The molecular regulatory mechanism of reed canary grass under salt, waterlogging, and combined stress was analyzed by transcriptomic analysis.

BACKGROUND: Reed canary grass has been identified as a suitable species for restoring plateau wetlands and understanding plant adaptation mechanisms in wetland environments. In this study, we subjected a reed canary grass cultivar 'Chuanxi' to waterlogging, salt, and combined stresses to investigate its phenotypic characteristics, physiological indices, and transcriptome changes under these conditions. RESULTS: The results revealed that the growth rate was slower under salt stress than under waterlogging stress. The chlorophyll content and energy capture efficiency of the PS II reaction center decreased with prolonged exposure to each stress. Conversely, while the activities of enzymes associated with respiratory metabolism, as well as MDA, PRO, Na+, and K+-ATPase, increased. The formation of distinct aerenchyma was observed under waterlogging stress and combined stress. Transcriptome sequencing analysis identified 5,379, 4,169, and 14,993 DEGs under CK vs. W, CK vs. S, and CK vs. SW conditions, respectively. The WRKY was found to be the most abundant under waterlogging stress, whereas the MYB predominated under salt stress and combined stress. Glutathione metabolic pathways and Plant hormone signal transduction have also been found to play important roles in stress. CONCLUSION: By integrating phenotypic, physiological, anatomical, and transcriptomic, this research provides valuable insights into how reed canary grass responds to salt, waterlogging, and combined stresses. These findings may inform the ecological application of reed canary grass in high-altitude wetlands and for breeding purposes.

Exploring Evolutionary Pathways and Abiotic Stress Responses through Genome-Wide Identification and Analysis of the Alternative Oxidase (AOX) Gene Family in Common Oat (Avena sativa).

The alternative oxidase (AOX), a common terminal oxidase in the electron transfer chain (ETC) of plants, plays a crucial role in stress resilience and plant growth and development. Oat (Avena sativa), an important crop with high nutritional value, has not been comprehensively studied regarding the AsAOX gene family. Therefore, this study explored the responses and potential functions of the AsAOX gene family to various abiotic stresses and their potential evolutionary pathways. Additionally, we conducted a genome-wide analysis to explore the evolutionary conservation and divergence of AOX gene families among three Avena species (Avena sativa, Avena insularis, Avena longiglumis) and four Poaceae species (Avena sativa, Oryza sativa, Triticum aestivum, and Brachypodium distachyon). We identified 12 AsAOX, 9 AiAOX, and 4 AlAOX gene family members. Phylogenetic, motif, domain, gene structure, and selective pressure analyses revealed that most AsAOXs, AiAOXs, and AlAOXs are evolutionarily conserved. We also identified 16 AsAOX segmental duplication pairs, suggesting that segmental duplication may have contributed to the expansion of the AsAOX gene family, potentially preserving these genes through subfunctionalization. Chromosome polyploidization, gene structural variations, and gene fragment recombination likely contributed to the evolution and expansion of the AsAOX gene family as well. Additionally, we hypothesize that AsAOX2 may have potential function in resisting wounding and heat stresses, while AsAOX4 could be specifically involved in mitigating wounding stress. AsAOX11 might contribute to resistance against chromium and waterlogging stresses. AsAOX8 may have potential fuction in mitigating ABA-mediated stress. AsAOX12 and AsAOX5 are most likely to have potential function in mitigating salt and drought stresses, respectively. This study elucidates the potential evolutionary pathways of the AsAOXs gene family, explores their responses and potential functions to various abiotic stresses, identifies potential candidate genes for future functional studies, and facilitates molecular breeding applications in A. sativa.

Non-Volatile Component and Antioxidant Activity: A Comparative Analysis between Litsea cubeba Branches and Leaves.

