MicroProteinDB: A database to provide knowledge on sequences, structures and function of ncRNA-derived microproteins.

Omics-based technologies have revolutionized our comprehension of microproteins encoded by ncRNAs, revealing their abundant presence and pivotal roles within complex functional landscapes. Here, we developed MicroProteinDB (http://bio-bigdata.hrbmu.edu.cn/MicroProteinDB), which offers and visualizes the extensive knowledge to aid retrieval and analysis of computationally predicted and experimentally validated microproteins originating from various ncRNA types. Employing prediction algorithms grounded in diverse deep learning approaches, MicroProteinDB comprehensively documents the fundamental physicochemical properties, secondary and tertiary structures, interactions with functional proteins, family domains, and inter-species conservation of microproteins. With five major analytical modules, it will serve as a valuable knowledge for investigating ncRNA-derived microproteins.

Generate an AZFa deleted human embryonic stem cell line.

Y chromosome deletion and karyotype abnormalities are commonly associated with congenital non-obstructive azoospermia, impairing spermatogenesis. Specifically, the deletion of the Y chromosome Azoospermia factor a (AZFa) has been identified in infertile males with severely impaired spermatogenesis. AZFa, encompassing megabase-scale of the Y chromosome region, poses challenges in modeling AZFa deletion-related male infertility using gene editing tools. Here, we successfully created an AZFa-deleted human embryonic stem cell line utilizing the CRISPR/Cas9 gene editing tool. Our analysis indicates the AZFa-deleted stem cell line holds promise for differentiation into ectoderm, mesoderm, and endoderm, highlighting its potential for further comprehensive study.

The structures of two polysaccharides from Lepidium meyenii and their immunomodulatory effects via activating NF-κB signaling pathway.

Lepidium meyenii Walp., also known as the "Peruvian national treasure", is a popular functional food in the daily lives of Peruvian people due to its bioactive with main polysaccharides. However, studies on polysaccharides isolated from Lepidium meyenii were few. Two new highly heterogeneous polysaccharides, MCP-1a and MCP-2b, were isolated and purified from the tuber of Lepidium meyenii. The structure characterization revealed that MCP-1a primarily consisted of D-Glc and had a molecular weight of 6.6 kDa. Its backbone was composed of 1,4,6-α-D-Glc, while branches feature T-α-L-Ara, 1,5-α-L-Ara, and T-α-D-Glc attached to the O-6 positions. MCP-2b was a rare arabinogalactan with a molecular weight of 49.4 kDa. Interestingly, the backbone of MCP-2b was composed of 1,6-ß-D-Gal, 1,3,6-ß-D-Gal with a few 1,3-ß-D-GlcpA-4-OMe units inserted. Side chains of MCP-2b were mainly composed of 1,3-ß-D-Gal, T-ß-D-Gal, T-α-L-Ara, 1,5-α-L-Ara, with trace amounts of 1,4-ß-D-Glc and T-ß-D-Glc. The bioactivity assay results revealed that MCP-1a and MCP-2b increased the release of NO, IL-1ß, TNF-α, and IL-6 from RAW 264.7 cells at concentrations ranging from 50 µg/mL to 400 µg/mL. Furthermore, MCP-1a and MCP-2b could promote the expression of key transcription factors (IκB-α, p-IκB-α, p65, and p-p65) in the NF-κB pathway, indicating that MCP-1a and MCP-2b had potential immunomodulatory activities.

PeCIN8 expression correlates with flower size and resistance to yellow leaf disease in Phalaenopsis orchids.

