Bacterial effector restricts liquid-liquid phase separation of ZPR1 to antagonize host UPRER.

How pathogens manipulate host UPRER to mediate immune evasion is largely unknown. Here, we identify the host zinc finger protein ZPR1 as an interacting partner of the enteropathogenic E. coli (EPEC) effector NleE using proximity-enabled protein crosslinking. We show that ZPR1 assembles via liquid-liquid phase separation (LLPS) in vitro and regulates CHOP-mediated UPRER at the transcriptional level. Interestingly, in vitro studies show that the ZPR1 binding ability with K63-ubiquitin chains, which promotes LLPS of ZPR1, is disrupted by NleE. Further analyses indicate that EPEC restricts host UPRER pathways at the transcription level in a NleE-ZPR1 cascade-dependent manner. Together, our study reveals the mechanism by which EPEC interferes with CHOP-UPRER via regulating ZPR1 to help pathogens escape host defense.

YAP/miR-524-5p axis negatively regulates TXNIP expression to promote chondrosarcoma cell growth.

Chondrosarcoma (CHS) is the second most common bone malignant tumor and currently has limited treatment options. We have recently demonstrated that thioredoxin interacting protein (TXNIP) plays a crucial role in the oncogenesis of bone sarcoma, yet its implication in CHS is underdetermined. In the present study, we first found that knockdown of TXNIP promotes the proliferation of CHS cell largely through increasing their glycolytic metabolism, which is well-known as Warburg effect for providing energy. Consistent with our previous report that YAP is fundamental for CHS cell growth, herein we revealed that YAP functioned as an upstream molecule of TXNIP, and that YAP negatively regulated TXNIP mRNA and protein expression both in vitro and in vivo. Mechanistically, although knockdown of YAP upregulated both the nuclear and cytoplasmic TXNIP expression, we did not observe any obvious interaction between YAP and TXNIP; instead, miRNA-524-5p was demonstrated to be required for YAP-regulated TXNIP expression and thus controlling CHS cell growth. Together, our study reveals that TXNIP is a tumor suppressor in terms of CHS, and that the YAP/miRNA-524-5p/TXNIP signaling axis may provide a novel clue for CHS targeted therapy.

The boundary conditions by which body-esteem leads to eating disorders risk among adolescents.

Adolescents are believed to be susceptible to eating disorders (EDs) due to their serious fear of appearance evaluation from society. Related to this, low body-esteem has been found to be common among individuals with EDs. The present study mainly aimed to explore how emotional intelligence (EI), gender, and body size influence the relationship between body-esteem and EDs risk among adolescents. 128 middle school students classified as obese and 128 age-and gender-matched normal weight controls were included. All participants were asked to fill out self-report measures of body-esteem, EI, and EDs risk. The results showed that (1) both gender and body size directly influenced body-esteem and EDs risk; (2) EI acted as a moderator between body-esteem and EDs risk; and (3) both gender and body size interacted with EI and body-esteem to influence EDs risk. These findings contribute to our understanding of boundary conditions by which low body-esteem leads to EDs among adolescents, and help us to correspondingly conduct targeted intervention of adolescents' EDs.

KIF11 As a Potential Pan-Cancer Immunological Biomarker Encompassing the Disease Staging, Prognoses, Tumor Microenvironment, and Therapeutic Responses.

KIF11 is one of the 45 family members of kinesin superfamily proteins that functions as a motor protein in mitosis. Emerging evidence revealed that KIF11 plays pivotal roles in cancer initiation, development, and progression. However, the prognostic, oncological, and immunological values of KIF11 have not been comprehensively explored in pan-cancer. In present study, we comprehensively interrogated the role of KIF11 in tumor progression, tumor stemness, genomic heterogeneity, tumor immune infiltration, immune evasion, therapy response, and prognosis of cohorts from various cancer types. In general, KIF11 was significantly upregulated in tumors compared with paired normal tissues. KIF11 showed strong relationships with pathological stage, prognosis, tumor stemness, genomic heterogeneity, neoantigens, ESTIMATE, immune checkpoint, and drug sensitivity. The methylation level of KIF11 decreased in most cancers and was correlated with the survival probability in different human cancers. The expression of KIF11 was diverse in different molecular and immune subtypes and remarkably correlated with immune cell infiltration in the tumor microenvironment. Comparative study revealed that KIF11 was a powerful biomarker and associated with immune, targeted, and chemotherapeutic outcomes in various cancers. In addition, KIF11 interaction and coexpression networks mainly participated in the regulation of cell cycle, cell division, p53 signaling pathway, DNA repair and recombination, chromatin organization, antigen processing and presentation, and drug resistance. Our pan-cancer analysis provides a comprehensive understanding of the functions of KIF11 in oncogenesis, progression, and therapy in different cancers. KIF11 may serve as a potential prognostic and immunological pan-cancer biomarker. Moreover, KIF11 could be a novel target for tumor immunotherapy.