Litsea cubeba, which is found widely distributed across the Asian region, functions as both an economic tree and a medicinal plant with a rich historical background. Previous investigations into its chemical composition and biological activity have predominantly centered on volatile components, leaving the study of non-volatile components relatively unexplored. In this study, we employed UPLC-HRMS technology to analyze the non-volatile components of L. cubeba branches and leaves, which successfully resulted in identifying 72 constituents. Comparative analysis between branches and leaves unveiled alkaloids, organic acids, and flavonoids as the major components. However, noteworthy differences in the distribution of these components between branches and leaves were observed, with only eight shared constituents, indicating substantial chemical variations in different parts of L. cubeba. Particularly, 24 compounds were identified for the first time from this plant. The assessment of antioxidant activity using four methods (ABTS, DPPH, FRAP, and CUPRAC) demonstrated remarkable antioxidant capabilities in both branches and leaves, with slightly higher efficacy observed in branches. This suggests that L. cubeba may act as a potential natural antioxidant with applications in health and therapeutic interventions. In conclusion, the chemical composition and antioxidant activity of L. cubeba provides a scientific foundation for its development and utilization in medicine and health products, offering promising avenues for the rational exploitation of L. cubeba resources in the future.

A subset of VEGFR-TKIs activates AMPK in LKB1-mutant lung cancer.

The mutation of tumor suppressor gene liver kinase B1 (LKB1) has a prevalence of about 20% in non-small cell lung cancer (NSCLC). LKB1-mutant lung cancer is characterized by enhanced aggressiveness and immune escape and is associated with poor prognosis. Therefore, it is urgent to develop effective therapeutic methods for LKB1-mutant NSCLC. Recently, apatinib, a VEGFR-TKI, was found to significantly improve the outcome of LKB1-mutant NSCLC, but the mechanism is not completely clear. In this study, AMP-activated protein kinase (AMPK), the crucial downstream kinase of LKB1 was excavated as the potential target of apatinib. Biochemical experiments verified that apatinib is a direct AMPK activator. Moreover, clinically available VEGFR-TKIs were found to regulate AMPK differently: Apatinib and anlotinib can directly activate AMPK, while axitinib and sunitinib can directly inhibit AMPK. Activation of AMPK by apatinib leads to the phosphorylation of acetyl-CoA carboxylase (ACC) and inhibition of de novo fatty acid synthesis (FAsyn), which is upregulated in LKB1-null cancers. Moreover, the killing effect of apatinib was obviously enhanced under delipidated condition, and the combination of exogenous FA restriction with apatinib treatment can be a promising method for treating LKB1-mutant NSCLC. This study discovered AMPK as an important off-target of apatinib and elucidated different effects of this cluster of VEGFR-TKIs on AMPK. This finding can be the basis for the accurate and combined application of these drugs in clinic and highlights that the subset of VEGFR-TKIs including apatinib and anlotinib are potentially valuable in the treatment of LKB1-mutant NSCLC.

RAD-seq as an effective strategy for heterogenous variety identification in plants-a case study in Italian Ryegrass (Lolium multiflorum).

The primary approach for variety distinction in Italian ryegrass is currently the DUS (distinctness, uniformity and stability) test based on phenotypic traits. Considering the diverse genetic background within the population and the complexity of the environment, however, it is challenging to accurately distinguish varieties based on DUS criteria alone. In this study, we proposed the application of high-throughput RAD-seq to distinguish 11 Italian ryegrass varieties with three bulks of 50 individuals per variety. Our findings revealed significant differences among the 11 tested varieties. The PCA, DAPC and STRUCTURE analysis indicated a heterogeneous genetic background for all of them, and the AMOVA analysis also showed large genetic variance among these varieties (ΦST = 0.373), which were clearly distinguished based on phylogenetic analysis. Further nucleotide diversity (Pi) analysis showed that the variety 'Changjiang No.2' had the best intra-variety consistency among 11 tested varieties. Our findings suggest that the RAD-seq could be an effectively alternative method for the variety distinction of Italian ryegrass, as well as a potential tool for open-pollinated varieties (OPVs) of other allogamous species.

Comparative Metabolomic Studies of Siberian Wildrye (Elymus sibiricus L.): A New Look at the Mechanism of Plant Drought Resistance.

