Hsa_circ_0136666 stimulates gastric cancer progression and tumor immune escape by regulating the miR-375/PRKDC Axis and PD-L1 phosphorylation.

BACKGROUND: Targeted drugs are not quite effective for prolonging the survival of patients with gastric cancer due to off-target effects as well as tumor immune escape mechanisms. Circular RNAs widely exist in tumor regions as biomarkers and can be developed as effective drug targets. METHODS: Western blot, QRT-PCR, fluorescence in situ hybridization, and flow cytometry were used to investigate the function of hsa_circ_0136666 in promoting the proliferation of gastric cancer cells. Tissue immunofluorescence, enzyme-linked immunosorbent assay (ELISA), as well as flow cytometric analysis, was conducted to explore the process of tumor immune evasion in tumor-bearing mice. The differences of circRNA expression in clinical samples were analyzed through tissue microarray FISH. The effect of siRNA on improving the efficacy of anti-PDL1 drugs and suppressing the immune microenvironment was evaluated by the coadministration model. RESULTS: We demonstrated that hsa_circ_0136666 was widely and highly expressed in gastric cancer tissues and cells. Functionally, hsa_circ_0136666 promoted gastric cancer tumor proliferation and tumor microenvironment formation, leading to tumorigenesis immune escape, and this effect was dependent on CD8 + T cells. Mechanistically, we confirmed that hsa_circ_0136666 competitively upregulated PRKDC expression by sponging miR-375-3p, regulating immune checkpoint proteins, prompting phosphorylation of PD-L1 to preventing its degradation, driving PD-L1 aggregation and suppressing immune function, thereby impairing cancer immune responses. In terms of application, we found that LNP-siRNA effectively improved anti-PDL1 drug efficacy and inhibited immune escape. CONCLUSION: Our results reveal an oncogenic role played by hsa_circ_0136666 in gastric cancer, driving PD-L1 phosphorylation via the miR-375/PRKDC signaling axis, prompting immune escape. This work proposes a completely new pathogenic mechanism of gastric cancer, uncovers a novel role for hsa_circ_0136666 as an immune target, and provides a rationale for enhancing the efficacy of anti-PD-L1 therapy for gastric cancer.

IPOP: An Integrative Plant Multi-omics Platform for Cross-species Comparison and Evolutionary Study.

The advent of high-throughput sequencing technologies has led to the production of a significant amount of omics data in plants, which serves as valuable assets for conducting cross-species multi-omics comparative analysis. Nevertheless, the current dearth of comprehensive platforms providing evolutionary annotation information and multi-species multi-omics data impedes users from systematically and efficiently performing evolutionary and functional analysis on specific genes. In order to establish an advanced plant multi-omics platform that provides timely, accurate, and high-caliber omics information, we collected 7 distinct types of omics data from 6 monocots, 6 dicots, and 1 moss, and reanalyzed these data using standardized pipelines. Additionally, we furnished homology information, duplication events, and phylostratigraphic stages of 13 species to facilitate evolutionary examination. Furthermore, the integrative plant omics platform (IPOP) is bundled with a variety of online analysis tools that aid users in conducting evolutionary and functional analysis. Specifically, the Multi-omics Integration Analysis tool is available to consolidate information from diverse omics sources, while the Transcriptome-wide Association Analysis tool facilitates the linkage of functional analysis with phenotype. To illustrate the application of IPOP, we conducted a case study on the YTH domain gene family, wherein we observed shared functionalities within orthologous groups and discerned variations in evolutionary patterns across these groups. To summarize, the IPOP platform offers valuable evolutionary insights and multi-omics data to the plant sciences community, effectively addressing the need for cross-species comparison and evolutionary research platforms. All data and modules within IPOP are freely accessible for academic purposes (http://omicstudio.cloud:4012/ipod/).

Evolutionary Implications of the RNA N6-Methyladenosine Methylome in Plants.

