Overexpression of MsDREB1C Modulates Growth and Improves Forage Quality in Tetraploid Alfalfa (Medicago sativa L.).

DREB has been reported to be involved in plant growth and response to environmental factors. However, the function of DREB in growth and development has not been elucidated in alfalfa (Medicago sativa L.), a perennial tetraploid forage cultivated worldwide. In this study, an ortholog of MtDREB1C was characterized from alfalfa and named MsDREB1C accordingly. MsDREB1C was significantly induced by abiotic stress. The transcription factor MsDREB1C resided in the nucleus and had self-transactivation activity. The MsDREB1C overexpression (OE) alfalfa displayed growth retardation under both long-day and short-day conditions, which was supported by decreased MsGA20ox and upregulated MsGA2ox in the OE lines. Consistently, a decrease in active gibberellin (GA) was detected, suggesting a negative effect of MsDREB1C on GA accumulation in alfalfa. Interestingly, the forage quality of the OE lines was better than that of WT lines, with higher crude protein and lower lignin content, which was supported by an increase in the leaf-stem ratio (LSR) and repression of several lignin-synthesis genes (MsNST, MsPAL1, MsC4H, and Ms4CL). Therefore, this study revealed the effects of MsDREB1C overexpression on growth and forage quality via modifying GA accumulation and lignin synthesis, respectively. Our findings provide a valuable candidate for improving the critical agronomic traits of alfalfa, such as overwintering and feeding value of the forage.

The persistence and antitumor efficacy of CAR-T cells are modulated by tonic signaling within the CDR.

Chimeric antigen receptor (CAR) T cell therapy has demonstrated remarkable clinical efficacy, but challenges related to relapse and CAR-T cell exhaustion persist. One contributing factor to this exhaustion is CAR tonic signaling, where CAR-T cells self-activate without antigen stimulation, leading to reduced persistence and impaired antitumor activity. To address this issue, we conducted a preclinical study evaluating tonic signaling using nanobody-derived CAR-T cells. Our investigation revealed that specific characteristics of the complementary determining regions (CDRs), including low solubility, polarity, positive charge, energy, and area of ionic and positive CDR patches of amino acids, were associated with low antigen-independent tonic signaling. Significantly, we observed that stronger tonic signaling directly impacted CAR-T cell proliferation in vitro, consequently leading to CAR-T cell exhaustion and diminished persistence and effectiveness in vivo. Our findings provide compelling preclinical evidence and lay the foundation for the clinical assessment of CAR-T cells with distinct tonic signaling patterns. Understanding the role of CDRs in modulating tonic signaling holds promise for advancing the development of more efficient and durable CAR-T cell therapies, thereby enhancing the treatment of cancer and addressing the challenges of relapse in CAR-T cell therapy.

NMDAR antagonists suppress tumor progression by regulating tumor-associated macrophages.

Neurotransmitter receptors are increasingly recognized to play important roles in anti-tumor immunity. The expression of the ion channel N-methyl-D-aspartate receptor (NMDAR) on macrophages was reported, but the role of NMDAR on macrophages in the tumor microenvironment (TME) remains unknown. Here, we show that the activation of NMDAR triggered calcium influx and reactive oxygen species production, which fueled immunosuppressive activities in tumor-associated macrophages (TAMs) in the hepatocellular sarcoma and fibrosarcoma tumor settings. NMDAR antagonists, MK-801, memantine, and magnesium, effectively suppressed these processes in TAMs. Single-cell RNA sequencing analysis revealed that blocking NMDAR functionally and metabolically altered TAM phenotypes, such that they could better promote T cell- and Natural killer (NK) cell-mediated anti-tumor immunity. Treatment with NMDAR antagonists in combination with anti-PD-1 antibody led to the elimination of the majority of established preclinical liver tumors. Thus, our study uncovered an unknown role for NMDAR in regulating macrophages in the TME of hepatocellular sarcoma and provided a rationale for targeting NMDAR for tumor immunotherapy.

Improving the Initial Coulombic Efficiency of Sodium-Storage Antimony Anodes via Electrochemically Alloying Bismuth.