BACKGROUND: The orchid industry has seen a recent surge in export values due to the floral morphology and versatile applications of orchids in various markets for medicinal, food additive, and cosmetic usages. However, plant-related diseases, including the yellow leaf disease caused by Fusarium solani, have caused significant losses in the production value of Phalaenopsis (up to 30%). RESULTS: In this study, 203 Phalaenopsis cultivars were collected from 10 local orchid nurseries, and their disease severity index and correlation with flower size were evaluated. Larger flowers had weaker resistance to yellow leaf disease, and smaller flowers had stronger resistance. For the genetic relationship of disease resistance to flower size, the genetic background of all cultivars was assessed using OrchidWiz Orchid Database Software and principal component analysis. In addition, we identified the orthologous genes of BraTCP4, namely PeIN6, PeCIN7, and PeCIN8, which are involved in resistance to pathogens, and analyzed their gene expression. The expression of PeCIN8 was significantly higher in the most resistant cultivars (A7403, A11294, and A2945) relative to the most susceptible cultivars (A10670, A6390, and A10746). CONCLUSIONS: We identified a correlation between flower size and resistance to yellow leaf disease in Phalaenopsis orchids. The expression of PeCIN8 may regulate the two traits in the disease-resistant cultivars. These findings can be applied to Phalaenopsis breeding programs to develop resistant cultivars against yellow leaf disease.

ZSCAN4 interacts with PARP1 to promote DNA repair in mouse embryonic stem cells.

BACKGROUND: In eukaryotic cells, DNA double strand breaks (DSB) are primarily repaired by canonical non-homologous end joining (c-NHEJ), homologous recombination (HR) and alternative NHEJ (alt-NHEJ). Zinc finger and SCAN domain containing 4 (ZSCAN4), sporadically expressed in 1-5% mouse embryonic stem cells (mESCs), is known to regulate genome stability by promoting HR. RESULTS: Here we show that ZSCAN4 promotes DNA repair by acting with Poly (ADP-ribose) polymerase 1 (PARP1), which is a key member of the alt-NHEJ pathway. In the presence of PARP1, ZSCAN4-expressing mESCs are associated with lower extent of endogenous or chemical induced DSB comparing to ZSCAN4-negative ones. Reduced DSBs associated with ZSCAN4 are abolished by PARP1 inhibition, achieved either through small molecule inhibitor or gene knockout in mESCs. Furthermore, PARP1 binds directly to ZSCAN4, and the second ⍺-helix and the fourth zinc finger motif of ZSCAN4 are critical for this binding. CONCLUSIONS: These data reveal that PARP1 and ZSCAN4 have a protein-protein interaction, and shed light on the molecular mechanisms by which ZSCAN4 reduces DSB in mESCs.

Isolation and characterization of ZK002, a novel dual function snake venom protein from Deinagkistrodon acutus with anti-angiogenic and anti-inflammatory properties.

Introduction: Pathological angiogenesis, the abnormal or excessive generation of blood vessels, plays an important role in many diseases including cancer, diabetic retinopathy, psoriasis, and arthritis. Additionally, increasing evidence supports the close linkage between angiogenesis and inflammation. Snake venoms are a rich natural source of biologically active molecules and carry rich potential for the discovery of anti-angiogenic and anti-inflammatory modulators. Methods: Here, we isolated and purified a novel protein, ZK002, from the venom of the snake Deinagkistrodon acutus, and investigated its anti-angiogenic and anti-inflammatory activities and mechanisms. Results: ZK002 was identified as a 30 kDa heterodimeric protein of α and ß chains, which exhibited anti-angiogenic activity in various in vitro assays. Mechanistically, ZK002 inhibited activation of VEGF signaling and related mediators including eNOS, p38, LIMK, and HSP27. ZK002 also upregulated the metalloproteinase inhibitor TIMP3 and inhibited components of the VEGF-induced signaling cascade, PPP3R2 and SH2D2A. The anti-angiogenic activity of ZK002 was confirmed in multiple in vivo models. ZK002 could also inhibit the in vitro expression of pro-inflammatory cytokines, as well as in vivo inflammation in the carrageenin-induced edema rat model. Conclusion: Our findings highlight the potential for further development of ZK002 as a dual function therapeutic against diseases with involvement of pathogenic angiogenesis and chronic inflammation.

Structural characterizations and α-glucosidase inhibitory activities of four Lepidium meyenii polysaccharides with different molecular weights.

