scDOT: enhancing single-cell RNA-Seq data annotation and uncovering novel cell types through multi-reference integration.

The proliferation of single-cell RNA-seq data has greatly enhanced our ability to comprehend the intricate nature of diverse tissues. However, accurately annotating cell types in such data, especially when handling multiple reference datasets and identifying novel cell types, remains a significant challenge. To address these issues, we introduce Single Cell annotation based on Distance metric learning and Optimal Transport (scDOT), an innovative cell-type annotation method adept at integrating multiple reference datasets and uncovering previously unseen cell types. scDOT introduces two key innovations. First, by incorporating distance metric learning and optimal transport, it presents a novel optimization framework. This framework effectively learns the predictive power of each reference dataset for new query data and simultaneously establishes a probabilistic mapping between cells in the query data and reference-defined cell types. Secondly, scDOT develops an interpretable scoring system based on the acquired probabilistic mapping, enabling the precise identification of previously unseen cell types within the data. To rigorously assess scDOT's capabilities, we systematically evaluate its performance using two diverse collections of benchmark datasets encompassing various tissues, sequencing technologies and diverse cell types. Our experimental results consistently affirm the superior performance of scDOT in cell-type annotation and the identification of previously unseen cell types. These advancements provide researchers with a potent tool for precise cell-type annotation, ultimately enriching our understanding of complex biological tissues.

MGST3 regulates BACE1 protein translation and amyloidogenesis by controlling the RGS4-mediated AKT signaling pathway.

Microsomal glutathione transferase 3 (MGST3) regulates eicosanoid and glutathione metabolism. These processes are associated with oxidative stress and apoptosis, suggesting that MGST3 might play a role in the pathophysiology of Alzheimer's disease. Here, we report that knockdown (KD) of MGST3 in cell lines reduced the protein level of beta-site amyloid precursor protein cleaving enzyme 1 (BACE1) and the resulting amyloidogenesis. Interestingly, MGST3 KD did not alter intracellular reactive oxygen species level but selectively reduced the expression of apoptosis indicators which could be associated with the receptor of cysteinyl leukotrienes, the downstream metabolites of MGST3 in arachidonic acid pathway. We then showed that the effect of MGST3 on BACE1 was independent of cysteinyl leukotrienes but involved a translational mechanism. Further RNA-seq analysis identified that regulator of G-protein signaling 4 (RGS4) was a target gene of MGST3. Silencing of RGS4 inhibited BACE1 translation and prevented MGST3 KD-mediated reduction of BACE1. The potential mechanism was related to AKT activity, as the protein level of phosphorylated AKT was significantly reduced by silencing of MGST3 and RGS4, and the AKT inhibitor abolished the effect of MGST3/RGS4 on phosphorylated AKT and BACE1. Together, MGST3 regulated amyloidogenesis by controlling BACE1 protein expression, which was mediated by RGS4 and downstream AKT signaling pathway.

Fungal invasion-induced accumulation of salicylic acid promotes anthocyanin biosynthesis through MdNPR1-MdTGA2.2 module in apple fruits.

In the field, necrosis area induced by pathogens is usually surrounded by a red circle in apple fruits. However, the underlying molecular mechanism of this phenomenon remains unclear. In this study, we demonstrated that accumulated salicylic acid (SA) induced by fungal infection promoted anthocyanin biosynthesis through MdNPR1-MdTGA2.2 module in apple (Malus domestica). Inoculating apple fruits with Valsa mali or Botryosphaeria dothidea induced a red circle surrounding the necrosis area, which mimicked the phenotype observed in the field. The red circle accumulated a high level of anthocyanins, which was positively correlated with SA accumulation stimulated by fungal invasion. Further analysis showed that SA promoted anthocyanin biosynthesis in a dose-dependent manner in both apple calli and fruits. We next demonstrated that MdNPR1, a master regulator of SA signaling, positively regulated anthocyanin biosynthesis in both apple and Arabidopsis. Moreover, MdNPR1 functioned as a co-activator to interact with and enhance the transactivation activity of MdTGA2.2, which could directly bind to the promoters of anthocyanin biosynthetic and regulatory genes to promote their transcription. Suppressing expression of either MdNPR1 or MdTGA2.2 inhibited coloration of apple fruits, while overexpressing either of them significantly promoted fruit coloration. Finally, we revealed that silencing either MdNPR1 or MdTGA2.2 in apple fruits repressed SA-induced fruit coloration. Therefore, our data determined that fungal-induced SA promoted anthocyanin biosynthesis through MdNPR1-MdTGA2.2 module, resulting in a red circle surrounding the necrosis area in apple fruits.