Epigenetic modifications play important roles in genome evolution and innovation. However, most analyses have focused on the evolutionary role of DNA modifications, and little is understood about the influence of posttranscriptional RNA modifications on genome evolution. To explore the evolutionary significance of RNA modifications, we generated transcriptome-wide profiles of N6-methyladenosine (m6A), the most prevalent internal modification of mRNA, for 13 representative plant species spanning over half a billion years of evolution. These data reveal the evolutionary conservation and divergence of m6A methylomes in plants, uncover the preference of m6A modifications on ancient orthologous genes, and demonstrate less m6A divergence between orthologous gene pairs with earlier evolutionary origins. Further investigation revealed that the evolutionary divergence of m6A modifications is related to sequence variation between homologs from whole-genome duplication and gene family expansion from local-genome duplication. Unexpectedly, a significant negative correlation was found between the retention ratio of m6A modifications and the number of family members. Moreover, the divergence of m6A modifications is accompanied by variation in the expression level and translation efficiency of duplicated genes from whole- and local-genome duplication. Our work reveals new insights into evolutionary patterns of m6A methylomes in plant species and their implications, and provides a resource of plant m6A profiles for further studies of m6A regulation and function in an evolutionary context.

Tanshinone IIA inhibits gastric cancer cell stemness through inducing ferroptosis.

Tanshinone IIA is the active constituent extracted from Salvia Miltiorrhza. Numerous studies have shown that Tanshinone IIA could inhibit tumor proliferation and metastasis, including gastric cancer. However, the effect of Tanshinone IIA on gastric cancer cell stemness stays unclear. Here, we found that Tanshinone IIA could reduce gastric cancer cell stemness through detecting spheroid-forming, flow cytometry analysis, and the expression of stemness markers (OCT3/4, ALDH1A1, and CD44). Mechanistically, Tanshinone IIA increased the level of lipid peroxides and decreased glutathione level in gastric cancer cells, both of which are the markers of ferroptosis. Similarly, ferroptosis inducers (erastin, sulfasalazine, and sorafenib) reduced gastric cancer cell stemness. Additionally, the inhibitory effects of Tanshinone IIA on GC cell stemness were reversed by ferroptosis inhibitor (Fer-1) or overexpression of SLC7A11, which is a critical ferroptosis inhibitor. Therefore, we revealed that Tanshinone IIA inhibited the stemness of gastric cancer cells partly through inducing ferroptosis.

Integrated Analysis of Large-Scale Omics Data Revealed Relationship Between Tissue Specificity and Evolutionary Dynamics of Small RNAs in Maize (Zea mays).

The evolutionary dynamics and tissue specificity of protein-coding genes are well documented in plants. However, the evolutionary consequences of small RNAs (sRNAs) on tissue-specific functions remain poorly understood. Here, we performed integrated analysis of 195 deeply sequenced sRNA libraries of maize B73, representing more than 10 tissues, and identified a comprehensive list of 419 maize microRNA (miRNA) genes, 271 of which were newly discovered in this study. We further characterized the evolutionary dynamics and tissue specificity of miRNA genes and corresponding miRNA isoforms (isomiRs). Our analysis revealed that tissue specificity of isomiR events tends to be associated with miRNA gene abundance and suggested that the frequencies of isomiR types are affected by the local genomic regions. Moreover, genome duplication (GD) events have dramatic effect on evolutionary dynamics of maize miRNA genes, and the abundance divergence for tissue-specific miRNA genes is associated with GD events. Further study indicated that duplicate miRNA genes with tissue-specific expression patterns, such as miR2275a, a phased siRNA (phasiRNA) trigger, contribute to phenotypic traits in maize. Additionally, our study revealed the expression preference of 21- and 24-nt phasiRNAs in relation to tissue specificity. This large-scale sRNAomic study depicted evolutionary implications of tissue-specific maize sRNAs, which coordinate genome duplication, isomiR modification, phenotypic traits and phasiRNAs differentiation.

