Author Correction: The Apostasia genome and the evolution of orchids.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

An Enzyme-Cleavable Solubilizing-Tag Facilitates the Chemical Synthesis of Mirror-Image Proteins.

Mirror-image proteins (D-proteins) are useful in biomedical research for purposes such as mirror-image screening for D-peptide drug discovery, but the chemical synthesis of many D-proteins is often low yielding due to the poor solubility or aggregation of their constituent peptide segments. Here, we report a Lys-C protease-cleavable solubilizing tag and its use to synthesize difficult-to-obtain D-proteins. Our tag is easily installed onto multiple amino acids such as DLys, DSer, DThr, and/or the N-terminal amino acid of hydrophobic D-peptides, is impervious to various reaction conditions, such as peptide synthesis, ligation, desulfurization, and transition metal-mediated deprotection, and yet can be completely removed by Lys-C protease under denaturing conditions to give the desired D-protein. The efficacy and practicality of the new method were exemplified in the synthesis of two challenging D-proteins: D-enantiomers of programmed cell death protein 1 IgV domain and SARS-CoV-2 envelope protein, in high yield. This work demonstrates that the enzymatic cleavage of solubilizing tags under denaturing conditions is feasible, thus paving the way for the production of more D-proteins.

The Apostasia genome and the evolution of orchids.

Constituting approximately 10% of flowering plant species, orchids (Orchidaceae) display unique flower morphologies, possess an extraordinary diversity in lifestyle, and have successfully colonized almost every habitat on Earth. Here we report the draft genome sequence of Apostasia shenzhenica, a representative of one of two genera that form a sister lineage to the rest of the Orchidaceae, providing a reference for inferring the genome content and structure of the most recent common ancestor of all extant orchids and improving our understanding of their origins and evolution. In addition, we present transcriptome data for representatives of Vanilloideae, Cypripedioideae and Orchidoideae, and novel third-generation genome data for two species of Epidendroideae, covering all five orchid subfamilies. A. shenzhenica shows clear evidence of a whole-genome duplication, which is shared by all orchids and occurred shortly before their divergence. Comparisons between A. shenzhenica and other orchids and angiosperms also permitted the reconstruction of an ancestral orchid gene toolkit. We identify new gene families, gene family expansions and contractions, and changes within MADS-box gene classes, which control a diverse suite of developmental processes, during orchid evolution. This study sheds new light on the genetic mechanisms underpinning key orchid innovations, including the development of the labellum and gynostemium, pollinia, and seeds without endosperm, as well as the evolution of epiphytism; reveals relationships between the Orchidaceae subfamilies; and helps clarify the evolutionary history of orchids within the angiosperms.

The Euscaphis japonica genome and the evolution of malvids.

Malvids is one of the largest clades of rosids, includes 58 families and exhibits remarkable morphological and ecological diversity. Here, we report a high-quality chromosome-level genome assembly for Euscaphis japonica, an early-diverging species within malvids. Genome-based phylogenetic analysis suggests that the unstable phylogenetic position of E. japonica may result from incomplete lineage sorting and hybridization event during the diversification of the ancestral population of malvids. Euscaphis japonica experienced two polyploidization events: the ancient whole genome triplication event shared with most eudicots (commonly known as the γ event) and a more recent whole genome duplication event, unique to E. japonica. By resequencing 101 samples from 11 populations, we speculate that the temperature has led to the differentiation of the evergreen and deciduous of E. japonica and the completely different population histories of these two groups. In total, 1012 candidate positively selected genes in the evergreen were detected, some of which are involved in flower and fruit development. We found that reddening and dehiscence of the E. japonica pericarp and long fruit-hanging time promoted the reproduction of E. japonica populations, and revealed the expression patterns of genes related to fruit reddening, dehiscence and abscission. The key genes involved in pentacyclic triterpene synthesis in E. japonica were identified, and different expression patterns of these genes may contribute to pentacyclic triterpene diversification. Our work sheds light on the evolution of E. japonica and malvids, particularly on the diversification of E. japonica and the genetic basis for their fruit dehiscence and abscission.