Transcription factor PbNAC71 regulates xylem and vessel development to control plant height.

Dwarfism is an important agronomic trait in fruit breeding programs. However, the germplasm resources required to generate dwarf pear (Pyrus spp.) varieties are limited. Moreover, the mechanisms underlying dwarfism remain unclear. In this study, "Yunnan" quince (Cydonia oblonga Mill.) had a dwarfing effect on "Zaosu" pear. Additionally, the dwarfism-related NAC transcription factor gene PbNAC71 was isolated from pear trees comprising "Zaosu" (scion) grafted onto "Yunnan" quince (rootstock). Transgenic Nicotiana benthamiana and pear OHF-333 (Pyrus communis) plants overexpressing PbNAC71 exhibited dwarfism, with a substantially smaller xylem and vessel area relative to the wild-type controls. Yeast one-hybrid, dual-luciferase, chromatin immunoprecipitation-qPCR, and electrophoretic mobility shift assays indicated that PbNAC71 downregulates PbWalls are thin 1 expression by binding to NAC-binding elements in its promoter. Yeast two-hybrid assays showed that PbNAC71 interacts with the E3 ubiquitin ligase PbRING finger protein 217 (PbRNF217). Furthermore, PbRNF217 promotes the ubiquitin-mediated degradation of PbNAC71 by the 26S proteasome, thereby regulating plant height as well as xylem and vessel development. Our findings reveal a mechanism underlying pear dwarfism and expand our understanding of the molecular basis of dwarfism in woody plants.

Hypoxia-inducible factor prolyl hydroxylase inhibitor alleviates heatstroke-induced acute kidney injury by activating BNIP3-mediated mitophagy.

Hypoxia-induced inflammation and apoptosis are important pathophysiological features of heat stroke-induced acute kidney injury (HS-AKI). Hypoxia-inducible factor (HIF) is a key protein that regulates cell adaptation to hypoxia. HIF-prolyl hydroxylase inhibitor (HIF-PHI) stabilizes HIF to increase cell adaptation to hypoxia. Herein, we reported that HIF-PHI pretreatment significantly improved renal function, enhanced thermotolerance, and increased the survival rate of mice in the context of HS. Moreover, HIF-PHI could alleviate HS-induced mitochondrial damage, inflammation, and apoptosis in renal tubular epithelial cells (RTECs) by enhancing mitophagy in vitro and in vivo. By contrast, mitophagy inhibitors Mdivi-1, 3-MA, and Baf-A1 reversed the renoprotective effects of HIF-PHI. Mechanistically, HIF-PHI protects RTECs from inflammation and apoptosis by enhancing Bcl-2 adenovirus E18 19-kDa-interacting protein 3 (BNIP3)-mediated mitophagy, while genetic ablation of BNIP3 attenuated HIF-PHI-induced mitophagy and abolished HIF-PHI-mediated renal protection. Thus, our results indicated that HIF-PHI protects renal function by upregulating BNIP3-mediated mitophagy to improve HS-induced inflammation and apoptosis of RTECs, suggesting HIF-PHI as a promising therapeutic agent to treat HS-AKI.

Loss of Smooth Muscle Tenascin-X Inhibits Vascular Remodeling Through Increased TGF-ß Signaling.