Improving cycling stability while maintaining a high initial Coulombic efficiency (ICE) of the antimony (Sb) anode is always a trade-off for the design of electrodes of sodium-ion batteries (SIBs). Herein, we prepare a carbon-free Sb8Bi1 anode with an ICE of 87.1% at 0.1 A g-1 by a one-step electrochemical reduction of Sb2O3 and Bi2O3 in alkaline solutions. The improved ICE of the Sb8Bi1 anode is due to the alloying of bismuth (Bi) that prevents irreversible interfacial reactions during the sodiation process. Unlike carbon buffers, the use of Bi will reduce the number of side reactions between the carbon buffer and sodium. Moreover, Bi2O3 can promote the reduction of Sb2O3 and reduce the particle size of Sb from ∼20 µm to below 300 nm. The electrolytic products can be modulated by controlling the cell voltages and electrolysis time. The electrolytic Sb8Bi1 anode delivered a capacity of 625 mAh g-1 after 200 cycles with an ICE of 87.1% at 0.1 A g-1 and even 625 mAh g-1 at 1 A g-1 over 100 cycles. Hence, alloying Bi into Sb is an effective way to make a long-lasting Sb anode while maintaining a high Coulombic efficiency.

Characterization of the Heat Shock Transcription Factor Family in Medicago sativa L. and Its Potential Roles in Response to Abiotic Stresses.

Heat shock transcription factors (HSFs) are important regulatory factors in plant stress responses to various biotic and abiotic stresses and play important roles in growth and development. The HSF gene family has been systematically identified and analyzed in many plants but it is not in the tetraploid alfalfa genome. We detected 104 HSF genes (MsHSFs) in the tetraploid alfalfa genome ("Xinjiangdaye" reference genome) and classified them into three subgroups: 68 in HSFA, 35 in HSFB and 1 in HSFC subgroups. Basic bioinformatics analysis, including genome location, protein sequence length, protein molecular weight and conserved motif identification, was conducted. Gene expression analysis revealed tissue-specific expression for 13 MsHSFs and tissue-wide expression for 28 MsHSFs. Based on transcriptomic data analysis, 21, 11 and 27 MsHSFs responded to drought stress, cold stress and salt stress, respectively, with seven responding to all three. According to RT-PCR, MsHSF27/33 expression gradually increased with cold, salt and drought stress condition duration; MsHSF6 expression increased over time under salt and drought stress conditions but decreased under cold stress. Our results provide key information for further functional analysis of MsHSFs and for genetic improvement of stress resistance in alfalfa.

Review immune response of targeting CD39 in cancer.

The ATP-adenosine pathway has emerged as a promising target for cancer therapy, but challenges remain in achieving effective tumor control. Early research focused on blocking the adenosine generating enzyme CD73 and the adenosine receptors A2AR or A2BR in cancer. However, recent studies have shown that targeting CD39, the rate-limiting ecto-enzyme of the ATP-adenosine pathway, can provide more profound anti-tumor efficacy by reducing immune-suppressive adenosine accumulation and increasing pro-inflammatory ATP levels. In addition, combining CD39 blocking antibody with PD-1 immune checkpoint therapy may have synergistic anti-tumor effects and improve patient survival. This review will discuss the immune components that respond to CD39 targeting in the tumor microenvironment. Targeting CD39 in cancer has been shown to not only decrease adenosine levels in the tumor microenvironment (TME), but also increase ATP levels. Additionally, targeting CD39 can limit the function of Treg cells, which are known to express high levels of CD39. With phase I clinical trials of CD39 targeting currently underway, further understanding and rational design of this approach for cancer therapy are expected.

Transcriptome and GWAS Analyses Reveal Candidate Gene for Root Traits of Alfalfa during Germination under Salt Stress.