Development and characterization of a glycine biosensor system for fine-tuned metabolic regulation in Escherichia coli.

BACKGROUND: In vivo biosensors have a wide range of applications, ranging from the detection of metabolites to the regulation of metabolic networks, providing versatile tools for synthetic biology and metabolic engineering. However, in view of the vast array of metabolite molecules, the existing number and performance of biosensors is far from sufficient, limiting their potential applications in metabolic engineering. Therefore, we developed the synthetic glycine-ON and -OFF riboswitches for metabolic regulation and directed evolution of enzyme in Escherichia coli. RESULTS: The results showed that a synthetic glycine-OFF riboswitch (glyOFF6) and an increased-detection-range synthetic glycine-ON riboswitch (glyON14) were successfully screened from a library based on the Bacillus subtilis glycine riboswitch using fluorescence-activated cell sorting (FACS) and tetA-based dual genetic selection. The two synthetic glycine riboswitches were successfully used in tunable regulation of lactate synthesis, dynamic regulation of serine synthesis and directed evolution of alanine-glyoxylate aminotransferase in Escherichia coli, respectively. Mutants AGXT22 and AGXT26 of alanine-glyoxylate aminotransferase with an increase of 58% and 73% enzyme activity were obtained by using a high-throughput screening platform based on the synthetic glycine-OFF riboswitch, and successfully used to increase the 5-aminolevulinic acid yield of engineered Escherichia coli. CONCLUSIONS: A synthetic glycine-OFF riboswitch and an increased-detection-range synthetic glycine-ON riboswitch were successfully designed and screened. The developed riboswitches showed broad application in tunable regulation, dynamic regulation and directed evolution of enzyme in E. coli.

Effect of Cognitive-Behavioral Therapy or Mindfulness Therapy on Pain and Quality of Life in Patients with Diabetic Neuropathy: A Systematic Review and Meta-Analysis.

OBJECTIVES: This study aimed to evaluate the effectiveness of cognitive behavioral therapy (CBT) and mindfulness therapy (MT) for pain relief and quality of life (QOL) in patients with diabetic neuropathy. REVIEW/ANALYSIS METHODS: Four databases were systematically searched from their respective inception dates to 29 June 2021. Relevant randomized controlled trials (RCTs) were screened and assessed for risk of bias. Eight RCTs evaluating CBT or MT were included. Statistical analysis was performed using Review Manager 5.4. RESULTS: Eight RCTs involving 384 patients with painful diabetic neuropathy (PDN) tested psychological interventions, including three CBT and five MT studies. The results showed that patients' pain severity (standardized mean difference [SMD] = -0.60, 95% confidence interval [CI; -0.93 to -0.27], P = .0003) and QOL (SMD = -0.43, 95% CI [-0.83 to -0.04], p = .03) were improved immediately after treatment. Besides, the pain intensity (SMD = -0.67, 95% CI [-1.37 to 0.03], p = .06), pain interference (SMD = -0.75, 95% CI [-1.20 to -0.30], p = .001) and depressive symptoms (SMD = -0.62, 95% CI [-0.96 to -0.28], p = .0003) were superior to the control group after follow up. The subgroup analysis results of different intervention type showed that the CBT group could immediately improve pain (SMD = -0.44, 95% CI [-0.78 to -0.10], p = .01) after treatment. However, there was no statistically significant difference in the CBT group after follow-up (SMD = -0.15, 95% CI [-0.52 to 0.22], p = .42). CONCLUSIONS: Cognitive behavioral therapy or MT is effective for treating pain in patients with diabetic peripheral neuropathy, improving the QOL, and reducing depressive symptoms. However, large-scale, multi-centre, rigorously designed RCTs are needed to further verify the long-term effects.

Phosphorescent Calcium-Based Metal-Organic Framework with Second-Scale Long Afterglow.