Evolution of the RNA N 6-Methyladenosine Methylome Mediated by Genomic Duplication.

RNA N 6-methyladenosine (m6A) modification is the most abundant form of RNA epigenetic modification in eukaryotes. Given that m6A evolution is associated with the selective constraints of nucleotide sequences in mammalian genomes, we hypothesize that m6A evolution can be linked, at least in part, to genomic duplication events in complex polyploid plant genomes. To test this hypothesis, we presented the maize (Zea mays) m6A modification landscape in a transcriptome-wide manner and identified 11,968 m6A peaks carried by 5,893 and 3,811 genes from two subgenomes (maize1 and maize2, respectively). Each of these subgenomes covered over 2,200 duplicate genes. Within these duplicate genes, those carrying m6A peaks exhibited significant differences in retention rate. This biased subgenome fractionation of m6A-methylated genes is associated with multiple sequence features and is influenced by asymmetric evolutionary rates. We also characterized the coevolutionary patterns of m6A-methylated genes and transposable elements, which can be mediated by whole genome duplication and tandem duplication. We revealed the evolutionary conservation and divergence of duplicated m6A functional factors and the potential role of m6A modification in maize responses to drought stress. This study highlights complex interplays between m6A modification and gene duplication, providing a reference for understanding the mechanisms underlying m6A evolution mediated by genome duplication events.

Evolution of intron-poor clades and expression patterns of the glycosyltransferase family 47.

MAIN CONCLUSION: A large-scale bioinformatics analysis revealed the origin and evolution of GT47 gene family, and identified two clades of intron-poor genes with putative functions in drought stress responses and seed development in maize. Glycosyltransferase family 47 (GT47) genes encode ß-galactosyltransferases and ß-glucuronyltransferases that synthesize pectin, xyloglucans and xylan, which are important components of the plant cell wall. In this study, we performed a systematic and large-scale bioinformatics analysis of GT47 gene family using 352 GT47 proteins from 15 species ranging from cyanobacteria to seed plants. The analysis results showed that GT47 family may originate in cyanobacteria and expand along the evolutionary trajectory to moss. Further analysis of 47 GT47 genes in maize revealed that they can divide into five clades with diverse exon-intron structures. Among these five clades, two were mainly composed with intron-poor genes, which may originate in the moss. Gene duplication analysis revealed that the expansion of GT47 gene family in maize was significantly driven from tandem duplication events and segmental duplication events. Significantly, almost all duplicated genes are intron-poor genes. Expression analysis indicated that several intron-poor GT47 genes may be involved in the drought stress response and seed development in maize. This work provides insight into the origin and evolutionary process, expansion mechanisms and expression patterns of GT47 genes, thus facilitating their functional investigations in the future.

A systems approach to a spatio-temporal understanding of the drought stress response in maize.

Crops are often subjected to periods of drought stress during their life cycle. However, how stress response mechanisms contribute to the crosstalk between stress signaling pathways and developmental signaling pathways is still unknown. We built a gene co-expression network from a spatio-temporal transcriptomic map of the drought stress response in maize (Zea mays), profiled from three tissues and four developmental stages and characterized hub genes associated with duplication events, selection, and regulatory networks. Co-expression analysis grouped drought-response genes into ten modules, covering 844 highly connected genes (hub genes). Of these, 15.4% hub genes had diverged by whole-genome duplication events and 2.5% might then have been selected during natural domestication and artificial improvement processes, successively. We identified key transcription factor hubs in a transcriptional regulatory network, which may function as a crosstalk mechanism between drought stress and developmental signalling pathways in maize. Understanding the evolutionary biases that have evolved to enhance drought adaptation lays the foundation for further dissection of crosstalk between stress signalling pathways and developmental signalling pathways in maize, towards molecular design of new cultivars with desirable yield and greater stress tolerance.

Opposing Control by Transcription Factors MYB61 and MYB3 Increases Freezing Tolerance by Relieving C-Repeat Binding Factor Suppression.