Alfalfa growth and production in China are negatively impacted by high salt concentrations in soils, especially in regions with limited water supplies. Few reliable genetic markers are currently available for salt tolerance selection. As a result, molecular breeding strategies targeting alfalfa are hindered. Therefore, with the continuous increase in soil salinity in agricultural lands, it is indispensable that a salt-tolerant variety of alfalfa is produced. We collected 220 alfalfa varieties around the world for resequencing and performed genome-wide association studies (GWASs). Alfalfa seeds were germinated in saline water with different concentrations of NaCl, and the phenotypic differences in several key root traits were recorded. In the phenotypic analysis, the breeding status and geographical origin strongly affected the salt tolerance of alfalfa. Forty-nine markers were significantly associated with salt tolerance, and 103 candidate genes were identified based on linkage disequilibrium. A total of 2712 differentially expressed genes were upregulated and 3570 were downregulated based on transcriptomic analyses. Some candidate genes that affected root development in the seed germination stage were identified through the combination of GWASs and transcriptome analyses. These genes could be used for molecular breeding strategies to increase alfalfa's salt tolerance and for further research on salt tolerance in general.

Genome-Wide Identification and Phylogenetic and Expression Analyses of the PLATZ Gene Family in Medicago sativa L.

The PLATZ family is a novel class of plant-specific zinc finger transcription factors with important roles in plant growth and development and abiotic stress responses. PLATZ members have been identified in many plants, including Oryza sativa, Zea mays, Triticum aestivum, Fagopyrum tataricum, and Arabidopsis thaliana; however, due to the complexity of the alfalfa reference genome, the members of the PLATZ gene family in alfalfa (Medicago sativa L.) have not been systematically identified and analyzed. In this study, 55 Medicago sativa PLATZ genes (MsPLATZs) were identified in the alfalfa "Xinjiangdaye" reference genome. Basic bioinformatic analysis was performed, including the characterization of sequence lengths, protein molecular weights, genomic positions, and conserved motifs. Expression analysis reveals that 7 MsPLATZs are tissue-specifically expressed, and 10 MsPLATZs are expressed in all examined tissues. The transcriptomic expression of these genes is obvious, indicating that these MsPLATZs have different functions in the growth and development of alfalfa. Based on transcriptome data analysis and real-time quantitative PCR (RT-qPCR), we identified 22, 22, and 21 MsPLATZ genes that responded to salt, cold, and drought stress, respectively, with 20 MsPLATZs responding to all three stresses. This study lays a foundation for further exploring the functions of MsPLATZs, and provides ideas for the improvement of alfalfa varieties and germplasm innovation.

Optimization of PROTAC Ternary Complex Using DNA Encoded Library Approach.

The proteolysis targeting chimera (PROTAC) strategy results in the down-regulation of unwanted protein(s) for disease treatment. In the PROTAC process, a heterobifunctional degrader forms a ternary complex with a target protein of interest (POI) and an E3 ligase, which results in ubiquitination and proteasomal degradation of the POI. While ternary complex formation is a key attribute of PROTAC degraders, modification of the PROTAC molecule to optimize ternary complex formation and protein degradation can be a labor-intensive and tedious process. In this study, we take advantage of DNA-encoded library (DEL) technology to efficiently synthesize a vast number of possible PROTAC molecules and describe a parallel screening approach that utilizes DNA barcodes as reporters of ternary complex formation and cooperative binding. We use a designed PROTAC DEL against BRD4 and CRBN to describe a dual protein affinity selection method and the direct discovery of novel, potent BRD4 PROTACs that importantly demonstrate clear SAR. Such an approach evaluates all the potential PROTACs simultaneously, avoids the interference of PROTAC solubility and permeability, and uses POI and E3 ligase proteins in an efficient manner.

Heavy Metal Ions Trigger a Fluorescent Quenching in DNA-Organic Semiconductor Hybrid Assemblies.

The significance of DNA is no longer limited to its role as a biological information carrier; as a natural polymer, it also become in the field of materials. Single-stranded DNA (ssDNA) molecules with specific sequences can form a G-quadruplex or hairpin-shaped conformation with specific heavy metal ions through coordination bonds. In this study, ssDNA molecules of the four sequences were prepared into hybrid assemblies with one of the famous display materials, the tris-(8-hydroxyquinoline)aluminum (Alq3) semiconductor. Based on these hybrid assemblies, heavy metal ions, namely Pb2+, Hg2+, Cd2+ and As3+, were detected individually at the ppb level. Apart from this, in practical application, many samples containing heavy metal ions are digested with acid. By introducing MES buffer solution, the influence of acidity on the fluorescent signal of Alq3 was excluded. This strategy showed promising results in the practical application of detecting heavy metal ions in shrub branches and leaves.