BACKGROUND: Vascular smooth muscle cells (VSMCs) are highly plastic. Vessel injury induces a phenotypic transformation from differentiated to dedifferentiated VSMCs, which involves reduced expression of contractile proteins and increased production of extracellular matrix and inflammatory cytokines. This transition plays an important role in several cardiovascular diseases such as atherosclerosis, hypertension, and aortic aneurysm. TGF-ß (transforming growth factor-ß) is critical for VSMC differentiation and to counterbalance the effect of dedifferentiating factors. However, the mechanisms controlling TGF-ß activity and VSMC phenotypic regulation under in vivo conditions are poorly understood. The extracellular matrix protein TN-X (tenascin-X) has recently been shown to bind TGF-ß and to prevent it from activating its receptor. METHODS: We studied the role of TN-X in VSMCs in various murine disease models using tamoxifen-inducible SMC-specific knockout and adeno-associated virus-mediated knockdown. RESULTS: In hypertensive and high-fat diet-fed mice, after carotid artery ligation as well as in human aneurysmal aortae, expression of Tnxb, the gene encoding TN-X, was increased in VSMCs. Mice with smooth muscle cell-specific loss of TN-X (SMC-Tnxb-KO) showed increased TGF-ß signaling in VSMCs, as well as upregulated expression of VSMC differentiation marker genes during vascular remodeling compared with controls. SMC-specific TN-X deficiency decreased neointima formation after carotid artery ligation and reduced vessel wall thickening during Ang II (angiotensin II)-induced hypertension. SMC-Tnxb-KO mice lacking ApoE showed reduced atherosclerosis and Ang II-induced aneurysm formation under high-fat diet. Adeno-associated virus-mediated SMC-specific expression of short hairpin RNA against Tnxb showed similar beneficial effects. Treatment with an anti-TGF-ß antibody or additional SMC-specific loss of the TGF-ß receptor reverted the effects of SMC-specific TN-X deficiency. CONCLUSIONS: In summary, TN-X critically regulates VSMC plasticity during vascular injury by inhibiting TGF-ß signaling. Our data indicate that inhibition of vascular smooth muscle TN-X may represent a strategy to prevent and treat pathological vascular remodeling.

Metabolomic and transcriptomic analyses of peach leaves and fruits in response to pruning.

BACKGROUND: Pruning is an important cultivation management option that has important effects on peach yield and quality. However, the effects of pruning on the overall genetic and metabolic changes in peach leaves and fruits are poorly understood. RESULTS: The transcriptomic and metabolomic profiles of leaves and fruits from trees subjected to pruning and unpruning treatments were measured. A total of 20,633 genes and 622 metabolites were detected. Compared with those in the control, 1,127 differentially expressed genes (DEGs) and 77 differentially expressed metabolites (DEMs) were identified in leaves from pruned and unpruned trees (pdLvsupdL), whereas 423 DEGs and 29 DEMs were identified in fruits from the pairwise comparison pdFvsupdF. The content of three auxin analogues was upregulated in the leaves of pruned trees, the content of all flavonoids detected in the leaves decreased, and the expression of almost all genes involved in the flavonoid biosynthesis pathway decreased. The phenolic acid and amino acid metabolites detected in fruits from pruned trees were downregulated, and all terpenoids were upregulated. The correlation analysis revealed that DEGs and DEMs in leaves were enriched in tryptophan metabolism, auxin signal transduction, and flavonoid biosynthesis. DEGs and DEMs in fruits were enriched in flavonoid and phenylpropanoid biosynthesis, as well as L-glutamic acid biosynthesis. CONCLUSIONS: Pruning has different effects on the leaves and fruits of peach trees, affecting mainly the secondary metabolism and hormone signalling pathways in leaves and amino acid biosynthesis in fruits.

FUDNC1-dependent mitophagy ameliorate motor neuron death in an amyotrophic lateral sclerosis mouse model.