Drought is one of the most important factors affecting plant growth and production due to ongoing global climate change. Elymus sibiricus has been widely applied for ecological restoration and reseeding of degraded grassland in the Qinghai-Tibetan Plateau (QTP) because of its strong adaptability to barren, salted, and drought soils. To explore the mechanism of drought resistance in E. sibiricus, drought-tolerant and drought-sensitive genotypes of E. sibiricus were used in metabolomic studies under simulated long-term and short-term drought stress. A total of 1091 metabolites were detected, among which, 27 DMs were considered to be the key metabolites for drought resistance of E. sibiricus in weighted gene co-expression network analysis (WGCNA). Ten metabolites, including 3-amino-2-methylpropanoic acid, coniferin, R-aminobutyrate, and so on, and 12 metabolites, including L-Proline, L-histidine, N-acetylglycine, and so on, showed differential accumulation patterns under short-term and long-term drought stress, respectively, and thus, could be used as biomarkers for drought-tolerant and drought-sensitive E. sibiricus. In addition, different metabolic accumulation patterns and different drought response mechanisms were also found in drought-tolerant and drought-sensitive genotypes of E. sibiricus. Finally, we constructed metabolic pathways and metabolic patterns for the two genotypes. This metabolomic study on the drought stress response of E. sibiricus can provide resources and a reference for the breeding of new drought-tolerant cultivars of E. sibiricus.

Genomic survey sequencing, development and characterization of single- and multi-locus genomic SSR markers of Elymus sibiricus L.

BACKGROUND: Siberian wildrye (Elymus sibiricus L.) attracts considerable interest for grassland establishment and pasture recovery in the Qinghai-Tibet Plateau (QTP) due to its excellence in strong stress tolerance, high nutritional value and ease to cultivate. However, the lack of genomic information of E. sibiricus hampers its genetics study and breeding process. RESULTS: In this study, we performed a genome survey and developed a set of SSR markers for E. sibiricus based on Next-generation sequencing (NGS). We generated 469.17 Gb clean sequence which is 58.64× of the 6.86 Gb estimated genome size. We assembled a draft genome of 4.34 Gb which has 73.23% repetitive elements, a heterozygosity ratio of 0.01% and GC content of 45.68%. Based on the gnomic sequences we identified 67,833 SSR loci and from which four hundred were randomly selected to develop markers. Finally, 30 markers exhibited polymorphism between accessions and ten were identified as single-locus SSR. These newly developed markers along with previously reported 30 ones were applied to analyze genetic polymorphism among 27 wild E. sibiricus accessions. We found that single-locus SSRs are superior to multi-loci SSRs in effectiveness. CONCLUSIONS: This study provided insights into further whole genome sequencing of E. sibiricus in strategy selection. The novel developed SSR markers will facilitate genetics study and breeding for Elymus species.

A randomized phase 2 trial of apatinib vs observation as maintenance treatment following first-line induction chemotherapy in extensive- stage small cell lung cancer.

Background The 5-year survival rate for extensive-disease small-cell lung carcinoma (ED-SCLC) is only 1%. Recently, apatinib exerted promising effects on cancer patients after failure of first-line chemotherapy. Methods This study enrolled 24 ED-SCLC patients to study the efficacy and toxicity of apatinib in combination with chemotherapy and maintenance therapy. The primary endpoints were overall survival (OS) and progression-free survival (PFS). The secondary endpoints included toxicity and safety. Apatinib was given 250 mg/day during the chemotherapy interval, and as maintenance therapy after 4-6 cycles until the patient progressed, died, or was intolerant to drug toxicity. The study further evaluated the cytotoxicity, cell-cycle arrest and apoptotic induction of apatinib in A549 and H446 cells. Results There was no difference in short-term efficacy between combined and chemotherapy groups. Long-term efficacy showed that the median PFS was 7.8 months and 4.9 months in combination and chemotherapy groups, respectively [p = 0.002, HR(95%CI): 0.18(0.06-0.60)]. The median OS was 12.1 months and 8.2 months in combination and chemotherapy groups, respectively [p = 0.023, HR(95%CI): 0.38 (0.16-0.90)]. Multivariate Cox regression analysis showed that apatinib combined with chemotherapy was an independent prognostic factor for OS and PFS. The ECOG score was an independent prognostic factor affecting OS. In vitro analysis showed that apatinib inhibited cell proliferation and caused cell-cycle arrest and apoptosis. Conclusion Apatinib combination/maintenance therapy showed promising efficacy and safety to extend OS/PFS in ED-SCLC and will be a potent therapeutic option in future practice. Although the scale of this study is small, further research on large sample sizes is needed.