Capture and mass spectrometry analysis of effector-substrate complexes using genetically incorporated photo-crosslinkers in host cells.

Interactions between effectors and their host targets are often weak or transient, making them difficult to identify. We describe a protocol for covalent capture of effector substrates in living cells using genetic code expansion technology. The effector-substrate complexes are captured by the crosslinker and subsequently purified with tandem chromatography. We detail steps for mass spectrum analysis and substrate verification. While the steps here are specific for substrates of enteropathogenic E. coli in HEK293T cells, the protocol has broader applications. For complete details on the use and execution of this protocol, please refer to Li et al. (2021).1.

Network Pharmacology and Molecular Docking Study of Zhishi-Baizhu Herb Pair in the Treatment of Gastric Cancer.

OBJECTIVE: This study aimed to investigate the possible mechanism of the Zhishi and Baizhu herb pair in the treatment of gastric cancer by means of network pharmacology and molecular docking and to provide a theoretical basis for experiments and clinical application of traditional Chinese medicine for treating gastric cancer. METHODS: The main active chemical components of Zhishi and Baizhu were screened through Traditional Chinese Medicine Systems Pharmacology (TCMSP) database and selected by using the thresholds of oral bioavailability ≥30% and drug-likeness ≥18%. The targets of Zhishi and Baizhu were obtained from TCMSP, Therapeutic Targets Database (TTD), and the DrugBank database. The corresponding genes of the targets were retrieved from the UniProt database, and the gastric cancer targets were obtained from the GeneCards database and TTD. Subsequently, the networks were built between the main drug components, drug targets, and gastric cancer targets. Then, the enrichment analyses of GO and KEGG were applied to predict the potential roles of gastric cancer pathogenesis via the R package clusterProfiler. Finally, molecular docking was used to determine the affinity between the targets and components. RESULTS: Twenty-seven main active components were predicted from the Zhishi-Baizhu herb pair, and a total of 120 intersection genes were screened from 303 potential medicine genes and 1,839 disease genes. The enrichment included the PI3K-Akt and IL-17 signaling pathways, and the network analysis showed that the Zhishi-Baizhu herb pair acted on seven key targets, namely, AKT1, MMP9, IL-6, CCND1, BCL2, MTOR, and MDM2 (where they played a role in treating gastric cancer). Molecular docking showed that luteolin and naringenin could stably bind to the targets. CONCLUSION: The possible mechanisms of the components of the Zhishi-Baizhu herb pair in treating gastric cancer might be related to luteolin and naringenin, which intervened with the targets AKT1, MMP9, IL-6, CCND1, BCL2, MTOR, and MDM2, and are linked with the PI3K-Akt and IL-17 signaling pathways. This knowledge will lay a solid foundation for further experimental and clinical studies.

Genetically incorporated crosslinkers reveal NleE attenuates host autophagy dependent on PSMD10.

Autophagy acts as a pivotal innate immune response against infection. Some virulence effectors subvert the host autophagic machinery to escape the surveillance of autophagy. The mechanism by which pathogens interact with host autophagy remains mostly unclear. However, traditional strategies often have difficulty identifying host proteins that interact with effectors due to the weak, dynamic, and transient nature of these interactions. Here, we found that Enteropathogenic Escherichia coli (EPEC) regulates autophagosome formation in host cells dependent on effector NleE. The 26S Proteasome Regulatory Subunit 10 (PSMD10) was identified as a direct interaction partner of NleE in living cells by employing genetically incorporated crosslinkers. Pairwise chemical crosslinking revealed that NleE interacts with the N-terminus of PSMD10. We demonstrated that PSMD10 homodimerization is necessary for its interaction with ATG7 and promotion of autophagy, but not necessary for PSMD10 interaction with ATG12. Therefore, NleE-mediated PSMD10 in monomeric state attenuates host autophagosome formation. Our study reveals the mechanism through which EPEC attenuates host autophagy activity.

