Genomic sequence variation is the hallmark of life and is key to understanding diversity and adaptation among the numerous microorganisms on earth. Analysis of the sequenced microbial genomes suggests that genes are evolving at many different rates. We have attempted to derive a new classification of genes into three broad categories: lineage-specific genes that evolve rapidly and appear unique to individual species or strains; highly conserved genes that frequently perform housekeeping functions; and partially variable genes that contain highly variable regions, at least 70 amino acids long, interspersed among well-conserved regions. The latter we term segmentally variable genes (SVGs), and we suggest that they are especially interesting targets for biochemical studies. Among these genes are ones necessary to deal with the environment, including genes involved in host-pathogen interactions, defense mechanisms, and intracellular responses to internal and environmental changes. For the most part, the detailed function of these variable regions remains unknown. We propose that they are likely to perform important binding functions responsible for protein-protein, protein-nucleic acid, or protein-small molecule interactions. Discerning their function and identifying their binding partners may offer biologists new insights into the basic mechanisms of adaptation, context-dependent evolution, and the interaction between microbes and their environment.

Deficiency or mutation in the p53 tumor suppressor gene commonly occurs in human cancer and can contribute to disease progression and chemotherapy resistance. Currently, although the pro-survival or pro-death effect of autophagy remains a controversial issue, increasing data seem to support the idea that autophagy facilitates cancer cell resistance to chemotherapy treatment. Here we report that 5-FU treatment causes aberrant autophagosome accumulation in HCT116 p53(-/-) and HT-29 cancer cells. Specific inhibition of autophagy by 3-MA, CQ or small interfering RNA treatment targeting Atg5 or Beclin 1 can potentiate the re-sensitization of these resistant cancer cells to 5-FU. In further analysis, we show that JNK activation and phosphorylation of Bcl-2 are key determinants in 5-FU-induced autophagy. Inhibition of JNK by the compound SP600125 or JNK siRNA suppressed autophagy and phosphorylation of c-Jun and Bcl-2 but increased 5-FU-induced apoptosis in both HCT116 p53(-/-) and HT29 cells. Taken together, our results suggest that JNK activation confers 5-FU resistance in HCT116 p53(-/-) and HT29 cells by promoting autophagy as a pro-survival effect, likely via inducing Bcl-2 phosphorylation. These results provide a promising strategy to improve the efficacy of 5-FU-based chemotherapy for colorectal cancer patients harboring a p53 gene mutation.

TGF-β secreted by tumor stroma induces epithelial-to-mesenchymal transition (EMT) in cancer cells, a reversible phenotype linked to cancer progression and drug resistance. However, exposure to stromal signals may also lead to heritable changes in cancer cells, which are poorly understood. We show that epithelial cells failing to undergo proliferation arrest during TGF-β-induced EMT sustain mitotic abnormalities due to failed cytokinesis, resulting in aneuploidy. This genomic instability is associated with the suppression of multiple nuclear envelope proteins implicated in mitotic regulation and is phenocopied by modulating the expression of LaminB1. While TGF-β-induced mitotic defects in proliferating cells are reversible upon its withdrawal, the acquired genomic abnormalities persist, leading to increased tumorigenic phenotypes. In metastatic breast cancer patients, increased mesenchymal marker expression within single circulating tumor cells is correlated with genomic instability. These observations identify a mechanism whereby microenvironment-derived signals trigger heritable genetic changes within cancer cells, contributing to tumor evolution.