Nm23-H1 is involved in the repair of ionizing radiation-induced DNA double-strand breaks in the A549 lung cancer cell line.

BACKGROUND: Although originally identified as a putative metastasis suppressor, increasing studies have confirmed a possible role for Nm23-H1 in DNA repair, through the base excision repair and nucleotide excision repair pathways. In this study, we explored whether Nm23-H1 was also involved in double-strand break repair (DSBR). METHODS AND RESULTS: We constructed a stable A549-shNm23-H1 cell line with doxycycline-regulated expression of Nm23-H1, and a A549-nNm23-H1 cell line that over expressed a nucleus-localized version of Nm23-H1. Results from both lines confirmed that Nm23-H1 participated in the repair of double-strand breaks induced by X-rays, using Comet and γ-H2AX foci assays. Subsequent studies showed that Nm23-H1 activated the phosphorylation of checkpoint-related proteins including ATM serine/threonine kinase (on S1981), tumor protein p53 (on S15), and checkpoint kinase 2 (Chk2) (on T68). We also detected interactions between Nm23-H1 and the MRE11-RAD50-NBS1 (MRN) complex, as well as Ku80. Moreover, NBS1 and Ku80 levels were comparably higher in Nm23-H1 overexpressing cells than in control cells (t = 14.462, p < 0.001 and t = 5.347, p = 0.006, respectively). As Ku80 is the keystone of the non-homologous end joining (NHEJ) pathway, we speculate that Nm23-H1 promotes DSBR through NHEJ. CONCLUSIONS: The results indicate that Nm23-H1 participates in multiple steps of DSBR.

Association of DNA base excision repair genes (OGG1, APE1 and XRCC1) polymorphisms with outcome to platinum-based chemotherapy in advanced nonsmall-cell lung cancer patients.

Polymorphism of DNA base excision repair (BER) genes affects DNA repair capacity and may alter sensitivity to platinum-based chemotherapy regimens. This study investigated polymorphisms of OGG1 Ser326Cys, APE1 Asp148Glu APE1-141T/G and XRCC1 Arg399Gln for association with clinical outcome in 235 advanced inoperable nonsmall-cell lung cancer (NSCLC) patients after treatment with platinum-based chemotherapy. The multivariate analysis showed that OGG1 326 GC was associated with poor PFS [hazard ratio (HR) 1.730, p = 0.005], while XRCC1 399 GA, or GA+AA, was associated with poor OS in short-term period (HR 1.718, p = 0.003; HR 1.691, p = 0.003, respectively). Patients with OGG1 326/XRCC1 399 variant alleles had a higher risk to die early in short-term period (HR 1.929, p < 0.001). Furthermore, patients with XRCC1 399 variant allele (GA+AA) had higher risk of hematologic toxicity (p = 0.009), whereas patients carrying the OGG1 326 variant (GG), or the APE1-141 GG variant, had reduced risk of gastrointestinal toxicity (p = 0.015 and p = 0.023, respectively). The data from the current study provide evidence that OGG1 Ser326Cys, XRCC1 Arg399Gln, APE1 Asp148Glu, and APE1-141T/G polymorphisms may be useful in predicting clinical outcomes in patients with advanced inoperable NSCLC that will undergo platinum-based chemotherapy.

Hyperfractionated Accelerated Radiotherapy Versus Stereotactic Body Radiotherapy in the Treatment of Limited-Stage Small Cell Lung Cancer: A Matched-Pair Analysis.