Nucleus pulposus cell apoptosis is attenuated by CDMP-2 through regulating oxidative damage under the hyperosmotic environment.

Disc nucleus pulposus (NP) cell experiences periodic osmolarity alterations during daily activities, which has been proved to affect cell biology in vitro The present study was aimed to investigate the effects of cartilage-derived morphogenetic protein-2 (CDMP-2) on NP cell apoptosis under the hyperosmolarity culture and the potential mechanism. Isolated rat NP cells were cultured in the in situ-osmolarity medium or hyperosmolarity medium for 3 days. CDMP-2 was added into the hyperosmolarity medium to investigate its effects on NP cell apoptosis. Cell apoptosis rate, caspase-3 activity, gene expression of Bcl-2, Bax, and caspase-3, and protein expression of Bcl-2, Bax, and cleaved caspase-3 were analyzed to evaluate NP cell apoptosis. Additionally, the intracellular reactive oxygen species (ROS) and the total superoxide dismutase (SOD) activity were analyzed to investigate the potential role of oxidative damage in this process. In the hyperosmolarity culture, NP cells showed a significantly increased cell apoptosis rate and caspase-3 activity, an up-regulated expression of Bax and caspase-3/cleaved-caspase-3 and a down-regulated expression of Bcl-2. However, CDMP-2 partly inhibited these effects of hyperosmolarity culture on NP cells. Additionally, the hyperosmolarity culture significantly increased ROS content and decreased the total SOD activity compared with the in situ-osmolarity culture, whereas exogenous CDMP-2 partly decreased the ROS content and increased the total SOD activity in the hyperosmolarity culture. In conclusion, CDMP-2 is effective in attenuating hyperosmolarity environment-induced NP cell apoptosis, and this process may be mediated through inhibiting oxidative stress damage. The present study indicates that CDMP-2 may be helpful to retard hyperosmolarity niche-mediated disc degeneration.

The NK1 receptor antagonist NKP608 inhibits proliferation of human colorectal cancer cells via Wnt signaling pathway.

BACKGROUND: Neurokinin1 (NK1) receptor has played a vital role in the development of tumor. However, NKP608 as a NK1 receptor antagonist whether has the effect of the resistance of colorectal cancer is still unclear. Thereby, in this study, we investigated the role of NKP608 on human colorectal cancer and explored the underlying mechanism. METHODS: The cell proliferation of colorectal cancer cells was detected by cell counting kit-8 (CCK8) assay, cell migration and invasion were assessed by transwell assay, the apoptotic ratio of cells was assessed by Annexin V-fluorescein isothiocyanate/propidium iodide stained and flow cytometry. The involvement of molecular mechanisms was examined by western blot. RESULTS: In this study, we found that NKP608 inhibited the proliferation, migration/invasion of HCT116 cells. In addition, NKP608 reduced expressions of Wnt-3a, ß-catenin, Cyclin D1, and (vascular endothelial growth factor) VEGF while induced expression of E-Cadherin. Furthermore, flow cytometry analyzed that NKP608 induced apoptosis of HCT116 cells, consistently, western blotting detecting of apoptosis-related proteins revealed that NKP608 downregulated Bcl-2 while upregulated Bax and Active-Caspase-3. CONCLUSIONS: Taken together, our results demonstrated that NKP608 inhibited colorectal cancer cell proliferation, migration and invasion via suppressing the Wnt/ß-catenin signaling pathway. Therefore, NKP608 might represent a promising therapeutic agent in the treatment of colorectal cancer.