Four polysaccharides (MCPa, MCPb, MCPc, MCPd) were obtained from Lepidium meyenii Walp. Their structures were characterized by chemical and instrumental methods including total sugar, uronic acid and protein content determination, UV, IR and NMR spectroscopy, as well as monosaccharide composition determination and methylation analyses. Four polysaccharides were a group of glucans with different molecular weights ranging from 3.12 to 14.4 kDa, and shared a similar backbone chain consisting of (1→4)-glucose linkages with branches attached to C-3 and C-6. Furthermore, bioactivity assay showed that MCPs had concentration-dependent inhibitory activity on α-glucosidase. MCPb (Mw = 10.1 kDa) and MCPc (Mw = 5.62 kDa) with moderate molecular weights exhibited higher inhibitory activity compared with MCPa and MCPd.

NMR-tracking for 15,16-seco-cycloartane triterpenes from Cimicifuga acerina.

Twelve undescribed 15,16-seco-cycloartane triterpenoids, 15,16-seco-cimiterpenes C-N, as well as five previously reported analogs were isolated by NMR-tracking methods from the rhizomes of Cimicifuga acerina (Sieb. et Zucc.) Tanaka. Among them, 15,16-seco-cimiterpenes C-N were the first 15,16-seco-cycloartane triterpenoids featuring acetal or hemiacetal structures at C-15. The chemical structures of 15,16-seco-cimiterpenes C-N were determined based on comprehensive spectroscopic analysis, chemical method, and comparison with the previous literature data. After that, all these compounds were evaluated for their lipid-lowering effects on 3T3-L1 adipocytes.15,16-seco-cimiterpene D was found to exhibit a comparable reducing lipid effect at the concentration of 50 µM, with an inhibition rate at 35.96%.

MS/MS molecular networking-guided in-depth profiling of triterpenoid saponins from the fruit of Eleutherococcus senticosus and their neuroprotectivity evaluation.

INTRODUCTION: Eleutherococcus senticosus fruit (ESF) is a natural health supplement resource that has been extensively applied as a tonic for the nervous system. The structures and neural bioactivities of triterpenoid saponins (TS), which are the major constituents of ESF, have not been comprehensively analyzed thus far. OBJECTIVE: We conducted a complete in-depth MS/MS molecular networking (MN)-based targeted analysis of TS from the crude extract of ESF and investigated its neuroprotective value. METHODS: An MS/MS MN-guided strategy was used to rapidly present a series of precursor ions (PIs) of TS in a compound cluster as TS-targeted information used in the discovery and characterization of TS. In addition, a prepared TS-rich fraction of ESF was assayed for its restraining effects on ß-amyloid-induced inhibition of neurite outgrowth. RESULTS: A total of 87 TS were discovered using a PI tracking strategy, 28 of which were characterized as potentially undescribed structures according to their high-resolution MS values. Furthermore, the TS-rich fraction can significantly reduce ß-amyloid-induced damage to neural networks by promoting the outgrowth of neurites and axons. CONCLUSION: Our findings reveal the richness of TS in ESF and will accelerate their application in the treatment of neurodegenerative diseases.

Identification and localization of morphological feature-specific metabolites in Reynoutria multiflora roots.

Reynoutria multiflora roots are a classical herbal medicine with unique nourishing therapeutic effects. Anomalous vascular bundle (AVB) forming "cloudy brocade patterns" is a typical morphological feature of R. multiflora roots and has been empirically linked to its quality classification. However, scientific evidence, especially for AVB-specific specialised metabolites, has not been comprehensively revealed thus far. Herein, desorption electrospray ionization-mass spectrometry imaging (DESI-MSI) analysis was applied to carry out an in situ analysis of specialised metabolites distributed specifically at the AVB and cork of R. multiflora roots. To enlarge the scope of compounds by DESI detection, various solvent systems including acetone, acetonitrile, methanol, and water were used to assist in the discoveries of 40 specialised metabolites with determined localization. A series of bioactive constituents including stilbenes, flavonoids, anthraquinones, alkaloids, and naphthalenes were found specifically around the brocade patterns. Notably, phospholipids were detected from R. multiflora roots by in situ analysis for the first time and were found mainly in the phloem of AVB (PAB). This is the first study to use gradient solvent systems in DESI-MSI analysis to locate the specialised metabolites distribution. The discovery of feature-specific compounds will bridge the empirical identification to precision quality control of R. multiflora roots.

High-density genetic map and genome-wide association studies of aesthetic traits in Phalaenopsis orchids.

