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Cytokinins are a class of phytohormones that promote cell division and differentiation and are thought to affect plant immunity to multiple pathogens. However, a comprehensive analysis of cytokinin dehydrogenase/oxidase (CKX) family genes in cabbage has not been reported. In this study, a total of 36 CKX genes were identified using a genome-wide search method. Phylogenetic analysis classified these genes into three groups. They were distributed unevenly across nine chromosomes in B. oleracea, and 15 of them did not contain any introns. The results of colinearity analysis showed that 36 CKX gene in Arabidopsis was present in several copies in the Brassica oleracea genome. An analysis of cis-acting elements indicated that all genes possessed at least one stress or hormone responsive cis-acting element. A heatmap of CKX gene expression showed the patterns of expression of these genes in various tissues and organs. Three genes (Bol028363, Bol031036 and Bol018140) were relatively highly expressed in all of the investigated tissues under normal conditions, showing the expression profile of housekeeping genes. Generally, the expression patterns of CKX genes in Jingfeng 1 and Xiangan 336 were quite different under the same treatment. Notably, three genes (Bol020547, Bol028392 and Bol045724) were significantly down-regulated and up-regulated in the susceptible and resistant material, respectively, after inoculation, which may indicate their crucial roles in resistance to clubroot disease. The results provide insights for better understanding the roles of CKX genes in the B. oleracea–P. brassicae interaction.

Erythrokeratodermia variabilis et progressiva (EKVP) is a rare genodermatosis of clinical and genetic heterogeneity, characterized by the manifestations of localized or disseminated persistent hyperkeratotic plagues and stationary to migratory transient erythematous patches. The majority of EKVP cases display an autosomal dominant mode of inheritance with incomplete penetrance, although recessive transmission has also been described. Mutations associated with EKVP have been primarily detected in connexin (Cx) genes. We herein reported a Chinese sporadic case of late-onset EKVP with a novel heterozygous missense mutation c.109G>A (p.V37M) in GJB4 (Cx30.3) gene, which resulted in a significant reduction of GJB4 expression in the epidermis of the patient. In accordance, while wild-type GJB4 localized at the cell membrane of HeLa cells forming intercellular junctions and intracellular puncta, V37M mutant variant was diffusely expressed within HeLa cells at a considerably lower level. Our findings reveal an essential role of GJB4 in the pathogenesis of EKVP and provides insights into the therapeutic potential of the disease.

Most aldosterone-producing adenomas (APAs) have gain-of-function somatic mutations of ion channels or transporters. However, their frequency in aldosterone-producing cell clusters of normal adrenal gland suggests a requirement for codriver mutations in APAs. Here we identified gain-of-function mutations in both CTNNB1 and GNA11 by whole-exome sequencing of 3/41 APAs. Further sequencing of known CTNNB1-mutant APAs led to a total of 16 of 27 (59%) with a somatic p.Gln209His, p.Gln209Pro or p.Gln209Leu mutation of GNA11 or GNAQ. Solitary GNA11 mutations were found in hyperplastic zona glomerulosa adjacent to double-mutant APAs. Nine of ten patients in our UK/Irish cohort presented in puberty, pregnancy or menopause. Among multiple transcripts upregulated more than tenfold in double-mutant APAs was LHCGR, the receptor for luteinizing or pregnancy hormone (human chorionic gonadotropin). Transfections of adrenocortical cells demonstrated additive effects of GNA11 and CTNNB1 mutations on aldosterone secretion and expression of genes upregulated in double-mutant APAs. In adrenal cortex, GNA11/Q mutations appear clinically silent without a codriver mutation of CTNNB1. Sequence analysis identifies gain-of-function somatic mutations in GNA11 or GNAQ in CTNNB1-mutant aldosterone-producing adenomas. Most patients with these mutations presented during puberty, pregnancy or menopause, with elevated LHCGR expression.

MDM4 is a homologue of MDM2, serving cooperatively as the negative regulator of tumor suppressor p53. Under the shadow of MDM2 inhibitors, limited efforts had been put into the discovery of MDM4 modulators. Recent studies of the experimental drug ALRN-6924, a dual MDM4 and MDM2 inhibitor, suggest that concurrent inhibition of MDM4 and MDM2 might be beneficial over only MDM2 inhibition. In view of the present research progress, we summarized published inhibitors of MDM4/p53 interactions including both peptide-based compounds and small molecules. Cocrystal structures of ligand/MDM4 complexes have been examined, and their structural features were compiled and compared in order to show the molecular basis required for high MDM4 binding affinities. Representative examples of small-molecule MDM4 inhibitors were discussed, followed by clinical results of ALRN-6924, together, providing a consolidated reference for further development of MDM4 inhibitors, either dual or selective.

Background Increased aerobic glycolysis has been well-known as a hallmark of cancer, which is closely related to mitochondrial dysfunction. TFB2M (mitochondrial transcription factor B2) is a core mitochondrial transcription factor, which has been shown by us to play an oncogenic role in hepatocellular carcinoma (HCC). However, whether TFB2M contributes to the aerobic glycolysis in HCC cells remains unexplored. Methods The role and underlying molecular mechanisms of TFB2M in the regulation of aerobic glycolysis in HCC cells were systematically investigated by in vitro cell glucose metabolism and metabolomics analyses. Besides, the effects of TFB2M-regulated aerobic glycolysis in the growth and metastasis of HCC cells were also explored. Results Here, we show that TFB2M markedly enhanced the reprogramming of glucose metabolism from oxidative phosphorylation to aerobic glycolysis mainly through two mechanisms. On the one hand, TFB2M increased the expressions of glycolytic genes GAPDH, LDHA, GLUT1 and HK2. On the other hand, TFB2M decreased the expression of peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α), a critical regulator of mitochondrial respiration. Mechanistically, TFB2M regulates the up-regulation of glycolytic genes and down-regulation of PGC-1α mainly through NAD+ /SIRT3/HIF-1α signaling. Additionally, we found that TFBM2 promoted the progression of HCC cells through HIF-1α-regulated reprogramming of glucose metabolism. Conclusions Our findings indicate that TFB2M serves as a critical glucose metabolic reprogramming mechanism in tumorigenesis, which could be used as potential therapeutic target in HCC.

PURPOSE: TUBB8 is a gene that is frequently analysed in the genetic diagnosis of female infertility; 102 variants of this gene have been identified. However, the evaluation of its pathogenicity and the resulting phenotypes vary. Here, we aimed to identify novel TUBB8 variants as well as to summarize the reported variants and phenotypes in order for them to be included in genetic counselling analyses. METHODS: We performed whole exome sequencing to screen for candidate variants in 100 infertile female subjects and 100 controls who were able to conceive naturally. All variants were confirmed by Sanger sequencing. The effects of the variants in oocytes/arrested embryos were assessed by morphological observations, polar body biopsies, and chromosome analysis. A molecular modelling analysis was used to evaluate the possible effects of variants on protein secondary structure. RESULTS: We identified 29 TUBB8 variants, of which 20 were novel and five were maternally inherited. We identified three of a total of six recurrent variants that were specific for complete cleavage failure. Moreover, we obtained evidence that TUBB8 variants with large polar bodies had chromosome segregation errors. CONCLUSIONS: Our study expands the spectrum of TUBB8 variants, particularly for embryonic arrest. Together with the extant knowledge of TUBB8 variants, this study provides a foundation for the genetic counselling of female infertility. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10815-021-02219-9.