Mutations in human zinc-finger transcription factor WT1 result in abnormal development of the kidneys and genitalia and an array of pediatric problems including nephropathy, blastoma, gonadal dysgenesis and genital discordance. Several overlapping phenotypes are associated with WT1 mutations, including Wilms tumors, Denys-Drash syndrome (DDS), Frasier syndrome (FS) and WAGR syndrome (Wilms tumor, aniridia, genitourinary malformations, and mental retardation). These conditions vary in severity from individual to individual; they can be fatal in early childhood, or relatively benign into adulthood. DDS mutations cluster predominantly in zinc fingers (ZF) 2 and 3 at the C-terminus of WT1, which together with ZF4 determine the sequence-specificity of DNA binding. We examined three DDS associated mutations in ZF2 of human WT1 where the normal glutamine at position 369 is replaced by arginine (Q369R), lysine (Q369K) or histidine (Q369H). These mutations alter the sequence-specificity of ZF2, we find, changing its affinity for certain bases and certain epigenetic forms of cytosine. X-ray crystallography of the DNA binding domains of normal WT1, Q369R and Q369H in complex with preferred sequences revealed the molecular interactions responsible for these affinity changes. DDS is inherited in an autosomal dominant fashion, implying a gain of function by mutant WT1 proteins. This gain, we speculate, might derive from the ability of the mutant proteins to sequester WT1 into unproductive oligomers, or to erroneously bind to variant target sequences.

Externally controlled site specific drug delivery could potentially provide a means of reducing drug related side effects whilst maintaining, or perhaps increasing therapeutic efficiency. The aim of this work was to develop a nanoscale drug carrier, which could be loaded with an anti-cancer drug and be directed by an external magnetic field. Using a single, commercially available monomer and a simple one-pot reaction process, a polymer was synthesized and crosslinked within the pores of an anodized aluminum oxide template. These polymer nanotubes (PNT) could be functionalized with iron oxide nanoparticles for magnetic manipulation, without affecting the large internal pore, or inherent low toxicity. Using an external magnetic field the nanotubes could be regionally concentrated, leaving areas devoid of nanotubes. Lastly, doxorubicin could be loaded to the PNTs, causing increased toxicity towards neuroblastoma cells, rendering a platform technology now ready for adaptation with different nanoparticles, degradable pre-polymers, and various therapeutics.

This study investigated the clinical significance of serum soluble Toll-like receptor 4 (sTLR4) in non-small cell lung cancer (NSCLC). A total of 54 NSCLC patients and 13 healthy volunteers were enrolled from January 2012 to December 2013. The patients with NSCLC were characterized by significantly higher serum levels of sTLR4 compared with those in healthy controls (P < 0.01). A positive correlation between serum sTLR4 and tumor stage was found in patients with stages I-III NSCLC. However, serum sTLR4 in patients with metastatic NSCLC was significantly decreased compared with those with stage III NSCLC (P < 0.05). Furthermore, low serum sTLR4 was identified as a prognostic marker for poor survival of early-stage NSCLC patients who received surgical resection. In conclusion, our present study identified sTLR4 as a potential serum biomarker of NSCLC.

Immune checkpoints associate with dysfunctional T cells, which have a reduced ability to clear pathogens or cancer cells. T-cell checkpoint blockade may improve patient survival. However, checkpoint molecules on cytokine-induced killer (CIK) cell, a non-specific adoptive immunotherapy, remain unknown. In present study, we detected the dynamic expression of eight major checkpoint molecules (CTLA-4, PD-1, PD-L1, TIM- 3, CEACAM-1, LAG-3, TIGIT and BTLA) on CIK cells from NSCLC patients. The majority of these molecules, except BTLA, were sharply elevated during the early stage of CIK cell culture. Thereafter, PD-1 and TIGIT expressions decreased gradually towards the initial level (day 0). Moreover, CTLA-4 faded away during the later stage of CIK culture. LAG-3 expression decreased but was still significantly higher than the initial level. Of note, PD-L1 remained stably upregulated during CIK culture compared with PD-1, indicating that PD-L1 might act as an inhibitory molecule on CIK cells instead of PD-1. Furthermore, TIM-3 and CEACAM1 were strongly expressed simultaneously during long-term CIK culture and showed a significant and mutually positive correlation. BTLA displayed a distinct pattern, and its expression gradually decreased throughout the CIK culture. These observations suggested that CIK cells might be partly exhausted before clinical transfusion, characterized by the high expression of PD-L1, LAG-3, TIM- 3, and CEACAM-1 and the low expression of TIGIT, BTLA, PD-1, and CTLA-4 compared with initial culture. Our results imply that implementing combined treatment on CIK cells before transfusion via antibodies targeting PD-L1, LAG-3, TIM-3, and CEACAM-1 might improve the efficiency of CIK therapy for NSCLC patients.