Cold acclimation is an important process by which plants respond to low temperature and enhance their winter hardiness. C-REPEAT BINDING FACTOR1 (CBF1), CBF2, and CBF3 genes were shown previously to participate in cold acclimation in Medicago truncatula In addition, MtCBF4 is transcriptionally induced by salt, drought, and cold stresses. We show here that MtCBF4, shown previously to enhance drought and salt tolerance, also positively regulates cold acclimation and freezing tolerance. To identify molecular factors acting upstream and downstream of the MtCBF4 transcription factor (TF) in cold responses, we first identified genes that are differentially regulated upon MtCBF4 overexpression using RNAseq Digital Gene Expression Profiling. Among these, we showed that MtCBF4 directly activates the transcription of the COLD ACCLIMATION SPECIFIC15 (MtCAS15) gene. To gain insights into how MtCBF4 is transcriptionally regulated in response to cold, an R2R3-MYB TF, MtMYB3, was identified based on a yeast one-hybrid screen as binding directly to MYB cis-elements in the MtCBF4 promoter, leading to the inhibition of MtCBF4 expression. In addition, another MYB TF, MtMYB61, identified as an interactor of MtMYB3, can relieve the inhibitory effect of MtMYB3 on MtCBF4 transcription. This study, therefore, supports a model describing how MtCBF4 is regulated by antagonistic MtMYB3/MtMYB61 TFs, leading to the up-regulation of downstream targets such as MtCAS15 acting in cold acclimation in M. truncatula.

De novo transcriptome analysis of Medicago falcata reveals novel insights about the mechanisms underlying abiotic stress-responsive pathway.

BACKGROUND: The entire world is facing a deteriorating environment. Understanding the mechanisms underlying plant responses to external abiotic stresses is important for breeding stress-tolerant crops and herbages. Phytohormones play critical regulatory roles in plants in the response to external and internal cues to regulate growth and development. Medicago falcata is one of the stress-tolerant candidate leguminous species and is able to fix atmospheric nitrogen. This ability allows leguminous plants to grow in nitrogen deficient soils. METHODS: We performed Illumina sequencing of cDNA prepared from abiotic stress treated M. falcata. Sequencedreads were assembled to provide a transcriptome resource. Transcripts were annotated using BLASTsearches against the NCBI non-redundant database and gene ontology definitions were assigned. Acomparison among the three abiotic stress treated samples was carried out. The expression of transcriptswas confirmed with qRT-PCR. RESULTS: We present an abiotic stress-responsive M. falcata transcriptome using next-generation sequencing data from samples grown under standard, dehydration, high salinity, and cold conditions. We combined reads from all samples and de novo assembled 98,515 transcripts to build the M. falcata gene index. A comprehensive analysis of the transcriptome revealed abiotic stress-responsive mechanisms underlying the metabolism and core signalling components of major phytohormones. We identified nod factor signalling pathways during early symbiotic nodulation that are modified by abiotic stresses. Additionally, a global comparison of homology between the M. falcata and M. truncatula transcriptomes, along with five other leguminous species, revealed a high level of global sequence conservation within the family. CONCLUSIONS: M. falcata is shown to be a model candidate for studying abiotic stress-responsive mechanisms in legumes. This global gene expression analysis provides new insights into the biochemical and molecular mechanisms involved in the acclimation to abiotic stresses. Our data provides many gene candidates that might be used for herbage and crop breeding. Additionally, FalcataBase ( http://bioinformatics.cau.edu.cn/falcata/ ) was built for storing these data.

GmHs1-1, encoding a calcineurin-like protein, controls hard-seededness in soybean.

