion, such as BoPNPO. These findings provide a comprehensive framework for understanding the molecular mechanisms of male sterility and offer valuable insights into future breeding strategies in cruciferous vegetables. Full article

β-1,3-Glucanases (Glu) are key enzymes involved in plant defense and physiological processes through the hydrolysis of β-1,3-glucans. This study provides a comprehensive analysis of the β-1,3-glucanase gene family in wolfberry (Lycium barbarum), including their chromosomal distribution, evolutionary relationships, and expression profiles. A total of 58 Glu genes were identified, distributed across all 12 chromosomes. Evolutionary analysis revealed six distinct branches within wolfberry and nine distinct branches when compared with Arabidopsis thaliana. Expression analysis showed that 45 Glu genes were expressed in berries, with specific genes also being expressed in flowers and leaves. Notably, LbaGlu28 exhibited significant expression during the tetrad stage of pollen development and was localized in the cell wall. These findings provide valuable insights into the functional significance of Glu genes in wolfberry, highlighting their roles in development and their potential involvement in reproductive processes, particularly in pollen development. Full article

G-quadruplex (G4), an important secondary structure of nucleic acids, is polymorphic in structure. G4 monomers can associate with each other to form multimers, which show better application performance than monomers in some aspects. G4 dimers, the simplest and most widespread multimeric structures, are often used as a representative for studying multimers. RHAU, a G4 ligand, has been reported to recognize G4 dimers. However, there are few reports focusing on interactions between RHAU and different G4 dimers. In this work, interactions between RHAU peptide and six G4 dimers were investigated by size-exclusion chromatography (SEC). It was revealed that compared to the hybrid G4 monomer, the hybrid tandem unstacked G4 dimer could form special binding sites, leading to a weak interaction with RHAU. It was also found that the steric hindrance at terminal G-tetrads of a special Z-G4 structure greatly weakened their interactions with RHAU. Additionally, RHAU exhibited stronger interactions with intermolecular stacked/interlocked parallel dimers than with intramolecular tandem stacked parallel dimers. This work enriches the understanding of interactions between RHAU and G4 dimers, which is conducive to the elucidation of G4 polymorphism, and provides a strong reference for studying G4 multimer–peptide interactions. Full article

Male sterility is an essential trait for minimizing costs and ensuring seed purity in the production of tomato hybrid seeds. However, its application in commercial breeding faces challenges such as instability, low stigma exsertion rates, and the lack of molecular markers to facilitate the efficient backcrossing of male sterile loci into elite tomato lines. This study characterized the male sterile-24 (ms-24) mutant, which demonstrated exserted stigmas and complete male sterility. Our histological examination revealed that ms-24 anthers displayed dysfunctional development of the tetrads and tapetum. The ms-24 locus was finely mapped to a 149 kb interval containing 22 putative genes. Among these, Solyc02g079810, also known as MS-10, encodes a basic helix–loop–helix (bHLH) transcription factor essential for tapetum and pollen development. A sequencing analysis revealed an approximately 4.9 kb retrotransposon insertion in the first exon of the MS-10 gene in ms-24. Allelism tests confirmed that ms-24 was allelic to ms-10, which is also allelic to ms-35 and ms-36. The same retrotransposon insertion was also identified in the ms-10 mutant, and a similar insertion of retrotransposon was detected in the second exon of the MS-10 gene in the ms-36 mutant. Based on these retrotransposon sequences, codominant insertion–deletion (InDel) markers, MS-24I and MS-35I, were developed for the precise identification of the ms-24, ms-10, ms-35, and ms-36 alleles. These findings establish a foundational basis for subsequent investigations into the molecular mechanisms underlying male sterility and enhance the selection process of male sterile lines in tomato hybrid breeding programs. Full article

We demonstrate that a short oligonucleotide complementary to a G-quadruplex domain can invade this iconic, noncanonical DNA secondary structure in ways that profoundly influence the properties and differential occupancies of the resulting DNA polymorphic products. Our spectroscopic mapping of the conformational space of the associated reactants and products, both before and after strand invasion, yield unanticipated outcomes which reveal several overarching features. First, strand invasion induces the disruption of DNA secondary structural elements in both the invading strand (which can assume an iDNA tetrad structure) and the invaded species (a G-quadruplex). The resultant cascade of coupled alterations represents a potential pathway for the controlled unfolding of kinetically trapped DNA states, a feature that may be characteristic of biological regulatory mechanisms. Furthermore, the addition of selectively designed, exogenous invading oligonucleotides can enable the manipulation of noncanonical DNA conformations for biomedical applications. Secondly, our results highlight the importance of metastability, including the interplay between slower and faster kinetic processes in determining preferentially populated DNA states. Collectively, our data reveal the importance of sample history in defining state populations, which, in turn, determine preferred pathways for further folding steps, irrespective of the position of the thermodynamic equilibrium. Finally, our spectroscopic data reveal the impact of topological constraints on the differential stabilities of base-paired domains. We discuss how our collective observations yield insights into the coupled and uncoupled cascade of strand-invasion-induced transformations between noncanonical DNA forms, potentially as components of molecular wiring diagrams that regulate biological processes. Full article