The MspJI modification-dependent restriction endonuclease recognizes 5-methylcytosine or 5-hydroxymethylcytosine in the context of CNN(G/A) and cleaves both strands at fixed distances (N(12)/N(16)) away from the modified cytosine at the 3'-side. We determined the crystal structure of MspJI of Mycobacterium sp. JLS at 2.05-Å resolution. Each protein monomer harbors two domains: an N-terminal DNA-binding domain and a C-terminal endonuclease. The N-terminal domain is structurally similar to that of the eukaryotic SET and RING-associated domain, which is known to bind to a hemi-methylated CpG dinucleotide. Four protein monomers are found in the crystallographic asymmetric unit. Analytical gel-filtration and ultracentrifugation measurements confirm that the protein exists as a tetramer in solution. Two monomers form a back-to-back dimer mediated by their C-terminal endonuclease domains. Two back-to-back dimers interact to generate a tetramer with two double-stranded DNA cleavage modules. Each cleavage module contains two active sites facing each other, enabling double-strand DNA cuts. Biochemical, mutagenesis and structural characterization suggest three different monomers of the tetramer may be involved respectively in binding the modified cytosine, making the first proximal N(12) cleavage in the same strand and then the second distal N(16) cleavage in the opposite strand. Both cleavage events require binding of at least a second recognition site either in cis or in trans.

In current studies, the influence of tumor immune microenvironment on tumorigenesis and tumor progression has been widely explored. In the present study, we investigated the expression and significance of high mobility group box 1 (HMGB1), HMG nucleosome-binding protein 1 (HMGN1), the receptor programmed cell death 1 (PD-1) and its ligand programmed cell death ligand 1 (PD-L1) in head and neck squamous cell carcinoma (HNSCC). We explored whether HMGB1 and HMGN1 take part in recruiting T cells to HNSCC microenvironment. Furthermore, we assessed the prognostic value of HMG proteins, TILs, and PD-1/PD-L1 in postoperative patients. Tumor tissue sections were collected from 81 cases of patients with resectable HNSCC. All patients' information was integrated with clinical and pathological records, as well as follow-up data. We used immunohistochemistry to examine the subcellular localization and expression levels of HMGB1 and HMGN1, as well as tumor CD3+, CD8+, FOXP3+ lymphocyte infiltration, and the expression of immune inhibiting molecules PD-1/PD-L1. Results showed that there was no significant difference in the number of CD8+ and FOXP3+ T cells between the two groups with or without HMGB1 cytoplasmic expression in tumor tissues. The number of CD3+ T cells in HMGB1 cytoplasmic expression group (339.39 ± 230.76) was more than that in group without HMGB1 cytoplasmic expression (233.30 ± 230.91, P < 0.05). The number of CD3+, CD8+, and FOXP3+ T cells in HMGN1 cytoplasmic expression group [400.74 ± 224.04, 158.10 ± 112.10, 36.00(15.00, 69.00)] was more than that in group without HMGN1-cytoplasmic expression [222.84 ± 217.78, P < 0.01; 105.10 ± 108.25, P < 0.05; 13.00(6.75, 32.25), P < 0.01]. The positive rates of PD-1 and PD-L1 in tumor tissues were 29.6 and 67.9%, respectively. Multivariate analysis suggested that tumor expression of PD-L1 was an independent prognostic factor and PD-L1 overexpression indicated a poor overall survival (OS) and disease-free survival (DFS). Taken together, we concluded that HMGB1 and HMGN1 secreted by cancer cells may relate to recruitment of tumor infiltrating lymphocytes (TILs) in HNSCC. PD-1/PD-L1 axis, rather than HMG proteins or CD8+ tumor-infiltrating lymphocytes, has a critical role in tumor immune microenvironment and could predict the outcome of HNSCC patients who received surgical resection.