BACKGROUND: Concurrent chemoradiotherapy based on hyperfractionated accelerated radiotherapy (HART) is the first-line recommended regimen for the treatment of small-cell lung cancer (SCLC). However, Stereotactic Body Radiotherapy (SBRT) is also regarded as an effective treatment for limited-stage (LS) SCLC, and the efficacy and safety of HART versus SBRT stay controversial. METHODS: In this study, 188 LS-SCLC patients were retrospectively divided into two groups receiving chemotherapy combined with either HART or SBRT. In HART group, patients received 4500 cGy in 30 fractions, administered twice daily for 3 weeks. Whereas in the SBRT group, a total radiation dose of 4000-4500 cGy was delivered in 10 fractions over 2 weeks. Thirty-three pairs of patients were finally included for next analysis. RESULTS: The estimated objective response rates were 63.6 % (21/33) and 78.8 % (26/33) in HART group and SBRT group, respectively (P = 0.269). Furthermore, there was no significant difference between HART and SBRT groups in overall survival (26 months vs. 29 months, P = 0.362) and progression free survival (11 months vs. 15 months, P = 0.223). As for the adverse events, toxicity of both groups is similar and slight that no grade 4 event was observed. Grade 3 pneumonitis cases were all occurred in the HART group (9.1%, 3/33, P = 0.238), and grade 3 esophagitis cases were all occurred in the SBRT group (6.1%, 2/33, P = 0.492). CONCLUSION: Compared with HART, SBRT could be another effective treatment with satisfactory safety for the concurrent chemoradiotherapy in patients with LS-SCLC.

The Ability of Clinically Relevant Chemotherapeutics to Induce Immunogenic Cell Death in Squamous Cell Carcinoma.

BACKGROUND: Immunogenic cell death (ICD) is a crucial mechanism for triggering the adaptive immune response in cancer patients. Damage-associated molecular patterns (DAMPs) are critical factors in the detection of ICD. Chemotherapeutic drugs can cause ICD and the release of DAMPs. The aim of this study was to assess the potential for paclitaxel and platinum-based chemotherapy regimens to induce ICD in squamous cell carcinoma (SCC) cell lines. In addition, we examined the immunostimulatory effects of clinically relevant chemotherapeutic regimens utilized in the treatment of SCC. METHODS: We screened for differentially expressed ICD markers in the supernatants of three SCC cell lines following treatment with various chemotherapeutic agents. The ICD markers included Adenosine Triphosphate (ATP), Calreticulin (CRT), Annexin A1 (ANXA 1), High Mobility Group Protein B1 (HMGB1), and Heat Shock Protein 70 (HSP70). A vaccination assay was also employed in C57BL/6J mice to validate our in vitro findings. Lastly, the levels of CRT and HMGB1 were evaluated in Serum samples from SCC patients. RESULTS: Addition of the chemotherapy drugs cisplatin (DDP), carboplatin (CBP), nedaplatin (NDP), oxaliplatin (OXA) and docetaxel (DOC) increased the release of ICD markers in two of the SCC cell lines. Furthermore, mice that received vaccinations with cervical cancer cells treated with DDP, CBP, NDP, OXA, or DOC remained tumor-free. Although CBP induced the release of ICD-associated molecules in vitro, it did not prevent tumor growth at the vaccination site in 40% of mice. In addition, both in vitro and in vivo results showed that paclitaxel (TAX) and LBP did not induce ICD in SCC cells. CONCLUSION: The present findings suggest that chemotherapeutic agents can induce an adjuvant effect leading to the extracellular release of DAMPs. Of the agents tested here, DDP, CBP, NDP, OXA and DOC had the ability to act as inducers of ICD.

WTAP/YTHDF1-mediated m6A modification amplifies IFN-γ-induced immunosuppressive properties of human MSCs.

INTRODUCTION: The immunosuppressive capacity of mesenchymal stem cells (MSCs) is dependent on the "license" of several pro-inflammatory factors to express immunosuppressive molecular profiles, which determines the therapeutic efficacy of MSCs in immune-mediated inflammatory diseases. Of those, interferon-γ (IFN-γ) is a key inducer for the expression of immunosuppressive molecular profiles; however, the mechanism underlying this effect is unknown. OBJECTIVES: To elucidate the regulation mechanism and biological functions of N6-methyladenosine (m6A) modification in the immunosuppressive functions by the IFN-γ-licensing MSCs. METHODS: Epitranscriptomic microarray analysis and MeRIP-qPCR assay were performed to identify the regulatory effect of WTAP in the IFN-γ-licensing MSCs. RIP-qPCR, western blot, qRT-PCR and RNA stability assays were used to determine the regulation of WTAP/m6A/YTHDF1 signaling axis in the expression of immunosuppressive molecules. Further, functional capacity of T cells was tested using flow cytometry, and both DSS-induced colitis mice and CIA mice were constructed to clarify the effect of WTAP and YTHDF1 in MSC-mediated immunosuppression. RESULTS: We identified that IFN-γ increased the m6A methylation levels of immunosuppressive molecules, while WTAP deficiency abolished the IFN-γ-induced promotion of m6A modification. IFN-γ activated ERK signaling, which induced WTAP phosphorylation. Additionally, the stabilization of WTAP post-transcriptionally increased the mRNA expression of immunosuppressive molecules (IDO1, PD-L1, ICAM1, and VCAM1) in an m6A-YTHDF1-dependent manner; this effect further impacted the immunosuppressive capacity of IFN-γ licensing MSCs on activated T cells. Notably, WTAP/YTHDF1 overexpression enhanced the therapeutic efficacy of IFN-γ licensing MSCs and restructures the ecology of inflammation in both colitis and arthritis models. CONCLUSION: Our results showed that m6A modification of IDO1, PD-L1, ICAM1, and VCAM1 mRNA mediated by WTAP-YTHDF1 is involved in the regulation of IFN-γ licensing MSCs immunosuppressive abilities, and shed a light to enhance the clinical therapeutic potential of IFN-γ-licensing MSCs.