Presented here is a calcium-based metal-organic framework (Ca-MOF) with obvious room temperature phosphorescence. Notably, a long afterglow can be observed by the naked eye and lasts about 4 s, which is mainly attributed to the unique framework structure of the Ca-MOF.

Molecular cloning, characterization and expression profiles of CD2AP in Nile tilapia (Oreochromis niloticus) responding to Streptococcus agalactiae infection and interaction with CD2 cytoplasmic segment.

The interaction between CD2-associated protein (CD2AP) and CD2 plays a vital role in lymphocyte adhesion and T cells activation in mammals. In this study, a CD2AP gene (GenBank accession number: MK579862; designated as On-CD2AP) was identified from tilapia (Oreochromis niloticus). Sequence analysis showed that On-CD2AP protein shares high similarity with mammals, including three Src homology 3 (SH3) domains, a section of poly proline motif and a coiled coil region. Transcription levels of On-CD2AP were detected in nine tissues of healthy Nile tilapia, and the highest expression levels were detected in the spleen and gill. On-CD2AP were significantly up-regulated in thymus, head kidney and brain after infected by Streptococcus agalactiae, as well as in head kidney leukocytes (HKLs) with LPS and LTA stimulation. Moreover, a section conserved pro-rich motif that are responsible for binding of CD2 to CD2AP were found in the CD2 cytoplasmic sequence of Nile tilapia (On-CD2C). A weak interaction between On-CD2AP and On-CD2C was proved by yeast two-hybrid assay. In addition, the recombinant proteins of CD2AP-His (rOn-CD2AP-His) and GST-CD2C (GST-rOn-CD2C) were obtained through prokaryotic expression system. His pull-down assay showed that rOn-CD2AP-His and GST-rOn-CD2C could bind to each other. These findings indicate that CD2AP is crucial in immune response during S.agalactiae infection, and the mechanism of interaction between CD2AP and CD2 is conservative in Nile tilapia.

Intrinsic interaction between in-plane ferroelectric polarization and surface adsorption.

The chemical properties of a ferroelectric surface are polarization dependent, the underlying nature of which is, however, far from being completely understood. One of the reasons is that when the polarization direction is perpendicular to the surface, the depolarization field may induce electronic or atomic reconstruction and thus change the chemistry of the ferroelectric surface in complicated ways. Instead, the in-plane polarization results in no depolarization field. Therefore, the chemical properties of a ferroelectric surface can be more intrinsically reflected by the interplay between the in-plane polarization and the surface adsorption. By using first-principles calculations, we study the effect of the strain-induced in-plane polarization on the adsorption of a series of molecules on the reduced rutile TiO2(110) surface. We reveal that it is the surface doping caused by the charge transfer between the adsorbates and the TiO2(110) surface that dominates the polarization-induced change of the adsorption energy, as a result of screening long-range Coulomb interactions. The electrostatic interaction between the polarization of the substrate and the polar molecule is of relatively less importance. We propose that charge transfer effects generally occur for ferroelectric surfaces with no localized surface states.

Recent advances in CRISPR/Cas9 mediated genome editing in Bacillus subtilis.

Genome editing using engineered nucleases has rapidly transformed from a niche technology to a mainstream method used in various host cells. Its widespread adoption has been largely developed by the emergence of the clustered regularly interspaced short palindromic repeats (CRISPR) system, which uses an easily customizable specificity RNA-guided DNA endonuclease, such as Cas9. Recently, CRISPR/Cas9 mediated genome engineering has been widely applied to model organisms, including Bacillus subtilis, enabling facile, rapid high-fidelity modification of endogenous native genes. Here, we reviewed the recent progress in B. subtilis gene editing using CRISPR/Cas9 based tools, and highlighted state-of-the-art strategies for design of CRISPR/Cas9 system. Finally, future perspectives on the use of CRISPR/Cas9 genome engineering for sequence-specific genome editing in B. subtilis are provided.

Evaluation of the Sensitivity and Specificity of the New Clinical Diagnostic and Classification Criteria for Kashin-Beck Disease, an Endemic Osteoarthritis, in China.