Background: As a major subtype of ovarian cancer, high grade FIGO stage IIIc serous ovarian carcinoma (HG3cSOC), has various prognosis due to genetic heterogeneity. Methods: The transcriptome of 401 primary FIGO IIIc serous ovarian samples was screened, seven genes based prognostic model was developed. The prognostic valueof risk score in four different cohorts (TCGA-cohort, Poland-cohort, Japan-cohort and USA-cohort) was validated. The relationship between risk score and other clinical indicators were analyzed. The guide value of risk score for platinum-taxol chemotherapy was also assayed. Tissue microenvironment difference among samples with different risk scores was investigated. Results: High-risk group (N=200, median survival months: 39.6, 95% CI: 35.9-46.3 months) had a significantly worse prognosis than low-risk group (N=201, median survival months: 52.6, 95% CI: 45.2-64.9 months;). The risk score's performance was validated in Japan-cohort (N=90, Poland-cohort (N=48) and USA-cohort (N=84). The risk score is independent from age, primary tumor size, grade and treatment methods and the performance of risk score is uniform in subgroups. Furthermore, the risk score predicted the response of HG3cSOC to platinum-based regimen after surgery, and this finding was further validated in newly collected China-cohort (N=102). Gene Set Enrichment Analysis (GSEA) and tumor infiltration analysis revealed that risk score reflected the immune infiltration and cell-cell interaction status, and the migration function of candidate genes were also verified. Conclusions: The optimized seven genes-based model is a valuable and robust model in predicting the survival of HG3cSOC, and served as a valuable marker for the response to platinum-based chemotherapy.

Vitamin D co-regulates cell proliferation, differentiation and apoptosis in numerous tissues, including cancers. The known anti-proliferative and pro-apoptotic actions of the active metabolite of vitamin D, 1,25-dihydroxy-vitamin D [1,25(OH)2D] are mediated through binding to the vitamin D receptor (VDR). Here, we report on the unexpected finding that stable knockdown of VDR expression in the human breast and prostate cancer cell lines, MDA-MB-231 and PC3, strongly induces cell apoptosis and inhibits cell proliferation in vitro. Implantation of these VDR knockdown cells into the mammary fat pad (MDA-MB-231), subcutaneously (PC3) or intra-tibially (both cell lines) in immune-incompetent nude mice resulted in reduced tumor growth associated with increased apoptosis and reduced cell proliferation compared with controls. These growth-retarding effects of VDR knockdown occur in the presence and absence of vitamin D and are independent of whether cells were grown in bone or soft tissues. Transcriptome analysis of VDR knockdown and non-target control cell lines demonstrated that loss of the VDR was associated with significant attenuation in the Wnt/β-catenin signaling pathway. In particular, cytoplasmic and nuclear β-catenin protein levels were reduced with a corresponding downregulation of downstream genes such as Axin2, Cyclin D1, interleukin-6 (IL-6), and IL-8. Stabilization of β-catenin using the GSK-3β inhibitor BIO partly reversed the growth-retarding effects of VDR knockdown. Our results indicate that the unliganded VDR possesses hitherto unknown functions to promote breast and prostate cancer growth, which appear to be operational not only within but also outside the bone environment. These novel functions contrast with the known anti-proliferative nuclear actions of the liganded VDR and may represent targets for new diagnostic and therapeutic approaches in breast and prostate cancer.

Acetic acid bacteria (AAB) are widely applied in food, bioengineering and medicine fields. However, the acid stress at low pH conditions limits acetic acid fermentation efficiency and high concentration of vinegar production with AAB. Therefore, how to enhance resistance ability of the AAB remains as the major challenge. Amino acids play an important role in cell growth and cell survival under severe environment. However, until now the effects of amino acids on acetic fermentation and acid stress resistance of AAB have not been fully studied.