Phalaenopsis spp. represent the most popular orchids worldwide. Both P. equestris and P. aphrodite are the two important breeding parents with the whole genome sequence available. However, marker-trait association is rarely used for floral traits in Phalaenopsis breeding. Here, we analyzed markers associated with aesthetic traits of Phalaenopsis orchids by using genome-wide association study (GWAS) with the F1 population P. Intermedia of 117 progenies derived from the cross between P. aphrodite and P. equestris. A total of 113,517 single nucleotide polymorphisms (SNPs) were identified in P. Intermedia by using genotyping-by-sequencing with the combination of two different restriction enzyme pairs, Hinp1 I/Hae III and Apek I/Hae III. The size-related traits from flowers were negatively related to the color-related traits. The 1191 SNPs from Hinp1 I/ Hae III and 23 simple sequence repeats were used to establish a high-density genetic map of 19 homolog groups for P. equestris. In addition, 10 quantitative trait loci were highly associated with four color-related traits on chromosomes 2, 5 and 9. According to the sequence within the linkage disequilibrium regions, 35 candidate genes were identified and related to anthocyanin biosynthesis. In conclusion, we performed marker-assisted gene identification of aesthetic traits with GWAS in Phalaenopsis orchids.

Memory enhancement effect of saponins from Eleutherococcus senticosus leaves and blood-brain barrier-permeated saponins profiling using a pseudotargeted monitoring strategy.

Dried Eleutherococcus senticosus leaves (ESL), also known as Siberian ginseng tea, are beneficial for human neural disorders. Our previous studies showed that the aqueous extract of ESL enhanced memory in mice, and its saponin fraction (ESL-SAP) exhibited promising neuroprotective activities in vitro; however, the in vivo neurally related effect, bioactive material basis, and possible mechanism of action of ESL-SAP have not been investigated. Here, a series of memory and learning tests were carried out, and the results evidenced a significant enhancement effect of ESL-SAP. Furthermore, an in vivo saponin library-guided pseudotargeted strategy was established to support the rapid monitoring of 26 blood-brain barrier (BBB)-permeated saponins from ESL-SAP-administered rats. A further network pharmacology analysis was conducted on BBB-permeated compounds, which indicated that the in vivo mechanism of ESL-SAP might be effective through multiple targets and pathways, such as the AGE-RAGE signaling pathway and PI3K-Akt signaling pathway, to exert neuroprotective effects. Moreover, the molecular docking experiments demonstrated that key BBB-transferred saponins primarily interacted with targets HRAS, MAPK1, and MAPK8 to produce the neuroprotective effect.

Delivery of CRISPR/Cas9 Plasmid DNA by Hyperbranched Polymeric Nanoparticles Enables Efficient Gene Editing.

Gene editing nucleases such as CRISPR/Cas9 have enabled efficient and precise gene editing in vitro and hold promise of eventually achieving in vivo gene editing based therapy. However, a major challenge for their use is the lack of a safe and effective virus-free system to deliver gene editing nuclease elements. Polymers are a promising class of delivery vehicle due to their higher safety compared to currently used viral vectors, but polymers suffer from lower transfection efficiency. Polymeric vectors have been used for small nucleotide delivery but have yet to be used successfully with plasmid DNA (pDNA), which is often several hundred times larger than small nucleotides, presenting an engineering challenge. To address this, we extended our previously reported hyperbranched polymer (HP) delivery system for pDNA delivery by synthesizing several variants of HPs: HP-800, HP-1.8K, HP-10K, HP-25K. We demonstrate that all HPs have low toxicity in various cultured cells, with HP-25K being the most efficient at packaging and delivering pDNA. Importantly, HP-25K mediated delivery of CRISPR/Cas9 pDNA resulted in higher gene-editing rates than all other HPs and Lipofectamine at several clinically significant loci in different cell types. Consistently, HP-25K also led to more robust base editing when delivering the CRISPR base editor "BE4-max" pDNA to cells compared with Lipofectamine. The present work demonstrates that HP nanoparticles represent a promising class of vehicle for the non-viral delivery of pDNA towards the clinical application of gene-editing therapy.

Exploring the Effects of Magnesium Deficiency on the Quality Constituents of Hydroponic-Cultivated Tea (Camellia sinensis L.) Leaves.