CD155 in tumor progression and targeted therapy.

CD155, also known as the poliovirus receptor (PVR), has received considerable attention in recent years because of its intrinsic and extrinsic roles in tumor progression. Although barely expressed in host cells, CD155 is upregulated in tumor-infiltrating myeloid cells. High expression of CD155 in tumor cells across multiple cancer types is common and associated with poor patient outcomes. The intrinsic functions of CD155 in tumor cells promote tumor progression and metastasis, whereas its extrinsic immunoregulatory functions in the tumor microenvironment (TME) involve interaction with the upregulated inhibitory immune cell receptor and checkpoint TIGIT, suggesting that CD155 and CD155 pathways are promising tumor immunotherapy targets. Preclinical studies demonstrate that targeting CD155 and its receptor (anti-TIGIT) using a single treatment or in combination with anti-PD-1 can improve immune-mediated tumor control. However, there is still a limited understanding of CD155 and its associated targeting strategies, especially antibody and immune cell editing-related strategies of CD155 in cancer. Here, we review the role of CD155 in host and tumor cells in controlling tumor progression and discuss the potential of targeting CD155 for tumor therapy.

Clinically approved combination immunotherapy: Current status, limitations, and future perspective.

Immune-checkpoint inhibitor-based combination immunotherapy has become a first-line treatment for several major types of cancer including hepatocellular carcinoma (HCC), renal cell carcinoma, lung cancer, cervical cancer, and gastric cancer. Combination immunotherapy counters several immunosuppressive elements in the tumor microenvironment and activates multiple steps of the cancer-immunity cycle. The anti-PD-L1 antibody, atezolizumab, plus the anti-vascular endothelial growth factor antibody, bevacizumab, represents a promising class of combination immunotherapy. This combination has produced unprecedented clinical efficacy in unresectable HCC and become a landmark in HCC therapy. Advanced HCC patients treated with atezolizumab plus bevacizumab demonstrated impressive improvements in multiple clinical endpoints including overall survival, progress-free survival, objective response rate, and patient-reported quality of life when compared to current first-line treatment with sorafenib. However, atezolizumab plus bevacizumab first-line therapy has limitations. First, cancer patients falling into the criteria for the combination therapy may need to be further selected to reap benefits while avoiding some potential pitfalls. Second, the treatment regimen of atezolizumab plus bevacizumab at a fixed dose may require adjustment for optimal normalization of the tumor microenvironment to obtain maximum efficacy and reduce adverse events. Third, utilization of predictive biomarkers is urgently needed to guide the entire treatment process. Here we review the current status of clinically approved combination immunotherapies and the underlying immune mechanisms. We further provide a perspective analysis of the limitations for combination immunotherapies and potential approaches to overcome the limitations.

The association of IGF1 rs35767 polymorphism with colorectal cancer risk in the Chinese Han population.

Insulin-like growth factor 1 (IGF1) plays an important role in the development and growth of colorectal cancer (CRC). Hence, potential functional polymorphisms of the IGF1 gene may be involved in CRC risk. This study mainly aimed to assess the association of IGF1 rs35767 polymorphism with CRC risk in the Chinese Han population by a case-control study and a pooled analysis. In a case-control study with 208 CRC patients and 312 healthy individuals, the rs35767 polymorphism was genotyped by DNA sequencing. Furthermore, a pooled analysis of two case-control studies was performed using Stata software. IGF1 rs35767 polymorphism was significantly associated with CRC risk in both a case-control study (AA vs. GG: OR = 2.26, 95% CI = 1.35-3.80, P = 0.003; AA vs. (GG + GA): OR = 2.32, 95% CI = 1.44-3.74, P = 0.001; A vs. G: OR = 1.43, 95% CI = 1.11-1.85, P = 0.007) and a pooled analysis [(GA + AA) versus GG: OR = 1.30, 95% CI = 1.03-1.63, P = 0.03; A versus G: OR = 1.28, 95% CI = 1.08-1.53, P = 0.01]. In addition, the IGF1 rs35767 polymorphism was also significantly associated with the stage of CRC. CRC patients with the rs35767 A allele were more likely to have a high tumor stage. These findings indicated that IGF1 rs35767 polymorphism was linked to CRC risk and tumor stage in the Chinese Han population, and might serve as a valuable biomarker.