Helminths and their products can shape immune responses by modulating immune cells, which are dysfunctional in inflammatory diseases such as asthma. We previously identified SJMHE1, a small molecule peptide from the HSP60 protein of Schistosoma japonicum. SJMHE1 can inhibit delayed-type hypersensitivity and collagen-induced arthritis in mice. In the present study, we evaluated this peptide's potential intervention effect and mechanism on ovalbumin-induced asthma in mice. SJMHE1 treatment suppressed airway inflammation in allergic mice, decreased the infiltrating inflammatory cells in the lungs and bronchoalveolar lavage fluid, modulated the production of pro-inflammatory and anti-inflammatory cytokines in the splenocytes and lungs of allergic mice, reduced the percentage of Th2 cells and increased the proportion of Th1 and regulatory T cells (Tregs). At the same time, Foxp3 and T-bet expression increased, and GATA3 and RORγt decreased in the lungs of allergic mice. We proved that SJMHE1 can interrupt the development of asthma by diminishing airway inflammation in mice. The down-regulation of Th2 response and the up-regulation of Th1 and Tregs response may contribute to the protection induced by SJMHE1 in allergic mice. SJMHE1 can serve as a novel therapy for asthma and other allergic or inflammatory diseases.

Cowpea [Vigna unguiculata (L.) Walp.] is an annual legume of economic importance and widely grown in the semi-arid tropics. However, high-density genetic maps of cowpea are still lacking. Here, we identified 34,868 SNPs (single nucleotide polymorphisms) that were distributed in the cowpea genome based on the RAD sequencing (restriction-site associated DNA sequencing) technique using a population of 170 individuals (two cowpea parents and 168 F2:3 progenies). Of these, 17,996 reliable SNPs were allotted to 11 consensus linkage groups (LGs). The length of the genetic map was 1,194.25 cM in total with a mean distance of 0.066 cM/SNP marker locus. Using this map and the F2:3 population, combined with the CIM (composite interval mapping) method, eleven quantitative trait loci (QTL) of yield-related trait were detected on seven LGs (LG4, 5, 6, 7, 9, 10, and 11) in cowpea. These QTL explained 0.05-17.32% of the total phenotypic variation. Among these, four QTL were for pod length, four QTL for thousand-grain weight (TGW), two QTL for grain number per pod, and one QTL for carpopodium length. Our results will provide a foundation for understanding genes related to grain yield in the cowpea and genus Vigna.

Metastasis-competent circulating tumour cells (CTCs) experience oxidative stress in the bloodstream, but their survival mechanisms are not well defined. Here, comparing single-cell RNA-Seq profiles of CTCs from breast, prostate and lung cancers, we observe consistent induction of β-globin (HBB), but not its partner α-globin (HBA). The tumour-specific origin of HBB is confirmed by sequence polymorphisms within human xenograft-derived CTCs in mouse models. Increased intracellular reactive oxygen species (ROS) in cultured breast CTCs triggers HBB induction, mediated through the transcriptional regulator KLF4. Depletion of HBB in CTC-derived cultures has minimal effects on primary tumour growth, but it greatly increases apoptosis following ROS exposure, and dramatically reduces CTC-derived lung metastases. These effects are reversed by the anti-oxidant N-Acetyl Cysteine. Conversely, overexpression of HBB is sufficient to suppress intracellular ROS within CTCs. Altogether, these observations suggest that β-globin is selectively deregulated in cancer cells, mediating a cytoprotective effect during blood-borne metastasis.

The whitefly Bemisia tabaci (Gennadius) is an important agricultural insect pest worldwide. The B and Q biotypes are the two most predominant and devastating biotypes prevalent across China. However, there are few studies regarding the occurrence of the Q biotype in Fujian Province, China, where high insecticide resistance has been reported in the B biotype. Differences in some biological characteristics between the B and Q biotypes, especially insecticide resistance, are considered to affect the outcome of their competition. Extensive surveys in Fujian revealed that the B biotype was predominant during 2005-2014, whereas the Q biotype was first detected in some locations in 2013 and widely detected throughout the province in 2014. Resistance to neonicotinoids (that have been used for more than 10 years) exhibited fluctuations in open fields, but showed a continual increasing trend in protected areas. Resistance to lambda-cyhalothrin, chlorpyrifos, and abamectin exhibited a declining trend. Resistance to novel insecticides, such as nitenpyram, pymetrozine, sulfoxaflor, and cyantraniliprole, in 2014 was generally below a moderate level. A decline in insecticide resistance in the B biotype and the rapid buildup of protected crops under global temperature increase may have promoted the establishment of the Q biotype in Fujian.