Identification and Validation of Reference Genes for RT-qPCR Analysis in Reed Canary Grass during Abiotic Stress.

Reed canary grass (Phalaris arundinacea L.) is known for its tolerance to drought, heavy metals, and waterlogging, making it a popular choice for forage production and wetland restoration in the Qinghai-Tibet Plateau (QTP). To accurately assess gene expression in reed canary grass under different abiotic stresses, suitable reference genes need to be identified and validated. Thirteen candidate reference gene sequences were selected and screened using RT-qPCR to detect their expression levels in reed canary grass leaves under drought, salt, cadmium, and waterlogging stresses. Four algorithms were used to assess the stability of the expression levels of the candidate reference genes. The most stably expressed genes were UBC and H3 under drought Cd, ETF and CYT under salt stress, and ETF and TUB under waterlogging stress. GAPDH was found to be less stable under abiotic stresses. PIP-1, PAL, NAC 90, and WRKY 72A were selected as response genes for quantitative expression assessment under drought, salt, Cd, and waterlogging stresses to confirm the accuracy of the selected stable reference genes. These results provide a theoretical reference for assessing gene expression in reed canary grass under abiotic stresses.

Genetic diversity analysis and molecular characteristics of wild centipedegrass using sequence-related amplified polymorphism (SRAP) markers.

Centipedegrass (Eremochloa ophiuroides (Munro) Hack.) is commonly used as a low-maintenance warm-season turfgrass owing to its excellent adaptation to various soil types. A better understanding of the genetic diversity pattern of centipedegrass is essential for the efficient development and utilization of accessions. This study used fifty-five pairs of primers to detect the genetic variation and genetic structure of twenty-three wild centipedegrass accessions by Sequence-related amplified polymorphism (SRAP) markers. A total of 919 reliable bands were amplified, among which 606 (65.80%) were polymorphic and 160 (2.91%) were the monomorphic loci. The average polymorphic information content (PIC) value was 0.228. The unweighted pair group method with arithmetic mean (UPGMA) clustering analysis grouped the twenty-three accessions into two clusters. Meanwhile, the structure analysis showed that the tested accessions possessed two main genetic memberships (K = 2). The Mantel test significantly correlated the genetic and geographic distance matrices (r = 0.3854, p = 0.000140). Furthermore, geographical groups showed moderate genetic differentiation, and the highest intragroup genetic diversity was found in the Sichuan group (He = 0.201). Overall, the present research findings could promote the protection and collection of centipedegrass and provide comprehensive information to develop novel breeding strategies.

Selection of Suitable Reference Genes for RT-qPCR Gene Expression Analysis in Centipedegrass under Different Abiotic Stress.