Elongated hypocotyl5 (HY5) is a key transcription factor that promotes photomorphogenesis. Constitutive photomorphogenic1 (COP1)–Suppressor of phytochrome A-105 (SPA) E3 ubiquitin ligase complex promotes ubiquitination and degradation of HY5 to repress photomorphogenesis in darkness. HY5 is also regulated by phosphorylation at serine 36 residue. However, the kinase responsible for phosphorylation of HY5 remains unknown. Here, using extensive in vitro and in vivo biochemical, genetic, and photobiological techniques, we have identified a new kinase that phosphorylates HY5 and demonstrated the significance of phosphorylation of HY5 in Arabidopsis thaliana. We show that SPA proteins are the missing kinases necessary for HY5 phosphorylation. SPAs can directly phosphorylate HY5 in vitro, and the phosphorylated HY5 is absent in the spaQ background in vivo. We also demonstrate that the unphosphorylated HY5 interacts strongly with both COP1 and SPA1 and is the preferred substrate for degradation, whereas the phosphorylated HY5 is more stable in the dark. In addition, the unphosphorylated HY5 actively binds to the target promoters and is the physiologically more active form. Consistently, the transgenic plants expressing the unphosphorylated form of HY5 display enhanced photomorphogenesis. Collectively, our study revealed the missing kinase responsible for direct phosphorylation of HY5 that fine-tunes its stability and activity to regulate photomorphogenesis.

The partial response of chronic hepatitis B virus (CHB) patients to interferon-α (IFN-α) therapy remains elusive, which requires a better understanding of the involved molecular mechanism. In our study, bioinformatics analysis was applied to relate IFN-α regulated candidate genes and RNA editing sites by RNA sequencing. Mitochondrial antiviral signaling protein (MAVS) antiviral effect was confirmed in HepG2.2.15 cells and in two mouse models. The associations between polymorphisms in MAVS gene and response to IFN-α therapy were confirmed in CHB patients. We found that IFN-α downregulates MAVS via RNA editing that was mediated by adenosine deaminase acting on RNA (ADAR1). ADAR1 inhibited MAVS expression via a human antigen R (HuR)-mediated post-transcriptional regulation. MAVS exerted an antiviral activity and reduced the level of hepatitis B virus (HBV) markers in vitro and in vivo. IFN-α antiviral effects were significantly enhanced by MAVS co-transfection. Hepatitis B core protein (HBc) interacted with SP1 to inhibit the promoter activity of MAVS that regulates its expression. CHB patients with a rs3746662A allele had higher MAVS expression and thus were more responsive to IFN-α treatment. In this work, we demonstrated that the decrease of MAVS expression is mediated by the IFN-α-ADAR1 axis. This study also highlighted the potential for the clinical application of MAVS in combination with IFN-α for the treatment of HBV infection.

Stress can induce microspores to change their developmental pathway from the gametophytic to the embryogenic pathway. To explore the molecular mechanisms of microspore embryogenesis, complement DNA-amplified fragment length polymorphism was used to isolate the transcript-derived fragments during microspore embryogenesis of the non-heading Brassica campestris L. ssp. chinensis ‘Wuyueman’. With 256 primer combinations screened, a total of 94 transcript-derived fragments were identified, and 15 were successfully sequenced. Based on a BLAST search in the Brassica database, 12 of the 15 sequenced transcript-derived fragments were homologous to genes with annotations; the remaining three transcript-derived fragments did not match any sequences. Transcript-derived fragments with annotations were involved in cell wall formation, hormones, and resistance. Analysis of cis-elements indicated that there were heat shock-related and stress-related cis-elements in the promoter sequences of 12 transcript-derived fragments. TDF1(Bra040720), TDF6(Bra013664), and TDF15(Bra022587) were selected for validation of complement DNA-amplified fragment length polymorphism expression patterns by real-time quantitative PCR. Results confirmed the altered expression patterns of three genes revealed by the complement DNA-amplified fragment length polymorphism. This study provides novel information on the molecular mechanism of microspore embryogenesis in non-heading Chinese cabbage.

Debris flows are abrupt geological hazards frequently resulting in destructive disasters in mountain areas. Thus, analysis of the acceptable risk of debris-flow disasters should be attentively considered, whereas the crucial problem of the acceptable disaster risk is how to know what the risk levels of debris-flow disasters are acceptable for the public. Thence, the questionnaire investigation of the acceptable risk preferences for different populations in various regions can be such a solution. The acceptable disaster risks are not only taking into account people's acceptance, but also related to government's disaster-reduction strategy and also referred to expert’s professional evaluation. Therefore, the establishment of guidelines and criterions for the acceptable risks of debris-flow disasters need all participations from multiple populations and different regions. In this survey, three-category populations including ordinary people, experts/scholars and administrators across China joined the questionnaire investigation. Five indicators (disaster-affected toll, death toll, death rate, personal direct economic loss, and total direct economic loss) were used to describe the acceptable risks of debris-flow disasters. Incorporated with frequency analysis, this work has explained the demographic characteristics for the acceptable risk preferences. The results indicate that ordinary people have relatively low acceptable risks, experts/scholars have slightly high acceptable risks and administrators have acceptable risks between both of them. Educational level, occupation, and income were the dominatingly demographic factors for the interpretations of the acceptable risk preferences.

The prevalence of drug-resistant Mycobacterium tuberculosis (Mtb) strains makes disease control more complicated, which is the main cause of death in tuberculosis (TB) patients. Early detection and timely standard treatment are the key to current prevention and control of drug-resistant TB. In recent years, despite the continuous advancement in drug-resistant TB diagnostic technology, the needs for clinical rapid and accurate diagnosis are still not fully met. With the development of sequencing technology, the research of human microecology has been intensified. This study aims to use 16 rRNA sequencing technology to detect and analyze upper respiratory flora of TB patients with anti-TB drug sensitivity (DS, n = 55), monoresistance isoniazide (MR-INH, n = 33), monoresistance rifampin (MR-RFP, n = 12), multidrug resistance (MDR, n = 26) and polyresistance (PR, n = 39) in southern China. Potential microbial diagnostic markers for different types of TB drug resistance are searched by screening differential flora, which provides certain guiding significance for drug resistance diagnosis and clinical drug use of TB. The results showed that the pulmonary microenvironment of TB patients was more susceptible to infection by external pathogens, and the infection of different drug-resistant Mtb leads to changes in different flora. Importantly, seven novel microorganisms (Leptotrichia, Granulicatella, Campylobacter, Delfitia, Kingella, Chlamydophila, Bordetella) were identified by 16S rRNA sequencing as diagnostic markers for different drug resistance types of TB. Leptotrichia, Granulicatella, Campylobacter were potential diagnostic marker for TB patients with INH single-resistance. Delftia was a potential diagnostic marker for TB patients with RFP single drug-resistance. Kingella and Chlamydophila can be used as diagnostic markers for TB patients with PR. Bordetella can be used as a potential diagnostic marker for identification of TB patients with MDR.

Background Ectopic expression of transcription elongation factor A (SII)-like 7 (TCEAL7) has been observed in several kinds of cancers, but its role in melanoma is still unclear. This study was carried out to investigate TCEAL7 role in melanoma progression, and uncover the underlying mechanisms. Methods TCEAL7 expression levels in melanoma tissues and cells were determined by using real-time quantitative PCR (RT-PCR) and western blotting. CCK-8, transwell chambers, flow cytometry, starch assay and tumorigenesis assay were applied to detect cell growth, invasion, apoptosis, migration and tumorigenesis, respectively. Results A low expression level of TCEAL7 was observed in melanoma tissues and cells, which was associated with malignant clinical process and poor prognosis. TCEAL7 negatively modulated AKT1, AKT2, c-Myc, N-cadherin and PCNA expression and inhibited cancer progression via decreasing AKT1 and c-Myc levels. In addition, TCEAL7 was negatively modulated by miR-758-3p which promoted melanoma progression. Moreover, overexpression of TCEAL7 abolished miR-758-3p role in promoting melanoma progression. Conclusion This study demonstrated that TCEAL7, regulated by miR-758-3p inhibited melanoma progression through decreasing the expression levels of c-Myc and AKT1.