This study aimed to evaluate the sensitivity and specificity of the new clinical diagnostic and classification criteria for Kashin-Beck disease (KBD) using six clinical markers: flexion of the distal part of fingers, deformed fingers, enlarged finger joints, shortened fingers, squat down, and dwarfism. One-third of the total population in Linyou County was sampled by stratified random sampling. The survey included baseline characteristics and clinical diagnoses, and the sensitivity and specificity of the new criteria was evaluated. We identified 3,459 KBD patients, of which 69 had early stage KBD, 1,952 had stage I, 1,132 had stage II, and 306 had stage III. A screening test classified enlarged finger joints as stage I KBD, with a sensitivity and specificity of 0.978 and 0.045, respectively. Shortened fingers were classified as stage II KBD, with a sensitivity and specificity of 0.969 and 0.844, respectively, and dwarfism was classified as stage III KBD with a sensitivity and specificity of 0.951 and 0.992, respectively. Serial screening test revealed that the new clinical classification of KBD classified stages I, II, and III KBD with sensitivities of 0.949, 0.945, and 0.925 and specificities of 0.967, 0.970, and 0.993, respectively. The screening tests revealed that enlarged finger joints, shortened fingers, and dwarfism were appropriate markers for the clinical diagnosis and classification of KBD with high sensitivity and specificity.

Comparative transcriptome analysis of eggplant (Solanum melongena L.) and turkey berry (Solanum torvum Sw.): phylogenomics and disease resistance analysis.

BACKGROUND: Eggplant (Solanum melongena L.) and turkey berry (S. torvum Sw.), a wild ally of eggplant with promising multi-disease resistance traits, are of great economic, medicinal and genetic importance, but genomic resources for these species are lacking. In the present study, we sequenced the transcriptomes of eggplant and turkey berry to accelerate research on these two non-model species. RESULTS: We built comprehensive, high-quality de novo transcriptome assemblies of the two Leptostemonum clade Solanum species from short-read RNA-Sequencing data. We obtained 34,174 unigenes for eggplant and 38,185 unigenes for turkey berry. Functional annotations based on sequence similarity to known plant datasets revealed a distribution of functional categories for both species very similar to that of tomato. Comparison of eggplant, turkey berry and another 11 plant proteomes resulted in 276 high-confidence single-copy orthologous groups, reasonable phylogenetic tree inferences and reliable divergence time estimations. From these data, it appears that eggplant and its wild Leptostemonum clade relative turkey berry split from each other in the late Miocene, ~6.66 million years ago, and that Leptostemonum split from the Potatoe clade in the middle Miocene, ~15.75 million years ago. Furthermore, 621 and 815 plant resistance genes were identified in eggplant and turkey berry respectively, indicating the variation of disease resistance genes between them. CONCLUSIONS: This study provides a comprehensive transcriptome resource for two Leptostemonum clade Solanum species and insight into their evolutionary history and biological characteristics. These resources establish a foundation for further investigations of eggplant biology and for agricultural improvement of this important vegetable. More generally, we show that RNA-Seq is a fast, reliable and cost-effective method for assessing genome evolution in non-model species.

[Construction and identification of anti-HER2 phage display single chain fragment of variable region library in human breast cancer].