As a C4 warm-season turfgrass, centipedegrass (Eremochloa ophiuroides (Munro) Hack.) is known for its exceptional resilience to intensive maintenance practices. In this research, the most stably expressed reference genes in the leaves of centipedegrass under different stress treatments, including salt, cold, drought, aluminum (Al), and herbicide, were screened by the quantitative real-time PCR (RT-qPCR) technique. The stability of 13 candidate reference genes was evaluated by software GeNorm V3.4, NormFinder V20, BestKeeper V1.0, and ReFinder V1.0. The results of this experiment demonstrated that the expression of the UBC (ubiquitin-conjugating enzyme) remained the most stable under cold and Al stress conditions. On the other hand, the MD (malate dehydrogenase) gene exhibited the best performance in leaf tissues subjected to salt and drought stresses. Under herbicide stress, the expression level of the RIP (60S ribosomal protein L2) gene ranked the highest. The expression levels of abiotic stress-associated genes such as PIP1, PAL, COR413, ALMT9, and BAR were assessed to validate the reliability of the selected reference genes. This study provides valuable information and reference points for gene expression under abiotic stress conditions in centipedegrass.

Analysis of Complete Chloroplast Genome: Structure, Phylogenetic Relationships of Galega orientalis and Evolutionary Inference of Galegeae.

Galega orientalis, a leguminous herb in the Fabaceae family, is an ecologically and economically important species widely cultivated for its strong stress resistance and high protein content. However, genomic information of Galega orientalis has not been reported, which limiting its evolutionary analysis. The small genome size makes chloroplast relatively easy to obtain genomic sequence for phylogenetic studies and molecular marker development. Here, the chloroplast genome of Galega orientalis was sequenced and annotated. The results showed that the chloroplast genome of G. orientalis is 125,280 bp in length with GC content of 34.11%. A total of 107 genes were identified, including 74 protein-coding genes, 29 tRNAs and four rRNAs. One inverted repeat (IR) region was lost in the chloroplast genome of G. orientalis. In addition, five genes (rpl22, ycf2, rps16, trnE-UUC and pbf1) were lost compared with the chloroplast genome of its related species G. officinalis. A total of 84 long repeats and 68 simple sequence repeats were detected, which could be used as potential markers in the genetic studies of G. orientalis and related species. We found that the Ka/Ks values of three genes petL, rpl20, and ycf4 were higher than one in the pairwise comparation of G. officinalis and other three Galegeae species (Calophaca sinica, Caragana jubata, Caragana korshinskii), which indicated those three genes were under positive selection. A comparative genomic analysis of 15 Galegeae species showed that most conserved non-coding sequence regions and two genic regions (ycf1 and clpP) were highly divergent, which could be used as DNA barcodes for rapid and accurate species identification. Phylogenetic trees constructed based on the ycf1 and clpP genes confirmed the evolutionary relationships among Galegeae species. In addition, among the 15 Galegeae species analyzed, Galega orientalis had a unique 30-bp intron in the ycf1 gene and Tibetia liangshanensis lacked two introns in the clpP gene, which is contrary to existing conclusion that only Glycyrrhiza species in the IR lacking clade (IRLC) lack two introns. In conclusion, for the first time, the complete chloroplast genome of G. orientalis was determined and annotated, which could provide insights into the unsolved evolutionary relationships within the genus Galegeae.

Pan-cancer analysis of chromothripsis-related gene expression patterns indicates an association with tumor immune and therapeutic agent responses.

Chromothripsis is a catastrophic event involving numerous chromosomal rearrangements in confined genomic regions of one or a few chromosomes, causing complex effects on cells via the extensive structural variation. The development of whole-genome sequencing (WGS) has promoted great progress in exploring the mechanism and effect of chromothripsis. However, the gene expression characteristics of tumors undergone chromothripsis have not been well characterized. In this study, we found that the transcriptional profile of five tumor types experiencing chromothripsis is associated with an immune evasion phenotype. A gene set variation analysis (GSVA) was used to develop a CHP score, which is based on differentially expressed gene sets in the TCGA database, revealing that chromothripsis status in multiple cancers is consistent with an abnormal tumor immune microenvironment and immune cell cytotoxicity. Evaluation using four immunotherapy datasets uncovered the ability of the CHP score to predict immunotherapy response in diverse tumor types. In addition, the CHP score was found to be related to resistance against a variety of anti-tumor drugs, including anti-angiogenesis inhibitors and platinum genotoxins, while EGFR pathway inhibitors were found to possibly be sensitizers for high CHP score tumors. Univariate COX regression analysis indicated that the CHP score can be prognostic for several types of tumors. Our study has defined gene expression characteristics of tumors with chromothripsis, supporting the controversial link between chromothripsis and tumor immunity. We also describe the potential value of the CHP score in predicting the efficacy of immunotherapy and other treatments, elevating chromothripsis as a tool in clinical practice.