Magnesium (Mg) plays important roles in photosynthesis, sucrose partitioning, and biomass allocation in plants. However, the specific mechanisms of tea plant response to Mg deficiency remain unclear. In this study, we investigated the effects of Mg deficiency on the quality constituents of tea leaves. Our results showed that the short-term (7 days) Mg deficiency partially elevated the concentrations of polyphenols, free amino acids, and caffeine but decreased the contents of chlorophyll and Mg. However, long-term (30 days) Mg-deficient tea displayed decreased contents of these constituents. Particularly, Mg deficiency increased the index of catechins' bitter taste and the ratio of total polyphenols to total free amino acids. Moreover, the transcription of key genes involved in the biosynthesis of flavonoid, caffeine, and theanine was differentially affected by Mg deficiency. Additionally, short-term Mg deficiency induced global transcriptome change in tea leaves, in which a total of 2522 differentially expressed genes were identified involved in secondary metabolism, amino acid metabolism, and chlorophyll metabolism. These results may help to elucidate why short-term Mg deficiency partially improves the quality constituents of tea, while long-term Mg-deficient tea may taste more bitter, more astringent, and less umami.

Breviscapine: A Review on its Phytochemistry, Pharmacokinetics and Therapeutic Effects.

Breviscapine is one of the extracts of several flavonoids of Erigeron breviscapus. Scutellarin is the main active component of breviscapine, and the qualitative or quantitative criteria as well. Scutellarin and its analogs share a similar skeleton of the flavonoids. Breviscapine has been widely used in the treatment of cerebral infarction and its sequelae, cerebral thrombus, coronary heart disease (CHD), and angina pectoris. Breviscapine has a broad spectrum of pharmacological activities, such as increasing blood flow, improving microcirculation, dilating blood vessels, decreasing blood viscosity, promoting fibrinolysis, inhibiting platelet aggregation, and thrombosis formation, etc. In addition, breviscapine and its analogs have significant value for drug research and development because of the superiority of those significant bioactivities. Furthermore, an increasing number of pharmacokinetic studies have explored the mechanism of scutellarin and its analogs. To provide a comprehensive understanding of the current research on breviscapine, scutellarin, and the analogs, the structural features, distribution situation, preparation method, content determination method, clinical applications, pharmacological action as well as pharmacokinetics are summarized in the present review.

Biodegradation of aromatic pollutants meets synthetic biology.

Ubiquitously distributed microorganisms are natural decomposers of environmental pollutants. However, because of continuous generation of novel recalcitrant pollutants due to human activities, it is difficult, if not impossible, for microbes to acquire novel degradation mechanisms through natural evolution. Synthetic biology provides tools to engineer, transform or even re-synthesize an organism purposefully, accelerating transition from unable to able, inefficient to efficient degradation of given pollutants, and therefore, providing new solutions for environmental bioremediation. In this review, we described the pipeline to build chassis cells for the treatment of aromatic pollutants, and presented a proposal to design microbes with emphasis on the strategies applied to modify the target organism at different level. Finally, we discussed challenges and opportunities for future research in this field.

Development of the Nude Rabbit Model.

Loss-of-function mutations in the forkhead box N1 (FOXN1) gene lead to nude severe combined immunodeficiency, a rare inherited syndrome characterized by athymia, severe T cell immunodeficiency, congenital alopecia, and nail dystrophy. We recently produced FOXN1 mutant nude rabbits (NuRabbits) by using CRISPR-Cas9. Here we report the establishment and maintenance of the NuRabbit colony. NuRabbits, like nude mice, are hairless, lack thymic development, and are immunodeficient. To demonstrate the functional applications of NuRabbits in biomedical research, we show that they can successfully serve as the recipient animals in xenotransplantation experiments using human induced pluripotent stem cells or tissue-engineered blood vessels. Our work presents the NuRabbit as a new member of the immunodeficient animal model family. The relatively large size and long lifespan of NuRabbits offer unique applications in regenerative medicine, cancer research, and the study of a variety of other human conditions, including immunodeficiency.

Clinical significance of LSECtin and its association with PVR in non-small-cell lung cancer patients.