NKG7 Is Required for Optimal Antitumor T-cell Immunity.

Tumor antigen-specific CD8+ T cells play a critical role in antitumor immunity. Clinical trials reinvigorating the immune system via immune checkpoint blockade (ICB) have shown remarkable clinical promise. Numerous studies have identified an association between NKG7 expression and patient outcome across different malignancies. However, aside from these correlative observations, very little is known about NKG7 and its role in antitumor immunity. Herein, we utilized single-cell RNA sequencing (scRNA-seq) datasets, NKG7-deficient mice, NKG7-reporter mice, and mouse tumor models to investigate the role of NKG7 in neoantigen-mediated tumor rejection and ICB immunotherapy. scRNA-seq of tumors from patients with metastatic melanoma or head and neck squamous cell carcinoma revealed that NKG7 expression is highly associated with cytotoxicity and specifically expressed by CD8+ T cells and natural killer (NK) cells. Furthermore, we identified a key role for NKG7 in controlling intratumor T-cell accumulation and activation. NKG7 was upregulated on intratumor antigen-specific CD8+ T cells and NK cells and required for the accumulation of T cells in the tumor microenvironment. Accordingly, neoantigen-expressing mouse tumors grew faster in Nkg7-deficient mice. Strikingly, efficacy of single or combination ICB was significantly reduced in Nkg7-deficient mice.See related article by Wen et al., p. 162.

The callus formation capacity of strawberry leaf explant is modulated by DNA methylation.

Shoot regeneration from leaf tissue requires de-differentiation of cells from a highly differentiated state into an active dividing state, but how this physiological transition occurs and is regulated especially at epigenetic level remains obscure. Here we have characterized the DNA methylome represented by 5-methylcytosine (5mC) in leaf and the callus tissue derived from the leaf explant of woodland strawberry Fragaria vesca. We detected an overall increase of DNA methylation and distinct 5mC enrichment patterns in the CG, CHG and CHH sequence contexts in genetic and transposable elements. Our analyses revealed an intricate relation between DNA methylation and gene expression levels in leaf or leaf-derived callus. However, when considering the genes involved in callus formation and shoot regeneration, e.g. FvePLT3/7, FveWIND3, FveWIND4, FveLOG4 and FveIAA14, their dynamic transcription levels were associated with the differentially methylated regions located in the promoters or gene bodies, indicating a regulatory role of DNA methylation in the transcriptional regulation of pluripotency acquisition in strawberry. Furthermore, application of a DNA methyltransferase inhibitor 5'-azacytidine (5'-Aza) hampered both callus formation and shoot regeneration from the leaf explant. We further showed that 5'-Aza down-regulated the genes involved in cell wall integrity, such as expansin, pectin lyase and pectin methylesterase genes, suggesting an essential role of cell wall metabolism during callus formation. This study reveals the contribution of DNA methylation in callus formation capacity and will provide a basis for developing a strategy to improve shoot regeneration for basic and applied research applications.

Immunotherapeutic approaches in EBV-associated nasopharyngeal carcinoma.