Lysine crotonylation (Kcrot), as a post-translational modification (PTM) originally identified at histone proteins, is involved in diverse biological processes. Several conventional machine-learning (ML) predictors were developed based on the Kcrot sites from histone proteins. Recently, thousands of Kcrot sites have been experimentally verified on non-histone proteins from multiple species. Accordingly, a few predictors have been developed for predicting the Krot sites for specific organisms (i.e. humans and papaya). Nevertheless, there is a lack of research on the comparison of the crotonylomes of different organisms. Here, we collected around 20,000 Kcrot sites experimentally identified from four different species as the benchmark data set. We present the deep-learning (DL) architecture dubbed DeepKcrot for predicting Kcrot sites on the proteomes across various species. DeepKcrot includes species-specific and general classifiers using a convolutional neural network with the word embedding (CNNWE) encoding approach. CNNWE performs better than both the traditional ML-based and other DL-based classifiers in terms of ten-fold cross-validation and independent test, independent of the size of the training set. Additionally, cross-species performance for each species-specific predictor is not as good as the self-species performance whereas the cross-species performance generally increases with the size of the training dataset. Moreover, the predictors developed based on the non-histone Kcrot sites are unsuccessful for the histone Kcrot prediction, suggesting that the Kcrot-containing peptides from non-histone and histone proteins have significantly different characteristics and data integration is required. Overall, DeepKcrot is an efficient prediction tool and freely available at http://www.bioinfogo.org/deepkcrot.

Sanghuangporus is a traditional medicine that has been used for nearly 2000 years in Asia. It has been found to enhance immunity and have anticancer properties. Some studies determined that Sanghuangporus polysaccharide can inhibit tumors in vitro, although the underlying mechanisms remain unknown. Therefore, the present study investigated the antitumor effects of polysaccharides from the fruiting body of S. vaninii and their associated mechanisms. Water-soluble S. vaninii polysaccharide (SVP) with a molecular weight of 3.156 × 104 Da was isolated and found to be mainly composed of mannose, rhamnose, glucuronic acid, galacturonic acid, glucosamine, glucose, galactosamine, galactose, xylose, arabinose, and fucose in molar ratios of 1.63:0.04:0.36:0.03:0.13:8.39:0.08:1.08:0.25:1.07:0.40. SVP showed modest anti-proliferative activity against HeLa, SHG-44, SMMC-7721, and MCF-7 cells. Furthermore, it effectively regulated cell cycle, promoted apoptosis, and reduced the migratory and invasive capacities of MCF-7 cells. Mechanistically, SVP enhanced activation of p53-related genes and down-regulated MMP expression. These findings describe a potential natural polysaccharide for tumor therapy and a basis for the development of fungal polysaccharides as a type of health food.

Small intestinal neuroendocrine tumors (SI-NET) are serotonin-secreting well-differentiated neuroendocrine tumors of putative enterochromaffin (EC) cell origin. Recent studies recognize a subset of EC cells that is label-retaining at the +4 position in the crypt and functions as a reserve intestinal stem cell. Importantly, this +4 reserve EC cell subset not only contributes to regeneration of the intestinal epithelium during injury and inflammation but also to basal crypt homeostasis at a constant rate. The latter function suggests that the +4 EC cell subset serves as an active reserve stem cell via a constant rate of dedifferentiation. Characterization of early tumor formation of SI-NET, observed as crypt-based EC cell clusters in many cases of familial SI-NETs, suggests that the +4 active reserve EC cell subset is the cell of origin. This newly discovered active reserve stem cell property of EC cells can account for unique biological mechanisms and processes associated with the genesis and development of SI-NETs. The recognition of this property of the +4 active reserve EC cell subset may provide novel opportunities to explore NETs in the gastrointestinal tract and other organs.

To date, four species of porcine circoviruses (PCVs), including PCV1-4, have been reported to exist in the clinical cases. Fast and effective differential detection is critical to monitor the infection and co-infection status of PCVs for adopting reliable control strategies. However, currently available methods cannot simultaneously differentiate the four species of PCV strains. In this study, a quadruplex real-time PCR assay based on TaqMan probes was developed for differential detection of PCV1-4. The new quadruplex real-time PCR assay exhibited satisfied specificity, sensitivity, repeatability and reproducibility. In addition, the new assay was applied to the detection of 120 clinical samples collected from 2016 to 2020 in Jiangsu province of China and compared with previously reported PCV1-4 singleplex conventional PCR assays. Based on the clinical performance, the results from the quadruplex real-time PCR and conventional PCR assays showed 100% agreement. A total of 47 samples were detected as PCV positive by the quadruplex real-time PCR assay, including 1, 2, 1 samples were co-infected with PCV1 and PCV4, PCV2 and PCV3, PCV2 and PCV4, respectively. Full-length ORF2 sequencing and phylogenetic analysis supported the real-time PCR results that 5, 34, 8 and 4 of the 51 PCV sequences were PCV1, PCV2, PCV3 and PCV4, respectively. This study provides a promising alternative tool for rapid differential detection of PCVs and confirms the coexistence of all species of PCV1-4 strains in Jiangsu province in recent years.

Carpal tunnel syndrome (CTS) is the most common peripheral nerve entrapment syndrome, affecting a large proportion of the general population. Genetic susceptibility has been implicated in CTS, but the causative genes remain elusive. Here, we report the identification of two mutations in cartilage oligomeric matrix protein (COMP) that segregate with CTS in two large families with or without multiple epiphyseal dysplasia (MED). Both mutations impair the secretion of COMP by tenocytes, but the mutation associated with MED also perturbs its secretion in chondrocytes. Further functional characterization of the CTS-specific mutation reveals similar histological and molecular changes of tendons/ligaments in patients’ biopsies and the mouse models. The mutant COMP fails to oligomerize properly and is trapped in the ER, resulting in ER stress-induced unfolded protein response and cell death, leading to inflammation, progressive fibrosis and cell composition change in tendons/ligaments. The extracellular matrix (ECM) organization is also altered. Our studies uncover a previously unrecognized mechanism in CTS pathogenesis. Familial carpal tunnel syndrome (CTS) is common, but causal genes are not characterized. Here the authors report two CTS-related mutations in two large families that impair secretion of COMP in tenocytes, leading to ER stress-induced unfolded protein response, inflammation and fibrosis in patients and mouse models.

Background Epidermal growth factor receptor (EGFR) in trans-C797S alteration has been considered as one of the mechanisms, which leads to the resistance to third-generation tyrosine kinase inhibitors (TKI). Several reports have suggested that combination therapy with first- and third-generation TKIs might be effective for acquired trans-C797S alteration. Case presentation Herein, we report a patient diagnosed with lung adenocarcinoma harboring EGFR 19Del/T790M/in trans-C797S mutations with co-occurring CTNNB1 alteration showing a negative response to combination therapy with first- and third-generation TKIs. Conclusion Our report highlights the importance of deploying more informed molecular diagnosis and real-time monitoring strategies to detect co-alterations that drive disease progression and drug resistance, in order to find more effective treatment.

Dieldrin has been shown to induce liver tumors selectively in mice. Although the exact mechanism is not fully understood, previous studies from our laboratory and others have shown that dieldrin induced liver tumors in mice through a non-genotoxic mechanism acting on tumor promotion stage. Two studies were performed to examine the role of nuclear receptor activation as a possible mode of action (MOA) for dieldrin-induced mouse liver tumors. In the initial study, male C57BL/6 mice (6- to 8-week old) were treated with dieldrin in diet (10 ppm) for 7, 14, and 28 days. Phenobarbital (PB), beta-naphthoflavone (BNF) and Di (2-ethylhexyl) phthalate (DEHP) were included as positive controls in this study for evaluating the involvement of CAR (constitutive androstane receptor), AhR (aryl hydrocarbon receptor) or PPARα (peroxisome proliferator activated receptor alpha) in the MOA of dieldrin hepatocarcinogenesis. A significant increase in hepatocyte DNA synthesis (BrdU incorporation) was seen in treated mice compared with the untreated controls. Analysis of the expression of the nuclear receptor responsive genes revealed that dieldrin induced a significant increase in the expression of genes specific to CAR activation (Cyp2b10, up to 400- to 2700-fold) and PXR activation (Cyp3a11, up to 5- to 11-fold) over untreated controls. The AhR target genes Cyp1a1 and Cyp1a2 were also slightly induced (2.0- to 3.7-fold and 1.7- to 2.8-fold, respectively). PPARα activation was not seen in the liver following dieldrin treatment. In addition, consistent with previous studies in our lab, treatment with dieldrin produced significant elevation in the hepatic oxidative stress. In a subsequent study using CAR, PXR, and CAR/PXR knockout mice, we confirmed that the dieldrin-induced liver effects in mouse were only mediated by the activation of CAR receptor. Based on these findings, we propose that dieldrin induced liver tumors in mice through a nuclear receptor CAR-mediated mode of action. The previously observed oxidative stress/damage may be an associated or modifying factor in the process of dieldrin-induced liver tumor formation subsequent to the CAR activation.