OBJECTIVE: To construct human phage single-chain antibody (scFv) library against breast cancer, and to identify anti-HER2 specific antibodies from the human phage display scFv library to offer a stronger affinity sequence targeting HER2 for fusion protein targeting HER2 and CXCR4. METHODS: Total RNA was extracted from the adjacent lymphatic tissue harvested from breast cancer patients. The variable regions of the whole antibody were amplified by using RT-PCR and were cloned into the vector pCANTAB-5E through a linker. The products were electroporated into competent E.coli TG1 cells. Recombinant phages specific for breast cancer cells were enriched in SKBR-3 after four rounds. The antigen-positive clones were selected by ELISA and immunohistochemistry. RESULTS: The fragment of VH and VL were about 375 and 330 bp and were linked in vitro to form scFv of 750 bp that was resistant to the breast cancer HER2 single strand. A fusion phage display library that contained total of 2.48×10(8) pfu /ml was established. ELISA and immunohistochemical results confirmed that the antibody has a strong affinity with HER2 antigen in breast cancer tissue. Compared to human IgG antibody, a scFv phage library against human breast cancer was successfully constructed with high capacity. The scFv was highly specific to HER2 antigen and the sequencing results indicated that VL and VH genes were highly homologous with the variable region of human antibody. CONCLUSION: This strategy may achieve new targeted antibody resistant to the breast cancer for clinical treatment and provide a carrier that uses HER2 as a target of the fusion protein for anti-tumor therapy.

MAGE-A11 is a potential prognostic biomarker and immunotherapeutic target in gastric cancer.

Gastric cancer poses a serious threat to human health and affects the digestive system. The lack of early symptoms and a dearth of effective identification methods make diagnosis difficult, with many patients only receiving a definitive diagnosis at a malignant stage, causing them to miss out on optimal therapeutic interventions. Melanoma-associated antigen-A (MAGE-A) is part of the MAGE family and falls under the cancer/testis antigen (CTA) category. The MAGE-A subfamily plays a significant role in tumorigenesis, proliferation and migration. The expression, prognosis and function of MAGE-A family members in GC, however, remain unclear. Our research and screening have shown that MAGE-A11 was highly expressed in GC tissues and was associated with poor patient prognosis. Additionally, MAGE-A11 functioned as an independent prognostic factor in GC through Cox regression analysis, and its expression showed significant correlation with both tumour immune cell infiltration and responsiveness to immunotherapy. Our data further indicated that MAGE-A11 regulated GC cell proliferation and migration. Subsequently, our findings propose that MAGE-A11 may operate as a prognostic factor, having potential as an immunotherapy target for GC.

LINC00525 enhances ZNF460-regulated CD24 expression through the sponge miR-125a-5p to promote malignant progression of breast cancer.

INTRODUCTION: CD24 is a highly glycosylated glycosylphosphatidylinositol anchored membrane protein that plays an important role in tumor progression. The aim of this study was to investigate the effect of abnormal expression of CD24 on the proliferation, migration and invasion of breast cancer (BC) cells, and the molecular mechanism of regulating CD24 expression in breast cancer. METHODOLOGY: The bioinformatics method was used to predict the expression level of CD24 in BC and its relationship with the occurrence and development of BC. IHC, RT-qPCR and WB were used to detect the expression of CD24 in BC tissues and cells. The proliferation of CD24 was evaluated by CCK-8 and colony formation assay, and the migration and invasion of CD24 were evaluated by wound healing and transwell. In addition, the effect of CD24 on the malignancy of BC in vivo was further evaluated by subcutaneous tumorigenesis assay. Molecular mechanisms were measured by luciferase reporter assays, biotin-labeled miRNA pull-down assay, RIP, and western blotting. RESULTS: The results show that CD24 is highly expressed in breast cancer tissues and cell lines, and knockdown of CD24 in vivo and in vitro can inhibit the proliferation, migration and invasion of BC cells. Mechanistically, the transcription factor ZNF460 promotes its expression by binding to the CD24 promoter, and the expression of ZNF460 is regulated by miR-125a-5p, which inhibits its expression by targeting the 3'UTR of ZNF460. In addition, LINC00525 acts as a ceRNA sponge to adsorb miR-125a-5p and regulate its expression. CONCLUSIONS: Overexpression of CD24 is involved in the development and poor prognosis of BC, which can be used as a potential target for the treatment of BC and provide a theoretical basis for the treatment of BC.

Coping with alpine habitats: genomic insights into the adaptation strategies of Triplostegia glandulifera (Caprifoliaceae).