Amyotrophic lateral sclerosis (ALS) is one of the most common neurodegenerative diseases, yet effective treatment is lacking. Moreover, the underlying pathomechanisms of ALS remain unclear, with impaired mitophagy function being increasingly recognized as a contributing factor. FUN14 domain-containing protein 1 (FUNDC1) is an autophagy receptor localized to the outer mitochondrial membrane and a mitochondrial membrane protein that mediates mitophagy and therefore considered as important factor in neurodegenerative diseases. However, its specific role in ALS is not yet clear. Therefore, this study aimed to investigate the regulatory role of FUNDC1 in ALS and determine its regulatory mechanisms. ALS transgenic mice were obtained and maintained under standard conditions. Cell lines were generated by stable transfection with hSOD1G93A or control vectors. Mice received intrathecal injections of AAV9 vectors expressing FUNDC1 or EGFP. Motor function was assessed through behavioral tests, and histological and immunostaining analyses were performed. Colocalization analysis was conducted in transfected cells, and protein expression was evaluated via western blotting. We first observed that FUNDC1 was significantly downregulated in the spinal cord tissues of SOD1G93A mice. FUNDC1 overexpression considerably improved locomotor activity and prolonged survival time in SOD1G93A mice. Mechanistically, reduced expression of FUNDC1 resulted in decreased mitophagy, as indicated by decreased recruitment through LC3 in SOD1G93A mice and cellular models. Consequently, this led to increased mitochondrial accumulation and cell apoptosis, exacerbating the ALS phenotype. Furthermore, we identified transcription factor FOXD3 as an essential upstream factor of FUNDC1, resulting in reduced transcription of FUNDC1 in ALS lesions. This study suggests a novel strategy of targeting FUNDC1-mediated mitophagy for developing therapeutic interventions to mitigate disease progression and improve outcomes for ALS patients.

Disulfidptosis: A new type of cell death.

Disulfidptosis is a novel form of cell death that is distinguishable from established programmed cell death pathways such as apoptosis, pyroptosis, autophagy, ferroptosis, and oxeiptosis. This process is characterized by the rapid depletion of nicotinamide adenine dinucleotide phosphate (NADPH) in cells and high expression of solute carrier family 7 member 11 (SLC7A11) during glucose starvation, resulting in abnormal cystine accumulation, which subsequently induces andabnormal disulfide bond formation in actin cytoskeleton proteins, culminating in actin network collapse and disulfidptosis. This review aimed to summarize the underlying mechanisms, influencing factors, comparisons with traditional cell death pathways, associations with related diseases, application prospects, and future research directions related to disulfidptosis.

Epidermodysplasia Verruciformis and Vδ2 γδ T-cell Expansion in STK4 Deficiency.

The clinical penetrance of infectious diseases varies considerably among patients with inborn errors of immunity (IEI), even for identical genetic defects. This variability is influenced by pathogen exposure, healthcare access and host-environment interactions. We describe here a patient in his thirties who presented with epidermodysplasia verruciformis (EV) due to infection with a weakly virulent beta-papillomavirus (HPV38) and CD4+ T-cell lymphopenia. The patient was born to consanguineous parents living in the United States. Exome sequencing identified a previously unknown biallelic STK4 stop-gain mutation (p.Trp425X). The patient had no relevant history of infectious disease during childhood other than mild wart-like lesion on the skin, but he developed diffuse large B-cell lymphoma (DLBCL) and EBV viremia with a low viral load in his thirties. Despite his low CD4+ T-cell count, the patient had normal counts of CD3+ cells, predominantly double-negative T cells (67.4%), which turned out to be Vδ2+ γδ T cells. γδ T-cell expansion has frequently been observed in the 33 reported cases with STK4 deficiency. The Vδ2 γδ T cells of this STK4-deficient patient are mostly CD45RA-CD27+CCR7+ central memory γδT cells, and their ability to proliferate in response to T-cell activation was impaired, as was that of CD4+ T cells. In conclusion, γδ T-cell expansion may act as a compensatory mechanism to combat viral infection, providing immune protection in immunocompromised individuals.