Background & aims Human TFB2M (mitochondrial transcription factor B2) is a key regulator of mitochondria transcription. Our bioinformatic analysis based on the cancer genome atlas (TCGA) data revealed an aberrant over-expression of TFB2M in hepatocellular carcinoma (HCC). However, the functional roles of TFB2M in tumourigenesis remains unexplored, including HCC. Methods The expression and clinical significance of TFB2M were evaluated by qRT-PCR and western blot analysis. The biological effects and underlying mechanisms of TFB2M in HCC were determined by cell proliferation, colony formation, cell cycle, apoptosis, migration and invasion assays. Results TFB2M was commonly up-regulated in HCC mainly because of the down-regulation of miR101-3p, which significantly correlated with poor survival of HCC patients. Functional experiments revealed that TFB2M significantly promoted HCC cell proliferation, migration and invasion, while inhibited apoptosis in vitro and promoted xenograft tumourigenesis and lung metastasis in nude mice models in vivo. Mechanistically, increased production of reactive oxygen species (ROS) and subsequently activated Akt/NF-κB signalling was found to be involved in the promotion of growth and metastasis by TFB2M in HCC cells. Conclusions These findings suggest that TFB2M plays a pivotal oncogenic role in HCC cells through activating ROS-Akt-NF-κB signalling pathway.

The single and synergistic effects of dietary arabinoxylan-oligosaccharide (AXOS) and inulin at different doses (2, 4, and 8 mg/g diet) on survival, growth performance, gut microbiota, and immune response in Pacific white shrimp Litopenaeus vannamei were assessed. Singular application of either inulin or AXOS at doses of 4 mg/g diet showed the most stimulatory effects on the growth rate and gene expression levels of chitinase, cathepsin L, chymotrypsin, and extracellular signal-regulated kinase (ERK) in each single prebiotic feeding trial. Compared with single prebiotic treatments, simultaneous application of AXOS and inulin at 4 mg/g diet enhanced remarkably the growth parameters of shrimp and the expression of related genes (chitinase, cathepsin L, chymotrypsin, ERK, myeloid differentiation factor 88, and phenoloxidase) after 8-week feeding (P < 0.05). Additionally, gut microbiota analysis indicated the dietary supplementation with combined prebiotics increased significantly the bacterial community richness and relative abundance of Bacillus, Pseudomonas, Bacteriovorax, and Lactobacillus, and reduced the abundance of Vibrio, Rhodococcus, and Photobacterium in the digestive tract of L. vannamei. Compared with the single prebiotic treatment and the control, combined prebiotics supplementation boosted notably the survival rate and expression levels of immune-related genes in shrimp infected with Vibrio alginolyticus or white spot syndrome virus. Therefore, simultaneous application of AXOS and inulin shall have a great potential of dietary supplement in the culture of L. vannamei.

Background Patchouli alcohol is an effective component of the medicinal plant patchouli. Similar to other secondary metabolites, its synthesis is also regulated by transcription factors. Although the biosynthetic pathway of patchouli alcohol has been characterized, the regulatory mechanism of patchouli alcohol has not been fully revealed. Results This study combined the transcriptome data of patchouli leaves treated with different hormones and WGCNA to complete the coexpression network. The modules related to patchouli alcohol were identified, and PcMYB25 played a crucial role in regulating patchouli alcohol biosynthesis. The overexpression of PcMYB25 can promote the expression of PTS , thereby increasing the content of patchouli alcohol. Conclusions This is the first reporter that MYB25 regulates the secondary metabolism of patchouli. These experimental results lay the foundation for further analysis of the regulatory mechanism of patchouli alcohol synthesis.

Hepatocellular carcinoma (HCC) is a common malignancy in human. CD44 is a transmembrane glycoprotein which is frequently overexpressed in cancer of various origins. The function and mechanism of CD44 in HCC remains elusive. In this study, we reported that CD44 was overexpressed in HCC to promote the proliferation and migration of HCC cells via oncogenic YAP, which is the key downstream regulator in Hippo pathway. These findings suggest that CD44-YAP is a probable important axis in pathogenesis of HCC, providing an insight in to HCC pathogenesis as well as potential targets for the intervention of HCC. Supplementary Information The online version contains supplementary material available at 10.1186/s40164-021-00247-w.

Insulin-like growth factor 3 (IGF3), a newly discovered member of the IGF family in teleosts, plays a vital role in the reproduction system of fishes. Therefore, this study explored the spotted scat (Scatophagus argus), which is an aquaculture species of immense economic importance, but the regulatory mechanism of IGF3 in its ovarian development remains unknown. Here, we cloned igf3 cDNA in S. argus. Sequence analysis showed that the ORF of igf3 is 594 bp long and encodes 197 amino acid residues, possessing conserved characteristics of the IGF family. Tissue distribution revealed that igf3 is expressed in three different tissues: brain, gonad, and kidney. Following in situ hybridization, igf3 was detected in follicles, and its localization shifted during ovarian development. By qPCR analysis, the expression level of igf3 mRNA in the ovary increased as the ovary developed and was highest at stage Ⅳ which is approaching maturation. This study also produced recombinant S. argus IGF3 in bacteria to analyze the biological functions of IGF3. Incubating ovarian fragments with the recombinant IGF3 significantly promoted the cdc2, cyclinB1, 20β-HSD, egf, cyp11a1, cyp17a1, cyp19a1a, hsd17b1, and star1 expression levels, which are the key genes for ovarian development and maturation. The β-catenin signaling-sensitive genes, including cyclinD1 and axin2, were also promoted by IGF3 treatment. Collectively, our results demonstrate that IGF3 exerts functions on the development and maturation of ovaries in S. argus.

Melanocortin 1 receptor (MC1R) is thought to be a marker of poor prognosis and a potential target for the treatment of melanoma. Studies have found that MC1R promotes several tumor behaviors, including cell proliferation and differentiation, pigment formation, and genome damage repair. Some single-nucleotide polymorphisms (SNPs) of MC1R are involved in the occurrence and development of melanoma. A few studies have reported a relationship between MC1R and colorectal cancer (CRC). In this research, our objective was to examine MC1R expression and MC1R SNPs and investigate their correlation with the clinicopathological features of human CRC tissues. We evaluated MC1R mRNA expression by performing bioinformatic analyses on human CRC expression datasets. We used Western blotting and RT-qPCR to compare MC1R expression in CRC tissues with that in normal tissues, and MC1R SNPs in CRC tissues were detected by PCR-direct sequencing (DS). The expression of MC1R was significantly decreased in CRC tissues compared with normal tissue, and its expression was negatively associated with P53 expression, MLH1 expression, and PMS2 expression, and high MC1R expression was significantly associated with microsatellite instability (MSI). MC1R SNPs were also associated with the clinicopathological characteristics of CRC; for example, the rs2228479 locus genotype was correlated with Ki67 status, and the rs885479 locus genotype was correlated with age and T stage. In conclusion, MC1R plays a crucial role in the progression of CRC and may be a marker of poor prognosis in CRC.