Integrating Full-Length Transcriptome and RNA Sequencing of Siberian Wildrye (Elymus sibiricus) to Reveal Molecular Mechanisms in Response to Drought Stress.

Drought is one of the most significant limiting factors affecting plant growth and development on the Qinghai-Tibet Plateau (QTP). Mining the drought-tolerant genes of the endemic perennial grass of the QTP, Siberian wildrye (Elymus sibiricus), is of great significance to creating new drought-resistant varieties which can be used in the development of grassland livestock and restoring natural grassland projects in the QTP. To investigate the transcriptomic responsiveness of E. sibiricus to drought stress, PEG-induced short- and long-term drought stress was applied to two Siberian wildrye genotypes (drought-tolerant and drought-sensitive accessions), followed by third- and second-generation transcriptome sequencing analysis. A total of 40,708 isoforms were detected, of which 10,659 differentially expressed genes (DEGs) were common to both genotypes. There were 2107 and 2498 unique DEGs in the drought-tolerant and drought-sensitive genotypes, respectively. Additionally, 2798 and 1850 DEGs were identified in the drought-tolerant genotype only under short- and long-term conditions, respectively. DEGs numbering 1641 and 1330 were identified in the drought-sensitive genotype only under short- and long-term conditions, respectively. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed that all the DEGs responding to drought stress in E. sibiricus were mainly associated with the mitogen-activated protein kinase (MAKP) signaling pathway, plant hormone signal transduction, the linoleic acid metabolism pathway, the ribosome pathway, and plant circadian rhythms. In addition, Nitrate transporter 1/Peptide transporter family protein 3.1 (NPF3.1) and Auxin/Indole-3-Acetic Acid (Aux/IAA) family protein 31(IAA31) also played an important role in helping E. sibiricus resist drought. This study used transcriptomics to investigate how E. sibiricus responds to drought stress, and may provide genetic resources and references for research into the molecular mechanisms of drought resistance in native perennial grasses and for breeding drought-tolerant varieties.

Complete chloroplast genomes of three wild perennial Hordeum species from Central Asia: genome structure, mutation hotspot, phylogenetic relationships, and comparative analysis.

Hordeum L. is widely distributed in mountain or plateau of subtropical and warm temperate regions around the world. Three wild perennial Hordeum species, including H. bogdanii, H. brevisubulatum, and H. violaceum, have been used as forage and for grassland ecological restoration in high-altitude areas in recent years. To date, the degree of interspecies sequence variation in the three Hordeum species within existing gene pools is still not well-defined. Herein, we sequenced and assembled chloroplast (cp) genomes of the three species. The results revealed that the cp genome of H. bogdanii showed certain sequence variations compared with the cp genomes of the other two species (H. brevisubulatum and H. violaceum), and the latter two were characterized by a higher relative affinity. Parity rule 2 plot (PR2) analysis illuminated that most genes of all ten Hordeum species were concentrated in nucleotide T and G. Numerous single nucleotide polymorphism (SNP) and insertion/deletion (In/Del) events were detected in the three Hordeum species. A series of hotspots regions (tRNA-GGU ~ tRNA-GCA, tRNA-UGU ~ ndhJ, psbE ~ rps18, ndhF ~ tRNA-UAG, etc.) were identified by mVISTA procedures, and the five highly polymorphic genes (tRNA-UGC, tRNA-UAA, tRNA-UUU, tRNA-UAC, and ndhA) were proved by the nucleotide diversity (Pi). Although the distribution and existence of cp simple sequence repeats (cpSSRs) were predicted in the three Hordeum cp genomes, no rearrangement was found between them. A similar phenomenon has been found in the cp genome of the other seven Hordeum species, which has been published so far. In addition, evolutionary relationships were reappraised based on the currently reported cp genome of Hordeum L. This study offers a framework for gaining a better understanding of the evolutionary history of Hordeum species through the re-examination of their cp genomes, and by identifying highly polymorphic genes and hotspot regions that could provide important insights into the genetic diversity and differentiation of these species.