TBK1, a prioritized drug repurposing target for amyotrophic lateral sclerosis: evidence from druggable genome Mendelian randomization and pharmacological verification in vitro.

BACKGROUND: There is a lack of effective therapeutic strategies for amyotrophic lateral sclerosis (ALS); therefore, drug repurposing might provide a rapid approach to meet the urgent need for treatment. METHODS: To identify therapeutic targets associated with ALS, we conducted Mendelian randomization (MR) analysis and colocalization analysis using cis-eQTL of druggable gene and ALS GWAS data collections to determine annotated druggable gene targets that exhibited significant associations with ALS. By subsequent repurposing drug discovery coupled with inclusion criteria selection, we identified several drug candidates corresponding to their druggable gene targets that have been genetically validated. The pharmacological assays were then conducted to further assess the efficacy of genetics-supported repurposed drugs for potential ALS therapy in various cellular models. RESULTS: Through MR analysis, we identified potential ALS druggable genes in the blood, including TBK1 [OR 1.30, 95%CI (1.19, 1.42)], TNFSF12 [OR 1.36, 95%CI (1.19, 1.56)], GPX3 [OR 1.28, 95%CI (1.15, 1.43)], TNFSF13 [OR 0.45, 95%CI (0.32, 0.64)], and CD68 [OR 0.38, 95%CI (0.24, 0.58)]. Additionally, we identified potential ALS druggable genes in the brain, including RESP18 [OR 1.11, 95%CI (1.07, 1.16)], GPX3 [OR 0.57, 95%CI (0.48, 0.68)], GDF9 [OR 0.77, 95%CI (0.67, 0.88)], and PTPRN [OR 0.17, 95%CI (0.08, 0.34)]. Among them, TBK1, TNFSF12, RESP18, and GPX3 were confirmed in further colocalization analysis. We identified five drugs with repurposing opportunities targeting TBK1, TNFSF12, and GPX3, namely fostamatinib (R788), amlexanox (AMX), BIIB-023, RG-7212, and glutathione as potential repurposing drugs. R788 and AMX were prioritized due to their genetic supports, safety profiles, and cost-effectiveness evaluation. Further pharmacological analysis revealed that R788 and AMX mitigated neuroinflammation in ALS cell models characterized by overly active cGAS/STING signaling that was induced by MSA-2 or ALS-related toxic proteins (TDP-43 and SOD1), through the inhibition of TBK1 phosphorylation. CONCLUSIONS: Our MR analyses provided genetic evidence supporting TBK1, TNFSF12, RESP18, and GPX3 as druggable genes for ALS treatment. Among the drug candidates targeting the above genes with repurposing opportunities, FDA-approved drug-R788 and AMX served as effective TBK1 inhibitors. The subsequent pharmacological studies validated the potential of R788 and AMX for treating specific ALS subtypes through the inhibition of TBK1 phosphorylation.

Simultaneously Engineering the First and Second Coordination Shells of Single Iron Catalysts for Enhanced Oxygen Reduction.

The atomically dispersed Fe-N4 active site presents enormous potential for various renewable energy conversions. Despite its already remarkable catalytic performance, the local atomic microenvironment of each Fe atom can be regulated to further enhance its efficiency. Herein, a novel conceptual strategy that utilizes a simple salt-template polymerization method to simultaneously adjust the first coordination shell (Fe-N3S1) and second coordination shell (C-S-C, a structure similar to thiophene) of Fe-N4 isolated atoms is proposed. Theoretical studies suggest that this approach can redistribute charge density in the MN4 moiety, lowering the d-band center of the metal site. This weakens the binding of oxygenated intermediates, enhancing oxygen reduction reaction (ORR) activity when compared to only implementing coordination shell regulation. Based on the above discovery, a single Fe atom electrocatalyst with the optimal Fe-N3S1-S active moiety incorporated in nitrogen, sulfur co-doped graphene (Fe-SAc/NSG) is designed and synthesized. The Fe-SAc/NSG catalyst exhibits excellent alkaline ORR activity, exceeding benchmark Pt/C and most Fe-SAc ORR electrocatalysts, as well as superior stability in Zn-air battery. This work aims to pave the way for creating highly active single metal atom catalysts through the localized regulation of their atomic structure.