BackgroundBrain tumor ranks as the most devastating cancer type. The complex tumor immune microenvironment prevents brain tumor from receiving therapeutic benefits. The purpose of this study was to stratify brain tumors based on their distinct immune infiltration signatures to facilitate better clinical decision making and prognosis prediction.MethodsWe developed a deep learning model to characterize immune infiltration from transcriptome. The developed model was applied to distill expression signatures of transcriptome of brain tumor samples. We performed molecular subtyping with the extracted expression signatures to unveil brain tumor subtypes. Computational methods, including gene set enrichment analysis, Kaplan-Meier survival and multivariate Cox regression analyses, were employed.ResultsWe identified two distinctive subtypes (i.e. C1/2) of brain tumor featured by distinct immune infiltration signatures. The C1 subtype is characterized by protective immune infiltration signatures, including high infiltration of CD8+ T cells and activation of CX3CL1. The C2 subtype has an extensive infiltration of tumor-associated macrophages and microglia, and was enriched with immune suppressive, wound-healing, and angiogenic signatures. The C1 subtype had significantly better prognosis as compared with C2 (Log-rank test, HR: 2.5, 95% CI: 2.2 – 2.7; P = 8.2e-78). This difference remained statistically significant (multivariate Cox model, HR: 2.2, 95% CI: 1.7 – 2.9; P = 3.7e-10) by taking into account age, gender, recurrent/secondary status at sampling time, tumor grade, histology, radio-chemotherapy, IDH mutation, MGMT methylation, and co-deletion of 1p and 19q. This finding was validated in six datasets. The C2 subtype of glioblastoma patients with IDH mutation has poor survival analogous to those without IDH mutation (Log-rank test, adjusted P = 0.8), while C1 has favorable prognosis as compared with glioblastoma of C2 subtype with IDH mutation (Log-rank test, adjusted P = 1.2e-3) or without IDH mutation (Log-rank test, adjusted P = 1.3e-6).ConclusionsWe identified two distinctive subtypes of brain tumor with different immune infiltration signatures, which might be helpful as an independent prognosticator for brain tumor.

Simple Summary Skeletal muscle development and meat quality are key traits of considerable importance to consumers and farmers. In the past, second-generation sequencing technology has been used to study genes regulating muscle development and meat quality. However, with the limitation of read length, goat transcriptome was constructed mainly on the basis of the merging of short reads, resulting in an insufficient understanding of goat transcriptome structures. Identification of full-length transcript structure was still a challenge. Therefore, in this study, a hybrid sequencing was conducted that combined the long-reading character of third-generation sequencing with the quantification ability of second-generation sequencing. By comparing the longissimus dorsi and biceps femoris muscles, genes and transcript isoforms regulating meat quality and muscle development of goat were identified. A large number of novel loci and isoforms were identified in the goats. Functional annotation of these genes showed that they were associated with skeletal muscle development and lipid metabolism. Abstract Domestic goats are commonly reared for meat and milk production in several regions of the world. However, the genetic mechanism underlying muscle development and meat quality of goats is limited. Therefore, the aim of this study was to identify known and novel genes regulating muscle development and meat quality of goats using second- and third-generation sequencing technologies. To achieve this, the meat quality and transcriptomes of longissimus dorsi (LD) and biceps femoris (BF) muscle tissues of Lingqiu Greyback goats were examined and compared. Differentially expressed genes (DEGs) and isoforms (DEIs) were functionally annotated. Results showed that 45,574 full-length transcripts covering 18,491 loci were characterized, and 12,566 genes were co-expressed in all samples. Differential expression analysis identified 231 DEGs, including 45 novel genes in the LD and BF muscles of the goats. Additionally, 1173 DEIs were found, in which 642 novel isoforms were identified in this study. Functional annotation and pathway analysis of the DEGs and DEIs revealed that some of them were associated with muscle growth and lipid metabolism. Overall, the findings of this study contribute to the understanding of the transcriptomic diversity underlying meat quality and muscle development of goat.

Most studies of gut microbiota have focused on relationships between a specific disease and the presence/abundance of one or a few bacterial species/genera. Whether the spatial and temporal distribution of gut microbiota, as a whole, affects or correlates with health is unknown, largely due to the absence of tools for dynamically monitoring the overall gut microbiota landscape inside living subjects. Here, we describe a novel, noninvasive, live imaging method for gut microbiota using 2-deoxy-2-[18F]fluoro-d-sorbitol (18F-FDS), a compound that specifically labeled gut bacteria in mice and hamsters following oral administration. Positron emission tomography-computed tomography (PET-CT) scanning showed that the radiolabel signal was concentrated in the gut (especially the large intestine), was absent when mice gut microbiota was depleted by antibiotic treatment, and was restored after transplanting antibiotic-treated mice with a fecal or probiotic bacterial mixture. Thus, 18F-FDS images microbiota, not gut tissue. The tissue distribution of 18F-FDS was the highest in the gut (∼3-fold higher than average), in contrast to 2-deoxy-2-[18F]fluoro-d-glucose, which concentrated in brain and many other organs. 2-[18F]fluoro-aminobenzoic acid, another bacterium-specific radioactive tracer, was unsuited for gut microbiota imaging due to unexpected stomach retention following oral administration. When similar gut microbiota imaging was done with hamsters, the spatial resolution increased significantly over that with mice, suggesting that even higher spatial resolution can be achieved with humans or large animals. Thus, our work establishes a new tool for noninvasive, live imaging of gut microbiota; the new tool may enable exploration of relationships between gut microbiota landscape and diseases in clinical settings. IMPORTANCE Gut microbiota dysbiosis correlates with many diseases, but such correlations derive mostly from relationships between one or a few bacteria and a particular disease. Since microbiota resemble complex forest ecosystems more closely than individual patches of trees, the overall landscape (spatial and temporal distribution) of gut bacteria may also affect/reflect disease development. Such a possibility has not been explored due to a lack of tools for directly visualizing natural landscape patterns of gut microbiota. The present work identified 2-deoxy-2-[18F]fluoro-d-sorbitol as a gut microbiota-specific radioactive tracer and developed a novel PET-CT scan-based imaging method that enables noninvasive, real-time imaging of the overall gut bacterial landscape. The method showed increased spatial resolution when hamsters replaced mice, suggesting that even higher spatial resolution could be achieved with larger animals such as humans. This novel technology establishes the feasibility of investigating spatial-temporal distribution dynamics of gut microbiota with many human diseases.

Heterosis is a common biological phenomenon in nature. It substantially contributes to the biomass yield and grain yield of plants. Moreover, this phenomenon results in high economic returns in agricultural production. However, the utilization of heterosis far exceeds the level of theoretical research on this phenomenon. In this review, the recent progress in research on heterosis in plants was reviewed from the aspects of classical genetics, parental genetic distance, quantitative trait loci, transcriptomes, proteomes, epigenetics (DNA methylation, histone modification, and small RNA), and hormone regulation. A regulatory network of various heterosis-related genes under the action of different regulatory factors was summarized. This review lays a foundation for the in-depth study of the molecular and physiological aspects of this phenomenon to promote its effects on increasing the yield of agricultural production.

Flavonoids from plants play an important role in our diet. Watercress is a special plant that is rich in flavonoids. In this study, four important watercress varieties were compared with non-heading Chinese cabbage by ultra-high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UHPLC-ESI-MS/MS). A total of 132 flavonoid metabolites (including 8 anthocyanins, 2 dihydroflavone, 3 dihydroflavonol, 1 flavanols, 22 flavones, 11 flavonoid carbonosides, 82 flavonols, and 3 isoflavones) were detected. Flavonoid metabolites varied widely in different samples. Both the non-heading Chinese cabbage and the variety of watercress from Guangdong, China, had their own unique metabolites. This work is helpful to better understand flavonoid metabolites between the non-heading Chinese cabbage and the other four watercress varieties, and to provide a reliable reference value for further research.