Loss of seed-coat impermeability was essential in the domestication of many leguminous crops to promote the production of their highly nutritious seeds. Here we show that seed-coat impermeability in wild soybean is controlled by a single gene, GmHs1-1, which encodes a calcineurin-like metallophosphoesterase transmembrane protein. GmHs1-1 is primarily expressed in the Malpighian layer of the seed coat and is associated with calcium content. The transition from impermeability to permeability in domesticated soybean was caused by artificial selection of a point mutation in GmHs1-1. Interestingly, a number of soybean landraces evaded selection for permeability because of an alternative selection for seed-coat cracking that also enables seed imbibition. Despite the single origin of the mutant allele Gmhs1-1, the distribution pattern of allelic variants in the context of soybean population structure and the detected signature of genomic introgression between wild and cultivated soybeans suggest that Gmhs1-1 may have experienced reselection for seed-coat permeability.

Overexpression of MtCAS31 enhances drought tolerance in transgenic Arabidopsis by reducing stomatal density.

• Dehydrins are a type of late embryogenesis abundant protein. Some dehydrins are involved in the response to various abiotic stresses. Accumulation of dehydrins enhances the drought, cold and salt tolerances of transgenic plants, although the underlying mechanism is unclear. MtCAS31 (Medicago Truncatula cold-acclimation specific protein 31) is a Y(2)K(4)-type dehydrin that was isolated from Medicago truncatula. • We analyzed the subcellular and histochemical localization of MtCAS31, and the expression patterns of MtCAS31 under different stresses. Transgenic Arabidopsis that overexpressed MtCAS31 was used to determine the function of MtCAS31. A yeast two-hybrid assay was used to screen potential proteins that could interact with MtCAS31. The interaction was confirmed by bimolecular fluorescence complementation (BiFC) assay. • After a 3-h drought treatment, the expression of MtCAS31 significantly increased 600-fold. MtCAS31 overexpression dramatically reduced stomatal density and markedly enhanced the drought tolerance of transgenic Arabidopsis. MtCAS31 could interact with AtICE1 (inducer of CBF expression 1) and the AtICE1 homologous protein Mt7g083900.1, which was identified from Medicago truncatula both in vitro and in vivo. • Our findings demonstrate that a dehydrin induces decreased stomatal density. Most importantly, the interaction of MtCAS31 with AtICE1 plays a role in stomatal development. We hypothesize that the interaction of MtCAS31 and AtICE1 caused the decrease in stomatal density to enhance the drought resistance of transgenic Arabidopsis.

Medicago truncatula transporter database: a comprehensive database resource for M. truncatula transporters.

BACKGROUND: Medicago truncatula has been chosen as a model species for genomic studies. It is closely related to an important legume, alfalfa. Transporters are a large group of membrane-spanning proteins. They deliver essential nutrients, eject waste products, and assist the cell in sensing environmental conditions by forming a complex system of pumps and channels. Although studies have effectively characterized individual M. truncatula transporters in several databases, until now there has been no available systematic database that includes all transporters in M. truncatula. DESCRIPTION: The M. truncatula transporter database (MTDB) contains comprehensive information on the transporters in M. truncatula. Based on the TransportTP method, we have presented a novel prediction pipeline. A total of 3,665 putative transporters have been annotated based on International Medicago Genome Annotated Group (IMGAG) V3.5 V3 and the M. truncatula Gene Index (MTGI) V10.0 releases and assigned to 162 families according to the transporter classification system. These families were further classified into seven types according to their transport mode and energy coupling mechanism. Extensive annotations referring to each protein were generated, including basic protein function, expressed sequence tag (EST) mapping, genome locus, three-dimensional template prediction, transmembrane segment, and domain annotation. A chromosome distribution map and text-based Basic Local Alignment Search Tools were also created. In addition, we have provided a way to explore the expression of putative M. truncatula transporter genes under stress treatments. CONCLUSIONS: In summary, the MTDB enables the exploration and comparative analysis of putative transporters in M. truncatula. A user-friendly web interface and regular updates make MTDB valuable to researchers in related fields. The MTDB is freely available now to all users at http://bioinformatics.cau.edu.cn/MtTransporter/.