BACKGROUND: Liver and lymph node sinusoidal endothelial cell C-type lectin (LSECtin) is one of the new generation immune checkpoint ligand molecules and plays an important role in the immune environment. Poliovirus receptor (PVR), as another immunosuppression-related molecule, is upregulated in various malignant tumors. However, the clinical value of LSECtin and the correlation of LSECtin with PVR in non-small-cell lung cancer (NSCLC) remain to be elucidated. In this study, a retrospective study was performed to address these issues. METHODS: This retrospective study included 98 patients with NSCLC. Immunohistochemistry (IHC) was used to detect the expression of LSECtin and PVR in the paraffin-embedded tumor tissue specimens. LSECtin was analyzed for associations with the survival rate and overall survival (OS) of the subjects. The mRNA expression of LSECtin and PVR was assessed using the expression data from The Cancer Genome Atlas (TCGA) database. Clinical characteristics, prognosis, and the expression of LSECtin and PVR were included in the statistical analysis. RESULTS: High positive rates of LSECtin were found in the patients with NSCLC who were nonsmokers, at advanced stages, or had lung adenocarcinoma. Patients with positive LSECtin expression had a significantly lower survival rate (P=0.008) and shorter OS (P=0.017) than those with negative LSECtin. Significant correlation was found between the LSECtin and PVR expression in the patients with NSCLC (P<0.001). CONCLUSIONS: The increased expression of LSECtin was related to the poor prognosis of patients with NSCLC after tumor resection and has the potential value for predicting the prognosis of these patients. The positive correlation between LSECtin and PVR in NSCLC provides a theoretical basis for the future combination therapy of immune checkpoints.

MiCas9 increases large size gene knock-in rates and reduces undesirable on-target and off-target indel edits.

Gene editing nuclease represented by Cas9 efficiently generates DNA double strand breaks at the target locus, followed by repair through either the error-prone non-homologous end joining or the homology directed repair pathways. To improve Cas9's homology directed repair capacity, here we report the development of miCas9 by fusing a minimal motif consisting of thirty-six amino acids to spCas9. MiCas9 binds RAD51 through this fusion motif and enriches RAD51 at the target locus. In comparison to spCas9, miCas9 enhances double-stranded DNA mediated large size gene knock-in rates, systematically reduces off-target insertion and deletion events, maintains or increases single-stranded oligodeoxynucleotides mediated precise gene editing rates, and effectively reduces on-target insertion and deletion rates in knock-in applications. Furthermore, we demonstrate that this fusion motif can work as a "plug and play" module, compatible and synergistic with other Cas9 variants. MiCas9 and the minimal fusion motif may find broad applications in gene editing research and therapeutics.

Adjuvant herbal therapy for targeting susceptibility genes to Kawasaki disease: An overview of epidemiology, pathogenesis, diagnosis and pharmacological treatment of Kawasaki disease.

BACKGROUND: Kawasaki disease (KD) is a self-limiting acute systemic vasculitis occur mainly in infants and young children under 5 years old. Although the use of acetylsalicylic acid (AAS) in combination with intravenous immunoglobulin (IVIG) remains the standard therapy to KD, the etiology, genetic susceptibility genes and pathogenic factors of KD are still un-elucidated. PURPOSE: Current obstacles in the treatment of KD include the lack of standard clinical and genetic markers for early diagnosis, possible severe side effect of AAS (Reye's syndrome), and the refractory KD cases with resistance to IVIG therapy, therefore, this review has focused on introducing the current advances in the identification of genetic susceptibility genes, environmental factors, diagnostic markers and adjuvant pharmacological intervention for KD. RESULTS: With an overall update in the development of KD from different aspects, our current bioinformatics data has suggested CASP3, CD40 and TLR4 as the possible pathogenic factors or diagnostic markers of KD. Besides, a list of herbal medicines which may work as the adjunct therapy for KD via targeting different proposed molecular targets of KD have also been summarized. CONCLUSION: With the aid of modern pharmacological research and technology, it is anticipated that novel therapeutic remedies, especially active herbal chemicals targeting precise clinical markers of KD could be developed for accurate diagnosis and treatment of the disease.