Current COVID-19 vaccines need to take at least one month to complete inoculation and then become effective. Around 51% global population are still not fully vaccinated. Instantaneous protection is an unmet need among those who are not fully vaccinated. In addition, breakthrough infections caused by SARS-CoV-2 are widely reported. All these highlight the unmet needing for short-term instantaneous prophylaxis (STIP) in the communities where SARS-CoV-2 is circulating. Previously, we reported nanobodies isolated from an alpaca immunized with the spike protein, exhibiting ultrahigh potency against SARS-CoV-2 and its variants. Herein, we found that Nb22, among our previously reported nanobodies, exhibited ultrapotent neutralization against Delta variant with an IC50 value of 0.41 ng/ml (5.13 pM). Furthermore, the crystal structural analysis revealed that the binding of Nb22 to WH01 and Delta RBDs both effectively blocked the binding of RBD to hACE2. Additionally, intranasal Nb22 exhibited protection against SARS-CoV-2 Delta variant in the post-exposure prophylaxis (PEP) and pre-exposure prophylaxis (PrEP). Of note, intranasal Nb22 also demonstrated high efficacy against SARS-CoV-2 Delta variant in STIP for seven days administered by single dose and exhibited long-lasting retention in the respiratory system for at least one month administered by four doses, providing a means of instantaneous short-term prophylaxis against SARS-CoV-2. Thus, ultrahigh potency, long-lasting retention in the respiratory system as well as stability at room-temperature make the intranasal or inhaled Nb22 to be a potential therapeutic or STIP agent against SARS-CoV-2.Brief summaryNb22 exhibits ultrahigh potency against Delta variant in vitro and is exploited by crystal structural analysis; furthermore, animal study demonstrates high effectiveness in the treatment and short-term instantaneous prophylaxis in hACE2 mice via intranasal administration.HighlightsNb22 exhibits ultrapotent neutralization against Delta variant with an IC50 value of 0.41 ng/ml (5.13 pM).Structural analysis elucidates the ultrapotent neutralization of Nb22 against Delta variant.Nb22 demonstrates complete protection in the treatment of Delta variant infection in hACE2 transgenic mice.We complete the proof of concept of STIP against SARS-CoV-2 using intranasal Nb22 with ultrahigh potency and long-lasting retention in respiratory system.Graphic Abstract

Ciguatera poisoning is mainly caused by the consumption of reef fish that have accumulated ciguatoxins (CTXs) produced by the benthic dinoflagellates Gambierdiscus and Fukuyoa. China has a long history of problems with ciguatera, but research on ciguatera causative organisms is very limited, especially in the Beibu Gulf, where coral reefs have been degraded significantly and CTXs in reef fish have exceeded food safety guidelines. Here, five strains of Gambierdiscus spp. were collected from Weizhou Island, a ciguatera hotspot in the Beibu Gulf, and identified by light and scanning electron microscopy and phylogenetic analyses based on large and small subunit rDNA sequences. Strains showed typical morphological characteristics of Gambierdiscus caribaeus, exhibiting a smooth thecal surface, rectangular-shaped 2′, almost symmetric 4″, and a large and broad posterior intercalary plate. They clustered in the phylogenetic tree with G. caribaeus from other locations. Therefore, these five strains belonged to G. caribaeus, a globally distributed Gambierdiscus species. Toxicity was determined through the mouse neuroblastoma assay and ranged from 0 to 5.40 fg CTX3C eq cell−1. The low level of toxicity of G. caribaeus in Weizhou Island, with CTX-contaminated fish above the regulatory level in the previous study, suggests that the long-term presence of low toxicity G. caribaeus might lead to the bioaccumulation of CTXs in fish, which can reach dangerous CTX levels. Alternatively, other highly-toxic, non-sampled strains could be present in these waters. This is the first report on toxic Gambierdiscus from the Beibu Gulf and Chinese waters and will provide a basis for further research determining effective strategies for ciguatera management in the area.

Selecting superior clones is the first step for commercial short‐rotation coppice cultures to provide biomass and bioenergy. Till date, such selection for hybrid Populus clones in middle China is absent. Here we describe the growth, aboveground biomass production and cell wall composition of 27 hybrid poplar clones in Henan, China for two 3‐year rotations. Significant variation in these three characteristics over two triennial rotation coppices among the 27 poplar clones was observed. During two 3‐year rotation coppices, clones ‘276’, ‘02‐17’, and ‘599’ showed relatively higher tree heights and larger basal diameters than those of the other clones. However, the most productive clones were ‘36’ and ‘01‐30’. At the end of the second triennial rotation, the aboveground biomass production reached 18 Mg ha−1 year−1. For the cell wall composition analysis, the cellulose contents of clones ‘01‐243’ and ‘2001’ were relatively high, while the xylose contents of clones ‘01‐30’ and ‘65’ were relatively high. Cluster analysis based on height, basal diameter, biomass, heat value, cellulose content, and survival rate revealed five growth potential classes. Accordingly, clones ‘03‐332’, ‘36’, and ‘599’ exhibited high biomass and growth and had the greatest potential to serve as excellent biomass producers in Henan, China. In addition, the expression patterns of 20 key regulatory genes were analyzed, and an integrated coexpression network was constructed. This field trial provides a comprehensive quantification and evaluation of the agronomic performance of 27 poplar clones in Henan, China. The results of this study and the analytical strategies provide an efficient mechanism for selecting clones that will perform well agronomically in local environments. The expression of key genes and the integrated coexpression network provide the molecular mechanisms of poplar biomass performance.

We investigated the role of microRNA (miR)-9 in modulating chemoresistance in hepatocellular carcinoma (HCC) cells. MiR-9 was overexpressed or knocked down in HCC cell lines. Cell viability, cell proliferation, the expression of EIF5A2 and the epithelial-mesenchymal transition (EMT)-related proteins were examined. HCC cells overexpressing miR-9 were more sensitive to cisplatin; miR-9 knockdown yielded the opposite result. The in vivo nude mouse HCC xenograft tumors yielded the same results. EIF5A2 was identified as a potential target of miR-9, where miR-9 regulated EIF5A2 expression at mRNA and protein level. EIF5A2 knockdown reversed miR-9 inhibition-mediated cisplatin resistance. Altering miR-9 and EIF5A2 expression changed E-cadherin and vimentin expression. Furthermore, EIF5A2 mediated miR-9 EMT pathway regulation, indicating that miR-9 can enhance cisplatin sensitivity by targeting EIF5A2 and inhibiting the EMT pathway. Targeting miR-9 may be useful for overcoming drug resistance in HCC.

Breast cancer is one of the most common malignant cancers among females worldwide. This complex disease is not caused by a single gene, but resulted from multi-gene interactions, which could be represented by biological networks. Network modules are composed of genes with significant similarities in terms of expression, function and disease association. Therefore, the identification of disease risk modules could contribute to understanding the molecular mechanisms underlying breast cancer. In this paper, an integrated disease risk module identification strategy was proposed according to a multi-objective programming model for two similarity criteria as well as significance of permutation tests in Markov random field module score, function consistency score and Pearson correlation coefficient difference score. Three breast cancer risk modules were identified from a breast cancer-related interaction network. Genes in these risk modules were confirmed to play critical roles in breast cancer by literature review. These risk modules were enriched in breast cancer-related pathways or functions and could distinguish between breast tumor and normal samples with high accuracy for not only the microarray dataset used for breast cancer risk module identification, but also another two independent datasets. Our integrated strategy could be extended to other complex diseases to identify their risk modules and reveal their pathogenesis.

Genotoxic compounds may be detoxified to non-genotoxic metabolites while many pro-carcinogens require metabolic activation to exert their genotoxicity in vivo. Standard genotoxicity assays were developed and utilized for risk assessment for over 40 years. Most of these assays are conducted in metabolically incompetent rodent or human cell lines. Deficient in normal metabolism and relying on exogenous metabolic activation systems, the current in vitro genotoxicity assays often have yielded high false positive rates, which trigger unnecessary and costly in vivo studies. Metabolically active cells such as hepatocytes have been recognized as a promising cell model in predicting genotoxicity of carcinogens in vivo. In recent years, significant advances in tissue culture and biological technologies provided new opportunities for using hepatocytes in genetic toxicology. This review encompasses published studies (both in vitro and in vivo) using hepatocytes for genotoxicity assessment. Findings from both standard and newly developed genotoxicity assays are summarized. Various liver cell models used for genotoxicity assessment are described, including the potential application of advanced liver cell models such as 3D spheroids, organoids, and engineered hepatocytes. An integrated strategy, that includes the use of human-based cells with enhanced biological relevance and throughput, and applying the quantitative analysis of data, may provide an approach for future genotoxicity risk assessment.