A Noncentrosymmetric Metal-Free Borophosphate: Achieving a Large Birefringence and Excellent Stability by Covalent-Linkage.

Organic-inorganic hybrid linear and nonlinear optical (NLO) materials have received increasingly wide spread attention in recent years. Herein, the first hybrid noncentrosymmetric (NCS) borophosphate, (C5H6N)2B2O(HPO4)2 (4PBP), is rationally designed and synthesized by a covalent-linkage strategy. 4-pyridyl-boronic acid (4 PB) is considered as a bifunctional unit, which may effectively improve the optical properties and stability of the resultant material. On the one hand, 4 PB units are covalently linked with PO3(OH) groups via strong B-O-P connections, which significantly enhances the thermal stability of 4PBP (decomposition at 321, vs lower 200 °C of most of hybrid materials). On the other hand, the planar π-conjugated C5H6N units and their uniform layered arrangements represent large structural anisotropy and hyperpolarizability, achieving the largest birefringence (0.156 @ 546 nm) in the reported borophosphates and a second-harmonic generation response (0.7 × KDP). 4PBP also exhibits a wide transparency range (0.27-1.50 µm). This work not only provides a promising birefringent material, but also offers a practical covalent-attachment strategy for the rational design of new high-performance optical materials.

Transfemoral transcatheter aortic valve replacement with VitaFlowTM valve for pure native aortic regurgitation in patients with high surgical risk: Rationale and design of a prospective, multicenter, and randomized SEASON-AR trial.

BACKGROUND: Previous studies primarily demonstrated that transfemoral transcatheter aortic valve replacement (TAVR) with self-expanding valve appeared to be a safe and feasible treatment for patients with pure native aortic regurgitation (AR). However, the routine application of transfemoral TAVR for pure AR patients lacks support from randomized trials. TRIAL DESIGN: SEASON-AR trial is a prospective, multicenter, randomized, controlled, parallel-group, open-label trial, involving at least 20 sites in China, aiming to enroll 210 patients with pure native severe AR and high surgical risk. All enrolled patients are randomly assigned in a 1:1 fashion to undergo transfemoral TAVR with VitaFlowTM valve and receive guideline-directed medical therapy (GDMT) or to receive GDMT alone. The primary endpoint is the rate of major adverse cardiac events (MACE) at 12 months after the procedure, defined by the composite of all-cause mortality, disabling stroke, and rehospitalization for heart failure. The major secondary endpoints encompass various measures, including procedure-related complications, device success, 6-minute walk distance, and the occurrence of each individual component of the primary endpoint. After hospital discharge, follow-up was conducted through clinical visits or telephone contact at 1, 6, and 12 months. The follow-up will continue annually until 5 years after the index procedure to assess the long-term outcomes. CONCLUSION: SEASON-AR trial is the first study designed to investigate the clinical efficacy and safety of transfemoral TAVR with a self-expanding valve in patients with pure native severe AR with inoperable or high-risk, as compared to medical treatment only.

PCSK9 Inhibitor Added to High-Intensity Statin Therapy to Prevent Cardiovascular Events in Patients with Acute Coronary Syndrome after Percutaneous Coronary Intervention: A Randomized, Double- Blind, Placebo-Controlled, Multicenter SHAWN Study.