Gluconeogenesis, an essential metabolic process for hepatocytes, is downregulated in hepatocellular carcinoma (HCC). Here we show that the nuclear receptor Nur77 is a tumour suppressor for HCC that regulates gluconeogenesis. Low Nur77 expression in clinical HCC samples correlates with poor prognosis, and a Nur77 deficiency in mice promotes HCC development. Nur77 interacts with phosphoenolpyruvate carboxykinase (PEPCK1), the rate-limiting enzyme in gluconeogenesis, to increase gluconeogenesis and suppress glycolysis, resulting in ATP depletion and cell growth arrest. However, PEPCK1 becomes labile after sumoylation and is degraded via ubiquitination, which is augmented by the p300 acetylation of ubiquitin-conjugating enzyme 9 (Ubc9). Although Nur77 attenuates sumoylation and stabilizes PEPCK1 via impairing p300 activity and preventing the Ubc9-PEPCK1 interaction, Nur77 is silenced in HCC samples due to Snail-mediated DNA methylation of the Nur77 promoter. Our study reveals a unique mechanism to suppress HCC by switching from glycolysis to gluconeogenesis through Nur77 antagonism of PEPCK1 degradation.

The incidence of multiple primary malignant tumors (MPMTs) is rising due to the development of screening technologies, significant treatment advances and increased aging of the population. For patients with a prior cancer history, identifying the tumor origin of the second malignant lesion has important prognostic and therapeutic implications and still represents a difficult problem in clinical practice.

Study on the mechanism of high-frequency stimulation inhibiting low-Mg2+-induced epileptiform discharges in juvenile rat hippocampal slices High-frequency stimulation (HFS) has been demonstrated to be an effective treatment for inhibiting epilepsy in some clinical and laboratory studies. However, the mechanisms underlying the therapeutic effects of HFS are not yet fully understood. In our present study, epileptiform discharges (EDs) in acutely isolated hippocampal slices of male Sprague-Dawley (SD) juvenile rats induced by low-Mg2+ artificial cerebrospinal fluid (ACSF), and electrical stimulation (square wave, 900 pulses, 50 % duty-cycle, 130 Hz) was performed on the CA3 using concentric bipolar electrodes. EDs of neurons in hippocampal were recorded by multi-electrode arrays (MEA). After stable EDs events had been recorded for at least 20 min, HFS was added, followed by 10 μmol/L gamma-aminobutyric acid type A (GABAA) receptors blocker bicuculline (BIC). The results show that the HFS can increase the discharges frequency of inter-ictal discharges (IIDs) and decrease the duration of ictal discharges (IDs). However, the HFS had no effect on the slices with 10 μmol/L BIC. These results indicated that the GABAA receptors are activated when HFS inhibited EDs, thereby achieving the inhibition of low-Mg2+-induced EDs in slices.

Tumour-induced osteomalacia (TIO) is a very rare paraneoplastic syndrome with bone pain, fractures and muscle weakness, which is mostly caused by phosphaturic mesenchymal tumours (PMTs). Cell-free DNA (cfDNA) has been regarded as a non-invasive liquid biopsy for many malignant tumours. However, it has not been studied in benign tumours, which prompted us to adopt the targeted next-generation sequencing approach to compare cfDNAs of 4 TIO patients, four patients with bone metastasis (BM) and 10 healthy controls. The mutational landscapes of cfDNA in TIO and BM groups were similar in the spectrum of allele frequencies and mutation types. Markedly, deleterious missense mutations in FGFR1 and loss-of-function mutations in MED12 were found in 3/4 TIO patients but none of BM patients. The gene ontology analysis strongly supported that these mutated genes found in TIOs would play a potential role in PMTs' process. The genetic signatures and corresponding change in expression of FGFR1 and FGF23 were further validated in PMT tissues from a test cohort of another three TIO patients. In summary, we reported the first study of the mutational landscape and genetic signatures of cfDNA in TIO/PMTs.