Chinese cabbage (Brassica rapa L. ssp. pekinensis) is one of the most important economic crops. However, its yield and quality can be severely threatened by Turnip mosaic virus (TuMV). Emerging evidence indicates that epigenetic mechanisms, especially DNA methylation, play an important role in regulating gene expression. Therefore, identification of resistance genes modified by DNA methylation during the virus infection would provide a critical clue for improving disease resistance breeding programs. Here, we present detailed analysis for the correlation of DNA methylation and gene expression involved in several anti-pathogen pathways. We also found that different methylation patterns exist in different methylation sites (CG, CHG, and CHH, where H represents A, G, or T) and genomic regions. Furthermore, we identified disease-resistant genes related to the nucleotide binding site-leucine-rich repeats family, auxin, salicylic acid signaling transduction, cell wall biosynthesis, and protein degradation among the different methylated genes (DMGs) suggesting that these genes may be modified by DNA methylation and work together to activate an immune response. The identified DMGs are a valuable resource for discovering resistance genes. Our study not only provides valuable data for future biotechnology research and epigenetic studies, but also helps to explore how the epigenetic mechanisms modify antiviral pathways.

The MAF (MADS AFFECTING FLOWERING)/FLC (FLOWERING LOCUS C) subfamily is one of the key families that control flowering. This family has been well studied and characterized in Arabidopsis thaliana and other model plants. However, information regarding the evolution and functions of MAF/FLC genes in Brassica rapa (B. rapa) is limited. To further study the evolutionary history of MAF/FLC family during flowering of Pak-choi (B. rapa ssp. chinensis), the evolutionary patterns, footprints, and conservation of the MAF/FLC family in selected plants were investigated. The results showed that splitting of Brassica-Arabidopsis resulted in a preferential retention of the MAF/FLC family in Brassicas. Sequence similarity and the intracellular expression pattern of BrMAF2 suggested that the function of the MAF/FLC family might be conserved in B. rapa. However, protein interactions revealed that given the high sequence similarity between BrMAFs and AtMAFs, functional divisions of the BrMAF gene family still occurred. Additionally, the temporal and spatial expression of MAF/FLC genes in B. rapa further confirmed that a functional division occurred during gene duplication. Meanwhile, our finding partially confirmed the triplication event in B. rapa at the gene expression level. Furthermore, we found that high gene sequence homology does not indicate high function conservation. Thus, we suggest that when we name a gene in other plants, both gene sequence homology and function verification are needed. In addition, our studies also suggest that a new candidate BrMAF3 gene may be more important than other BrMAFs in the response to brief cold spells. Finally, the results of this study can help to better understand the evolution and function of MAF/FLC in plants.

Lung adenocarcinoma (LUAD), which comprises over 50% of all cases of non-small-cell lung cancer, has a poor prognosis and requires novel therapeutic approaches. The sonic Hedgehog (Shh) pathway, which plays a crucial role in differentiation, proliferation, and survival of cancer cells, is likely to be activated in LUADs, suggesting the Shh pathway as a potential therapeutic target for LUAD treatment. In this study, we reported that vismodegib, an inhibitor of the Shh pathway, only elicited minor antitumor efficacy in A549 and NCI-H1975 LUAD cells as well as in the xenograft tumors, with overexpressed GLI2 and increased autophagic activity. The aberrant autophagy in LUAD cells was further confirmed by the three main stages of autophagic flux, including the formation of autophagosomes, the fusion of autophagosomes with lysosomes, and degradation of autophagosomes in lysosomes. Furthermore, inhibition of autophagy by siRNA against ATG5 or ATG7 rescued the sensitivity of A549 and NCI-H1975 LUAD cells to vismodegib in vitro. Meanwhile, administration of the pharmaceutical inhibitor of autophagy, chloroquine, contributed to the enhanced anti-LUAD efficacy of vismodegib in vivo, probably through overproduction of ROS, acceleration of apoptosis, and suppression of GLI2 in LUAD tissues. In summary, our research revealed that downregulating autophagy facilitated the anti-LUAD efficacy of the Shh pathway suppression, thus highlighting a potential approach for LUAD therapy via simultaneously targeting the Shh signaling and autophagy pathway.

Plants have evolved various strategies to sense and respond to saline environments, which severely reduce plant growth and limit agricultural productivity. Alteration to the cell wall is one strategy that helps plants adapt to salt stress. However, the physiological mechanism of how the cell wall components respond to salt stress is not fully understood. Here, we show that expression of XTH30, encoding xyloglucan endotransglucosylase-hydrolase30, is strongly up-regulated in response to salt stress in Arabidopsis. Loss-of-function of XTH30 leads to increased salt tolerance and overexpression of XTH30 results in salt hypersensitivity. XTH30 is located in the plasma membrane and is highly expressed in the root, flower, stem, and etiolated hypocotyl. The NaCl-induced increase in xyloglucan (XyG)-derived oligosaccharide (XLFG) of the wild type is partly blocked in xth30 mutants. Loss-of-function of XTH30 slows down the decrease of crystalline cellulose content and the depolymerization of microtubules caused by salt stress. Moreover, lower Na+ accumulation in shoot and lower H2O2 content are found in xth30 mutants in response to salt stress. Taken together, these results indicate that XTH30 modulates XyG side chains, altered abundance of XLFG, cellulose synthesis, and cortical microtubule stability, and negatively affecting salt tolerance., Xyloglucan endotransglucosylase-hydrolase30 modulates xyloglucan side chains by altering abundance of xyloglucan-derived oligosaccharide, negatively affecting salt tolerance.

Background Rapid-cycling Brassica rapa (RCBr), also known as Wisconsin Fast Plants, are small robust plants with a short lifecycle that are widely used in biology teaching. RCBr have been used for decades but there are no published reports of RCBr genetic transformation. Agrobacterium-mediated vacuum infiltration has been used to transform pakchoi (Brassica rapa ssp. chinensis) and may be suitable for RCBr transformation. The floral dip transformation method, an improved version of vacuum infiltration, could make the procedure easier. Results Based on previous findings from Arabidopsis and pakchoi, plants of three different ages were inoculated with Agrobacterium. Kanamycin selection was suboptimal with RCBr; a GFP screen was used to identify candidate transformants. RCBr floral bud dissection showed that only buds with a diameter less than 1 mm carried unsealed carpels, a key point of successful floral dip transformation. Plants across a wide range of inflorescence maturities but containing these immature buds were successfully transformed, at an overall rate of 0.1% (one per 1000 T1 seeds). Transformation was successful using either vacuum infiltration or the floral dip method, as confirmed by PCR and Southern blot. Conclusion A genetic transformation system for RCBr was established in this study. This will promote development of new biology teaching tools as well as basic biology research on Brassica rapa. Electronic supplementary material The online version of this article (10.1186/s12870-019-1843-6) contains supplementary material, which is available to authorized users.

Carrots (Daucus carota L.), among the most important root vegetables in the Apiaceae family, are cultivated worldwide. The storage root is widely utilized due to its richness in carotenoids, anthocyanins, dietary fiber, vitamins and other nutrients. Carrot extracts, which serve as sources of antioxidants, have important functions in preventing many diseases. The biosynthesis, metabolism, and medicinal properties of carotenoids in carrots have been widely studied. Research on hormone regulation in the growth and development of carrots has also been widely performed. Recently, with the development of high-throughput sequencing technology, many efficient tools have been adopted in carrot research. A large amount of sequence data has been produced and applied to improve carrot breeding. A genome editing system based on CRISPR/Cas9 was also constructed for carrot research. In this review, we will briefly summarize the origins, genetic breeding, resistance breeding, genome editing, omics research, hormone regulation, and nutritional composition of carrots. Perspectives about future research work on carrots are also briefly provided., Carrots: past and future China accounts for nearly half of the total global production of carrots and advances in genetic technologies are contributing to improve both crop quality and yield. Cultivated carrot is the second most popular vegetable in the world after potato; this can be largely attributed to their taste and their nutritional benefits. Ai-Sheng Xiong and colleagues at Nanjing Agricultural University, China, review the latest studies on the origin and breeding of carrots. Understanding the genetic sequence and gene expression patterns in carrot plants provides valuable insights into their evolution as well as into the mechanisms underlying their resistance to disease, production of healthy carotenoids and environmental stress tolerance. Future application of gene editing technologies will help to further improve crop production and the nutritional value of carrots.