BACKGROUND: It is currently uncertain whether the combination of a proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitor and high-intensity statin treatment can effectively reduce cardiovascular events in patients with acute coronary syndrome (ACS) who have undergone percutaneous coronary intervention (PCI) for culprit lesions. METHODS: This study protocol describes a double-blind, randomized, placebo-controlled, multicenter study aiming to investigate the efficacy and safety of combining a PCSK9 inhibitor with high-intensity statin therapy in patients with ACS following PCI. A total of 1212 patients with ACS and multiple lesions will be enrolled and randomly assigned to receive either PCSK9 inhibitor plus high-intensity statin therapy or high-intensity statin monotherapy. The randomization process will be stratified by sites, diabetes, initial presentation and use of stable (≥4 weeks) statin treatment at presentation. PCSK 9 inhibitor or its placebo is injected within 4 hours after PCI for the culprit lesion. The primary endpoint is the composite of cardiovascular death, myocardial infarction, stroke, re-hospitalization due to ACS or heart failure, or any ischemia-driven coronary revascularization at one-year follow-up between two groups. Safety endpoints mean PCSK 9 inhibitor and statin intolerance. CONCLUSION: The SHAWN study has been specifically designed to evaluate the effectiveness and safety of adding a PCSK9 inhibitor to high-intensity statin therapy in patients who have experienced ACS following PCI. The primary objective of this study is to generate new evidence regarding the potential benefits of combining a PCSK9 inhibitor with high-intensity statin treatment in reducing cardiovascular events among these patients.

Immune infiltration is associated with pan-cancer prognostic biomarker RING finger protein 187.

Cancer is associated with the highest mortality rate globally. While life-saving screening and treatments exist, better awareness is needed. RNF187, an E3 ligase regulating biological processes, belongs to the RING domain-containing E3 ligase family. RNF187 may serve as an oncogene due to abnormal expression in tumors. However, its association with immune infiltration and prognosis across various cancers remains unclear. We searched several databases including TCGA, GTE x, CCLE, TIMER, and GSEA. R software was used to evaluate RNF187 differential expression, survival, pathology stage, DNA methylation, tumor mutational burden (TMB), microsatellite instability (MSI), gene co-expression analysis, mismatch repairs (MMRs), tumor microenvironment (TME), and immune cell infiltration. Clinicopathological data were collected, and immunohistochemistry was used to verify RNF187 expression in tumor tissues. RNF187 expression was up-regulated in various cancers compared to that in normal tissues and associated with poor patient outcomes. Dysregulation of RNF187 expression in multiple cancer types was strongly correlated with DNA methylation, MMR, MSI, and TMB. RNF187 could interact with different immune cells in cancers. Biomarkers associated with RNF187 may be helpful for prognosis and immunology in treating pan-cancer patients.

Cloning and functional identification of apple LATERAL ORGAN BOUNDARY DOMAIN 3 (LBD3) transcription factor in the regulation of drought and salt stress.

MAIN CONCLUSION: Overexpression of MdLBD3 in Arabidopsis reduced sensitivity to salt and drought stresses and was instrumental in promoting early flowering. Salt and drought stresses have serious effects on plant growth. LATERAL ORGAN BOUNDARY DOMAIN (LBD) proteins are a plant-specific transcription factors (TFs) family and play important roles in plants in resisting to abiotic stress. However, about the function of LBDs in apple and other woody plants is little known. In this study, protein sequences of the LBD family TFs in apples were identified which contained conserved LOB domains. The qRT-PCR analysis showed that the MdLBD3 gene was widely expressed in various tissues and organs. The subcellular localization assay showed that the MdLBD3 protein was localized in the nucleus. Ectopic expression of MdLBD3 in Arabidopsis positively regulated its salt and drought resistance, and promoted early flowering. Collectively, these results showed that MdLBD3 improved the abiotic stress resistance, plant growth and development. Overall, this study provided a new gene for breeding that can increase the abiotic stress tolerance in apple.

Generation of streptocarbazoles with cytotoxicities by pathway engineering and insights into their biosynthesis.