The NK1 receptor antagonist NKP608 inhibits proliferation of human colorectal cancer cells via Wnt signaling pathway

BACKGROUND: Neurokinin1 (NK1) receptor has played a vital role in the development of tumor. However, NKP608 as a NK1 receptor antagonist whether has the effect of the resistance of colorectal cancer is still unclear. Thereby, in this study, we investigated the role of NKP608 on human colorectal cancer and explored the underlying mechanism. METHODS: The cell proliferation of colorectal cancer cells was detected by cell counting kit-8 (CCK8) assay, cell migration and invasion were assessed by transwell assay, the apoptotic ratio of cells was assessed by Annexin V-fluorescein isothiocyanate/propidium iodide stained and flow cytometry. The involvement of molecular mechanisms was examined by western blot. RESULTS: In this study, we found that NKP608 inhibited the proliferation, migration/invasion of HCT116 cells. In addition, NKP608 reduced expressions of Wnt-3a, ß-catenin, Cyclin D1, and (vascular endothelial growth factor) VEGF while induced expression of E-Cadherin. Furthermore, flow cytometry analyzed that NKP608 induced apoptosis of HCT116 cells, consistently, western blotting detecting of apoptosis-related proteins revealed that NKP608 downregulated Bcl-2 while upregulated Bax and Active-Caspase-3. CONCLUSIONS: Taken together, our results demonstrated that NKP608 inhibited colorectal cancer cell proliferation, migration and invasion via suppressing the Wnt/ß-catenin signaling pathway. Therefore, NKP608 might represent a promising therapeutic agent in the treatment of colorectal cancer.

MiR-200a and miR-200b target PTEN to regulate the endometrial cancer cell growth in vitro.

OBJECTIVE: To study whether miR-200a and miR-200b target PTEN gene expression to regulate the endometrial cancer cell growth in vitro. METHODS: Endometrial cancer cells ECC-1 were cultured and transfected with the miR-200a and miR-200b mimics and inhibitors as well as the negative control mimics and inhibitors, and then the cell proliferation activity as well as the expression of PTEN and downstream genes in cells was determined; after transfection of miR-200a and miR-200b mimics as well as PTEN-3'UTR luciferase report gene plasmids, the fluorescence activity of luciferase reporter gene was determined. RESULTS: 12 h, 24 h and 48 h after transfection, the cell proliferation activity of miR-200a mimics group and miR-200b mimics group were significantly higher than those of NC mimics group while the cell proliferation activity of miR-200a inhibitor group and miR-200b inhibitor group were significantly lower than those of NC inhibitor group; 48 h after transfection, PTEN expression in cells and PTEN-3'UTR luciferase reporter gene fluorescence activity of miR-200a mimics group and miR-200b mimics group were significantly lower than those of NC mimics group while p-PI3K and p-Akt expression were significantly higher than those of NC mimics group; PTEN expression in cells and PTEN-3'UTR luciferase reporter gene fluorescence activity of miR-200a inhibitor group and miR-200b inhibitor group were significantly higher than those of NC inhibitor group while p-PI3K and p-Akt expression were significantly lower than those of NC inhibitor group. CONCLUSION: miR-200a and miR-200b can promote the endometrial cancer cell growth in vitro by targeted inhibition of PTEN gene expression.

Effect of hypoxia on expression of placental trophoblast cells SATB1 and ß-catenin and its correlation with the pathogenesis of preeclampsia.

OBJECTIVE: To study the effect of hypoxia on the expression of placental trophoblast cells SATB1 and ß-catenin and its correlation with the pathogenesis of preeclampsia. METHODS: Trophoblastic cell lines HRT8/SVneo were cultured, SATB1 and ß-catenin expression and cell biological behavior were determined after hypoxia reoxygenation treatment; cell biological behavior and the expression of related genes were determined after the transfection of SATB1 and ß-catenin siRNA; preeclampsia placenta and normal placenta tissues were collected and the expression of SATB1 and ß-catenin were determined. RESULTS: OD value, cell migration rate, mRNA contents of SATB1 and ß-catenin of H/R group were significantly lower than those of Nor group, cell apoptosis rate was higher than that of Nor group and the number of invasive cells was less than that of Nor group; OD value and bcl-2 mRNA content of SATB1-siRNA group were lower than those of NC group; cell apoptosis rate as well as Bax, Caspase-3, Caspase-6 and Caspase-9 mRNA contents were higher than those of NC group; cell migration rate as well as CTSB, CTSD, MMP2 and MMP9 mRNA contents of ß-catenin-siRNA group were lower than those of NC group; the number of invasive cells was less than that of NC group; the expression levels of SATB1 and ß-catenin in preeclampsia placenta tissue were significantly lower than those in normal placenta tissue. CONCLUSIONS: Hypoxia can inhibit the expression of SATB1 and ß-catenin in the pathogenesis of preeclampsia, which can affect the proliferation, apoptosis, migration and invasion of cells.