Epstein-Barr virus (EBV) was the first tumor virus in humans. Nasopharyngeal carcinoma (NPC) accounts for approximately 60% of the 200,000 new tumor cases caused by EBV infection worldwide each year. NPC has an insidious onset and is highly malignant, with more than 70% of patients having intermediate to advanced disease at the time of initial diagnosis, and is strongly implicated in epithelial cancers as well as malignant lymphoid and natural killer/T cell lymphomas. Over 90% of patients with confirmed undifferentiated NPC are infected with EBV. In recent decades, much progress has been made in understanding the molecular mechanisms of NPC and developing therapeutic approaches. Radiotherapy and chemotherapy are the main treatment options for NPC; however, they have a limited efficacy in patients with locally advanced or distant metastatic tumors. Tumor immunotherapy, including vaccination, adoptive cell therapy, and immune checkpoint blockade, represents a promising therapeutic approach for NPC. Significant breakthroughs have recently been made in the application of immunotherapy for patients with recurrent or metastatic NPC (RM-NPC), indicating a broad prospect for NPC immunotherapy. Here, we review important research findings regarding immunotherapy for NPC patients and provide insights for future research.

The Association of MEG3 Gene rs7158663 Polymorphism With Cancer Susceptibility.

Although the association of MEG3 gene rs7158663 polymorphism with cancer susceptibility has been investigated, the findings are inconsistent. The aim of this study was to analyze the association between the rs7158663 polymorphism and cancer susceptibility through a case-control study and meta-analysis. In a case-control study with 430 colorectal cancer (CRC) cases and 445 healthy controls, the rs7158663 polymorphism was genotyped by direct sequencing. STATA software was used to calculate the pooled odds ratio and 95% confidence interval in a meta-analysis including 4,649 cancer cases and 5,590 controls. Both the case-control study and meta-analysis showed that the rs7158663 polymorphism was associated with increased susceptibility to CRC. Individuals carrying the AA or GA genotype were more likely to develop CRC than those carrying the rs7158663 GG genotype. Interestingly, MEG3 expression was significantly lower in colorectal tissues of the AA or GA genotype compared to those of the rs7158663 GG genotype. In addition, the meta-analysis suggested that the rs7158663 polymorphism was also associated with increased susceptibility to breast cancer and gastric cancer. Bioinformatics analysis showed that the rs7158663 A allele contributed to the binding of hsa-miR-4307 and hsa-miR-1265 to MEG3. In conclusion, the current findings suggest that the MEG3 gene rs7158663 polymorphism may serve as a genetic marker for predicting the risk of cancers, such as breast cancer, gastric cancer and CRC. However, the sample size of the current study is still insufficient, especially in the subgroup analysis. Therefore large and well-designed studies are needed to validate our findings.

Characterization of expansin genes and their transcriptional regulation by histone modifications in strawberry.

MAIN CONCLUSION: The possible candidate expansin genes, which may be important for strawberry fruit softening, have been identified in the diploid woodland strawberry Fragaria vesca and the octoploid cultivated strawberry Fragaria × ananassa and their transcriptional regulation by histone modifications has been studied. Softening process greatly affects fruit texture and shelf life. Expansins (EXPs) are a group of structural proteins participating in cell wall loosening, which break the hydrogen bonding between cellulose microfibrils and hemicelluloses. However, our knowledge on how EXP genes are regulated in fruit ripening, especially in non-climacteric fleshy fruits, is limited. Here, we have identified the EXP genes in both the octoploid cultivated strawberry (Fragaria × ananassa) and one of its diploid progenitor species, woodland strawberry (Fragaria vesca). We found that EXP proteins in F. × ananassa were structurally more divergent than the ones in F. vesca. Transcriptome data suggested that FaEXP88, FaEXP114, FveEXP11 and FveEXP33 were the four candidate EXP genes more likely involved in fruit softening, whose transcript levels dramatically increased when firmness decreased during fruit maturation. Phylogenetic analyses showed that those candidate genes were closely clustered, indicating the presence of homoeolog expression dominance in the EXP gene family in strawberry. Moreover, we have performed chromatin immunoprecipitation (ChIP) experiments to investigate the distribution of histone modifications along the promoters and genic regions of the EXP genes in F. vesca. ChIP data revealed that the transcript levels of EXP genes were highly correlated with the enrichment of H3K9/K14 acetylation and H3K27 tri-methylation. Collectively, this study identifies the key EXP genes involved in strawberry fruit softening and reveals a regulatory role of histone modifications in their transcriptional regulation, which would facilitate functional studies of the EXP genes in the ripening of non-climacteric fruits.