Streptocarbazoles are a class of indolocarbazole (ICZ) compounds produced by Streptomyces strains that feature unique cyclic N-glycosidic linkages between the 1,3-carbon atoms of the glycosyl moiety and the two indole nitrogen atoms. Although several streptocarbazole compounds display effective cytotoxic activity, their biosynthesis remains unclear. Herein, through the inactivation of the aminotransferase gene spcI in the staurosporine biosynthetic gene cluster spc followed by heterologous expression, two new streptocarbazole derivatives (1 and 3) and three known ICZs (2, 4, and 5) were generated. Their structures were determined by a combination of spectroscopic methods, circular dichroism measurements, and single-crystal X-ray diffraction. Compounds 1-4 displayed moderate cytotoxicity against HCT-116 cell line, and compounds 3 and 4 were effective against Huh 7 cell line. Double-gene knockout experiments allowed us to propose a biosynthetic pathway for streptocarbazole productions. Furthermore, by overexpression of the involving key enzymes, the production of streptocarbazoles 1 and 3 were improved by approximately 1.5-2.5 fold. IMPORTANCE: Indolocarbazoles (ICZs) are a group of antitumor agents, with several analogs used in clinical trials. Therefore, the identification of novel ICZ compounds is important for drug discovery. Streptocarbazoles harbor unique N-glycosidic linkages (N13-C1' and N12-C3'), distinguishing them from the representative ICZ compound staurosporine; however, their biosynthesis remains unclear. In this study, two new streptocarbazoles (1 and 3) with cytotoxic activities were obtained by manipulating the staurosporine biosynthetic gene cluster spc followed by heterologous expression. The biosynthetic pathway of streptocarbazoles was proposed, and their productions were improved through the overexpression of the key enzymes involved. This study enriches the structural diversity of ICZ compounds and would facilitate the discovery of new streptocarbazoles via synthetic biological strategies.

A viroid-derived small interfering RNA targets bHLH transcription factor MdPIF1 to regulate anthocyanin biosynthesis in Malus domestica.

Fruit colour is a critical determinant for the appearance quality and commercial value of apple fruits. Viroid-induced dapple symptom severely affects the fruit coloration, however, the underlying mechanism remains unknown. In this study, we identified an apple dimple fruit viroid (ADFVd)-derived small interfering RNA, named vsiR693, which targeted the mRNA coding for a bHLH transcription factor MdPIF1 (PHYTOCHROME-INTERACTING FACTOR 1) to regulate anthocyanin biosynthesis in apple. 5' RLM-RACE and artificial microRNA transient expression system proved that vsiR693 directly targeted the mRNA of MdPIF1 for cleavage. MdPIF1 positively regulated anthocyanin biosynthesis in both apple calli and fruits, and it directly bound to G-box element in the promoter of MdPAL and MdF3H, two anthocyanin biosynthetic genes, to promote their transcription. Expression of vsiR693 negatively regulated anthocyanin biosynthesis in both apple calli and fruits. Furthermore, co-expression of vsiR693 and MdPIF1 suppressed MdPIF1-promoted anthocyanin biosynthesis in apple fruits. Infiltration of ADFVd infectious clone suppressed coloration surrounding the injection sites in apple fruits, while a mutated version of ADFVd, in which the vsiR693 producing region was mutated, failed to repress fruit coloration around the injection sites. These data provide evidence that a viroid-derived small interfering RNA targets host transcription factor to regulate anthocyanin biosynthesis in apple.

The AP2/ERF transcription factor MdDREB2A regulates nitrogen utilisation and sucrose transport under drought stress.

Drought stress is one of the main environmental factors limiting plant growth and development. Plants adapt to changing soil moisture by modifying root architecture, inducing stomatal closure, and inhibiting shoot growth. The AP2/ERF transcription factor DREB2A plays a key role in maintaining plant growth in response to drought stress, but the molecular mechanism underlying this process remains to be elucidated. Here, it was found that overexpression of MdDREB2A positively regulated nitrogen utilisation by interacting with DRE cis-elements of the MdNIR1 promoter. Meanwhile, MdDREB2A could also directly bind to the promoter of MdSWEET12, which may enhance root development and nitrogen assimilation, ultimately promoting plant growth. Overall, this regulatory mechanism provides an idea for plants in coordinating with drought tolerance and nitrogen assimilation to maintain optimal plant growth and development under drought stress.