Effect of the angiotensin II receptor blocker valsartan on cardiac hypertrophy and myocardial histone deacetylase expression in rats with aortic constriction.

The aim of the present study was to observe the myocardial expression of members of the histone deacetylase (HDAC) family (HDAC2, HDAC5 and HDAC9) in rats with or without myocardial hypertrophy (MH) in the presence and absence of the angiotensin II receptor blocker valsartan. Adult male Wistar rats were randomly divided into three groups (n=6/group): Sham-operated control rats, treated with distilled water (1 ml/day) through gavage; rats with MH (established through aortic constriction), treated with distilled water (1 ml/day) through gavage; and MH + valsartan rats, treated with 20 mg/kg/day valsartan through gavage. Treatments commenced one day after surgery and continued for eight weeks. Body weight (BW), heart weight (HW) and plasma atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) levels were determined, and the myocardial expression of HDAC2, HDAC5 and HDAC9 was analyzed through a reverse transcription semi-quantitative polymerase chain reaction. The BWs of the rats in the three groups were similar at baseline; however, after eight weeks the BW of the rats in the MH + valsartan group was significantly reduced compared with that of the MH rats. Furthermore, the HW/BW ratio and plasma ANP and BNP levels were increased, the myocardial HDAC2 expression was significantly upregulated and the HDAC5 and HDAC9 expression was significantly downregulated in the MH rats compared with those in the control rats; however, these changes were significantly attenuated by valsartan. Modulation of myocardial HDAC5, HDAC9 and HDAC2 expression may therefore be one of the anti-hypertrophic mechanisms of valsartan in this rat MH model.

A heterozygous moth genome provides insights into herbivory and detoxification.

How an insect evolves to become a successful herbivore is of profound biological and practical importance. Herbivores are often adapted to feed on a specific group of evolutionarily and biochemically related host plants, but the genetic and molecular bases for adaptation to plant defense compounds remain poorly understood. We report the first whole-genome sequence of a basal lepidopteran species, Plutella xylostella, which contains 18,071 protein-coding and 1,412 unique genes with an expansion of gene families associated with perception and the detoxification of plant defense compounds. A recent expansion of retrotransposons near detoxification-related genes and a wider system used in the metabolism of plant defense compounds are shown to also be involved in the development of insecticide resistance. This work shows the genetic and molecular bases for the evolutionary success of this worldwide herbivore and offers wider insights into insect adaptation to plant feeding, as well as opening avenues for more sustainable pest management.

Single-cell exome sequencing and monoclonal evolution of a JAK2-negative myeloproliferative neoplasm.

Tumor heterogeneity presents a challenge for inferring clonal evolution and driver gene identification. Here, we describe a method for analyzing the cancer genome at a single-cell nucleotide level. To perform our analyses, we first devised and validated a high-throughput whole-genome single-cell sequencing method using two lymphoblastoid cell line single cells. We then carried out whole-exome single-cell sequencing of 90 cells from a JAK2-negative myeloproliferative neoplasm patient. The sequencing data from 58 cells passed our quality control criteria, and these data indicated that this neoplasm represented a monoclonal evolution. We further identified essential thrombocythemia (ET)-related candidate mutations such as SESN2 and NTRK1, which may be involved in neoplasm progression. This pilot study allowed the initial characterization of the disease-related genetic architecture at the single-cell nucleotide level. Further, we established a single-cell sequencing method that opens the way for detailed analyses of a variety of tumor types, including those with high genetic complex between patients.

Single-cell exome sequencing reveals single-nucleotide mutation characteristics of a kidney tumor.