How plants find a way to thrive in alpine habitats remains largely unknown. Here we present a chromosome-level genome assembly for an alpine medicinal herb, Triplostegia glandulifera (Caprifoliaceae), and 13 transcriptomes from other species of Dipsacales. We detected a whole-genome duplication event in T. glandulifera that occurred prior to the diversification of Dipsacales. Preferential gene retention after whole-genome duplication was found to contribute to increasing cold-related genes in T. glandulifera. A series of genes putatively associated with alpine adaptation (e.g. CBFs, ERF-VIIs, and RAD51C) exhibited higher expression levels in T. glandulifera than in its low-elevation relative, Lonicera japonica. Comparative genomic analysis among five pairs of high- vs low-elevation species, including a comparison of T. glandulifera and L. japonica, indicated that the gene families related to disease resistance experienced a significantly convergent contraction in alpine plants compared with their lowland relatives. The reduction in gene repertory size was largely concentrated in clades of genes for pathogen recognition (e.g. CNLs, prRLPs, and XII RLKs), while the clades for signal transduction and development remained nearly unchanged. This finding reflects an energy-saving strategy for survival in hostile alpine areas, where there is a tradeoff with less challenge from pathogens and limited resources for growth. We also identified candidate genes for alpine adaptation (e.g. RAD1, DMC1, and MSH3) that were under convergent positive selection or that exhibited a convergent acceleration in evolutionary rate in the investigated alpine plants. Overall, our study provides novel insights into the high-elevation adaptation strategies of this and other alpine plants.

Comparative Efficacy of Six Active Warming Systems for Intraoperative Warming in Adult Patients Undergoing Laparoscopic Surgery: A Systematic Review and Network Meta-Analysis.

Intraoperative hypothermia is very common and harmful in adult patients undergoing laparoscopic surgery. A variety of active warming systems has received close attention and has been researched by related scholars. However, the relative efficacy of these systems and which active warming system is preferred for such patients remain unclear. The aim of this study was to compare and rank six active warming systems regarding intraoperative warming efficacy in adult patients undergoing laparoscopic surgery. Following the PRISMA 2020 guidelines, relevant randomized controlled trials (RCTs) on the efficacy of different active warming systems in warming adult patients undergoing laparoscopic surgery were searched from five English databases and three Chinese databases. The quality of the studies was assessed using the Cochrane Risk of Bias tool (RoB2). The outcome was the final intraoperative core temperature. We estimated direct effects by using pairwise meta-analysis, estimated relative effects and ranking with the consistency model to conduct an NetworkMeta-Analysis (NMA). We used GRADE (Grading of Recommendations Assessment, Development, and Evaluation) to assess the certainty of the evidence. Sensitivity analysis was performed to test the robustness of the results. This study is registered with PROSPERO, with number CRD42022309057. In total, 19 RCTs involving 6 active warming systems and comprising 1364 patients were included in this NMA. The NMA once again confirmed the validity of forced-air warming (FAW) systems compared with other active warming systems, and further showed that underbody FAW was associated with more remarkable warming efficacy in different types of FAW systems. NMA was used to perform an exhaustive comparison of the warming efficacy of six active warming systems and indicated that underbody FAW was most likely to be the most effective warming system in adult patients undergoing laparoscopic surgery; however, considering the sparsity of the network, our results should be cautiously interpreted. Furthermore, a large number of high-quality RCTs comparing the warming efficacy of different competitive active warming systems are needed.

GRB10 is a novel factor associated with gastric cancer proliferation and prognosis.

GRB10 and its family members GRB7 and GRB14 were important adaptor proteins. They regulated many cellular functions by interacting with various tyrosine kinase receptors and other phosphorus-containing amino acid proteins. More and more studies have shown that the abnormal expression of GRB10 is closely related to the occurrence and development of cancer. In our current research, expression data for 33 cancers from the TCGA database was downloaded for analysis. It was found that GRB10 was up-regulated in cholangiocarcinoma, colon adenocarcinoma, head and neck squamous carcinoma, renal chromophobe, clear renal carcinoma, hepatocellular carcinoma, lung adenocarcinoma, lung squamous carcinoma, gastric adenocarcinoma and thyroid carcinoma. Especially in gastric cancer, the high GRB10 expression was closely associated with poorer overall survival. Further research showed that the knockdown of GRB10 inhibited proliferation and migration ability in gastric cancer. Also, there was a potential binding site for miR-379-5p on the 3'UTR of GRB10. Overexpression of miR-379-5p in gastric cancer cells reduced GRB10-regulated gastric cancer proliferation and migration capacity. In addition, we found that tumor growth was slower in a mice xenograft model with knock down of GRB10 expression. These findings suggested that miR-379-5p suppresses gastric cancer development by downregulating GRB10 expression. Therefore, miR-379-5p and GRB10 were expected to be potential targets for the treatment of gastric cancer.

GNPNAT1 promotes the stemness of breast cancer and serves as a potential prognostic biomarker.

Glucosamine­phosphate N­acetyltransferase 1 (GNPNAT1) is a member of the acetyltransferase superfamily, related to general control non­depressible 5 (GCN5). It has been documented that GNPNAT1 expression is increased in lung cancer, whereas its involvement in breast cancer (BC) remains to be further investigated. The present study aimed to evaluate the expression levels of GNPNAT1 in BC and its effect on BC stem cells (BCSCs). The Cancer Genome Atlas (TCGA) database was used for the analysis of the expression of GNPNAT1 and its clinical significance. Cox regression and logistic regression analyses were used to evaluate prognosis­related factors. The GNPNAT1­binding protein network was constructed using the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) application. The biological signaling pathways implicated in GNPNAT1 were investigated through function enrichment analysis including Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, and gene set enrichment analysis. The single­sample GSEA method was used to investigate the connection between the level of immune infiltration and GNPNAT1 expression in BC. GNPNAT1 expression was upregulated in patients with BC and was significantly associated with a poor prognosis. GNPNAT1 and its co­expressed genes were mostly enriched in nuclear transport, Golgi vesicle transport, ubiquitin­like protein transferase activity and ribonucleoprotein complex binding, as determined using functional enrichment analysis. GNPNAT1 expression was positively associated with Th2 cells and T­helper cells, and negatively associated with plasmacytoid dendritic cells, CD8+ T­cells and cytotoxic cells. Additionally, the GNPNAT1 expression levels were considerably increased in BCSCs. GNPNAT1 knockdown markedly decreased the stemness ability of SKBR3 and Hs578T cells, including the production of CSC markers and mammosphere or clone formation, while GNPNAT1 overexpression increased the stemness level. Hence, the findings of the present study demonstrate that GNPNAT1 may be exploited as a novel prognostic biomarker and therapeutic target for BC.

Anomanolide C suppresses tumor progression and metastasis by ubiquitinating GPX4-driven autophagy-dependent ferroptosis in triple negative breast cancer.

Anomanolide C (AC), a natural withanolide isolated from Tubocapsicum anomalum, has been reported to have exhibits remarkable anti-tumour activities in several types of human cancers, particularly triple-negative breast cancer (TNBC). However, its intricate mechanisms still remain need to be clarified. Here, we evaluated whether AC could inhibit cell proliferation and the role of AC in ferroptosis induction and autophagy activation. Subsequently, the anti-migration potential of AC was found via autophagy-dependent ferroptosis. Additionally, we found that AC reduced the expression of GPX4 by ubiquitination and inhibited TNBC proliferation and metastasis in vitro and in vivo. Moreover, we demonstrated that AC induced autophagy-dependent ferroptosis, and led to Fe2+ accumulation via ubiquitinating GPX4. Moreover, AC was shown to induce autophagy-dependent ferroptosis as well as to inhibit TNBC proliferation and migration via GPX4 ubiquitination. Together, these results demonstrated that AC inhibited the progression and metastasis of TNBC by inducing autophagy-dependent ferroptosis via ubiquitinating GPX4, which might shed light on exploiting AC as a new drug candidate for the future TNBC therapy.