Clear cell renal cell carcinoma (ccRCC) is the most common kidney cancer and has very few mutations that are shared between different patients. To better understand the intratumoral genetics underlying mutations of ccRCC, we carried out single-cell exome sequencing on a ccRCC tumor and its adjacent kidney tissue. Our data indicate that this tumor was unlikely to have resulted from mutations in VHL and PBRM1. Quantitative population genetic analysis indicates that the tumor did not contain any significant clonal subpopulations and also showed that mutations that had different allele frequencies within the population also had different mutation spectrums. Analyses of these data allowed us to delineate a detailed intratumoral genetic landscape at a single-cell level. Our pilot study demonstrates that ccRCC may be more genetically complex than previously thought and provides information that can lead to new ways to investigate individual tumors, with the aim of developing more effective cellular targeted therapies.

Single-cell sequencing analysis characterizes common and cell-lineage-specific mutations in a muscle-invasive bladder cancer.

BACKGROUND: Cancers arise through an evolutionary process in which cell populations are subjected to selection; however, to date, the process of bladder cancer, which is one of the most common cancers in the world, remains unknown at a single-cell level. RESULTS: We carried out single-cell exome sequencing of 66 individual tumor cells from a muscle-invasive bladder transitional cell carcinoma (TCC). Analyses of the somatic mutant allele frequency spectrum and clonal structure revealed that the tumor cells were derived from a single ancestral cell, but that subsequent evolution occurred, leading to two distinct tumor cell subpopulations. By analyzing recurrently mutant genes in an additional cohort of 99 TCC tumors, we identified genes that might play roles in the maintenance of the ancestral clone and in the muscle-invasive capability of subclones of this bladder cancer, respectively. CONCLUSIONS: This work provides a new approach of investigating the genetic details of bladder tumoral changes at the single-cell level and a new method for assessing bladder cancer evolution at a cell-population level.

Resequencing 50 accessions of cultivated and wild rice yields markers for identifying agronomically important genes.

Rice is a staple crop that has undergone substantial phenotypic and physiological changes during domestication. Here we resequenced the genomes of 40 cultivated accessions selected from the major groups of rice and 10 accessions of their wild progenitors (Oryza rufipogon and Oryza nivara) to >15 × raw data coverage. We investigated genome-wide variation patterns in rice and obtained 6.5 million high-quality single nucleotide polymorphisms (SNPs) after excluding sites with missing data in any accession. Using these population SNP data, we identified thousands of genes with significantly lower diversity in cultivated but not wild rice, which represent candidate regions selected during domestication. Some of these variants are associated with important biological features, whereas others have yet to be functionally characterized. The molecular markers we have identified should be valuable for breeding and for identifying agronomically important genes in rice.

Frequent mutations of chromatin remodeling genes in transitional cell carcinoma of the bladder.

Transitional cell carcinoma (TCC) is the most common type of bladder cancer. Here we sequenced the exomes of nine individuals with TCC and screened all the somatically mutated genes in a prevalence set of 88 additional individuals with TCC with different tumor stages and grades. In our study, we discovered a variety of genes previously unknown to be mutated in TCC. Notably, we identified genetic aberrations of the chromatin remodeling genes (UTX, MLL-MLL3, CREBBP-EP300, NCOR1, ARID1A and CHD6) in 59% of our 97 subjects with TCC. Of these genes, we showed UTX to be altered substantially more frequently in tumors of low stages and grades, highlighting its potential role in the classification and diagnosis of bladder cancer. Our results provide an overview of the genetic basis of TCC and suggest that aberration of chromatin regulation might be a hallmark of bladder cancer.

Structural variation in two human genomes mapped at single-nucleotide resolution by whole genome de novo assembly.

Here we use whole-genome de novo assembly of second-generation sequencing reads to map structural variation (SV) in an Asian genome and an African genome. Our approach identifies small- and intermediate-size homozygous variants (1-50 kb) including insertions, deletions, inversions and their precise breakpoints, and in contrast to other methods, can resolve complex rearrangements. In total, we identified 277,243 SVs ranging in length from 1-23 kb. Validation using computational and experimental methods suggests that we achieve overall <6% false-positive rate and <10% false-negative rate in genomic regions that can be assembled, which outperforms other methods. Analysis of the SVs in the genomes of 106 individuals sequenced as part of the 1000 Genomes Project suggests that SVs account for a greater fraction of the diversity between individuals than do single-nucleotide polymorphisms (SNPs). These findings demonstrate that whole-genome de novo assembly is a feasible approach to deriving more comprehensive maps of genetic variation.