Watercress (Nasturtium officinale R. Br.), an aquatic leafy vegetable of the Brassicaceae family, is known as a nutritional powerhouse. Here, we de novo sequenced and assembled the complete chloroplast (cp) genome of watercress based on combined PacBio and Illumina data. The cp genome is 155,106 bp in length, exhibiting a typical quadripartite structure including a pair of inverted repeats (IRA and IRB) of 26,505 bp separated by a large single copy (LSC) region of 84,265 bp and a small single copy (SSC) region of 17,831 bp. The genome contained 113 unique genes, including 79 protein-coding genes, 30 tRNAs and 4 rRNAs, with 20 duplicate in the IRs. Compared with the prior cp genome of watercress deposited in GenBank, 21 single nucleotide polymorphisms (SNPs) and 27 indels were identified, mainly located in noncoding sequences. A total of 49 repeat structures and 71 simple sequence repeats (SSRs) were detected. Codon usage showed a bias for A/T-ending codons in the cp genome of watercress. Moreover, 45 RNA editing sites were predicted in 16 genes, all for C-to-U transitions. A comparative plastome study with Cardamineae species revealed a conserved gene order and high similarity of protein-coding sequences. Analysis of the Ka/Ks ratios of Cardamineae suggested positive selection exerted on the ycf2 gene in watercress, which might reflect specific adaptations of watercress to its particular living environment. Phylogenetic analyses based on complete cp genomes and common protein-coding genes from 56 species showed that the genus Nasturtium was a sister to Cardamine in the Cardamineae tribe. Our study provides valuable resources for future evolution, population genetics and molecular biology studies of watercress.

BcMAF2 plays a key role in flowering regulation by controlling BcTEM1, BcSOC1 and BCSPL15 in Pak-choi. Flowering is a key event in the life cycle of plants. Flowering time shows an extensive variation from different Pak-choi (Brassica rapa ssp. chinensis) cultivars. However, the regulation mechanism of flowering in Pak-choi remains rarely known. In this study, a systematic identification and functional analysis of a Pak-choi MADS Affecting Flowering (MAF) gene, BcMAF2, was carried out. BcMAF2 encoded a protein containing a conserved MADS-box domain, which was localized in the nucleus. QPCR analysis indicated that the expression of BcMAF2 was higher in the leaves and flowers. Overexpression of BcMAF2 in Arabidopsis showed that BcMAF2 repressed flowering, which was further confirmed by silencing endogenous BcMAF2 in Pak-choi. In addition, Tempranillo 1 (TEM1) expression was up-regulated and MAF2 expression was down-regulated in the BcMAF2-overexpressing Arabidopsis. The expression of BcMAF2 and BcTEM1 was down-regulated in BcMAF2-silencing Pak-choi plants. The yeast one-hybrid, dual luciferase and qPCR results revealed that BcMAF2 protein could directly bind to BcTEM1 promoter and activate its expression, which was not reported in Arabidopsis. Meanwhile, a self-inhibition was found in BcMAF2. Taken together, this work suggested that BcMAF2 could repress flowering by directly activating BcTEM1.

Antimicrobial efficacy, which is central to many aspects of medicine, is being rapidly eroded by bacterial resistance. Since new resistance can be induced by antimicrobial action, highly lethal agents that rapidly reduce bacterial burden during infection should help restrict the emergence of resistance. To improve lethal activity, recent work has focused on toxic reactive oxygen species (ROS) as part of the bactericidal activity of diverse antimicrobials. We report that when Escherichia coli was subjected to antimicrobial stress and the stressor was subsequently removed, both ROS accumulation and cell death continued to occur. Blocking ROS accumulation by exogenous mitigating agents slowed or inhibited poststressor death. Similar results were obtained with a temperature-sensitive mutational inhibition of DNA replication. Thus, bacteria exposed to lethal stressors may not die during treatment, as has long been thought; instead, death can occur after plating on drug-free agar due to poststress ROS-mediated toxicity. Examples are described in which ( i ) primary stress-mediated damage was insufficient to kill bacteria due to repair; ( ii ) ROS overcame repair (i.e., protection from anti-ROS agents was reduced by repair deficiencies); and ( iii ) killing was reduced by anti-oxidative stress genes acting before stress exposure. Enzymatic suppression of poststress ROS-mediated lethality by exogenous catalase supports a causal rather than a coincidental role for ROS in stress-mediated lethality, thereby countering challenges to ROS involvement in antimicrobial killing. We conclude that for a variety of stressors, lethal action derives, at least in part, from stimulation of a self-amplifying accumulation of ROS that overwhelms the repair of primary damage.

Functional connectomes computed from fMRI provide a means to characterize individual differences in the patterns of BOLD synchronization across regions of the entire brain. Using four resting-state fMRI datasets with a wide range of ages, we show that individual differences of the functional connectome are stable across three months to three years. Medial frontal and frontoparietal networks appear to be both unique and stable, resulting in high ID rates, as did a combination of these two networks. We conduct analyses demonstrating that these results are not driven by head motion. We also show that the edges demonstrating the most individualized features tend to connect nodes in the frontal and parietal cortices, while edges contributing the least tend to connect cross-hemispheric homologs. Our results demonstrate that the functional connectome is stable across years and is not an idiosyncratic aspect of a specific dataset, but rather reflects stable individual differences in the functional connectivity of the brain.Research highlightsWhole-brain functional connectivity profiles obtained from four resting-state fMRI datasets are unique and stable across 3 months-3 years in adolescents, young adults, and older adultsMedial frontal and frontoparietal networks tended to be both unique and stableIndividual edges in the frontal and parietal cortices tended to be most discriminative of individual subjects

Aging and apolipoprotein E4 (ApoE4) can increase the risk of cognitive impairment and neurodegenerative disorders, including Alzheimer's disease (AD), and patients with type 2 diabetes mellitus are highly susceptible to cognitive dysfunction. Recent research has indicated that metformin, a prescribed drug for type 2 diabetes, may affect cognitive function; however, findings regarding its efficacy are largely controversial. The current study reported that a 5-mo metformin administration (300 mg/kg/d) starting at 13 mo old improved the spatial memory of ApoE3-target replacement (TR) mice, not ApoE4-TR mice. It found that in aged ApoE3-TR mice, metformin treatment, at a molecular level, inhibited AMPK activity, increased insulin signaling, and activated mammalian target of rapamycin signaling, resulting in an enhanced expression of postsynaptic proteins; but the response of the neuronal AMPK activity and insulin signaling to metformin was blunt in aged ApoE4-TR mice. Meanwhile, metformin treatment also increased the phosphorylation of tau in both ApoE3-TR and ApoE4-TR mice, implying that metformin may have side effects in human. These findings suggest that metformin can improve the cognitive performance of aged mice in an APOE genotype-dependent manner, which provides empirical insights into the clinical value of metformin for ApoE4- and age-related AD prevention and treatment.-Zhang, J., Lin, Y., Dai, X., Fang, W., Wu, X., Chen, X. Metformin treatment improves the spatial memory of aged mice in an APOE genotype-dependent manner.

YABBY proteins are widely distributed among different plant species and exhibit a higher degree of prevalence in angiosperms than in gymnosperms; abaxial cell fate in the lateral organs of plants is mainly determined by the functions of YABBY genes. However, to date, no scientific study has been conducted on the functions and responses of these genes under different forms of abiotic stresses in Pak-choi (Brassica rapa ssp. chinensis). In this study, we cloned and identified 12 YABBY family genes from Pak-choi. The evolutionary history of the YABBY genes was studied in nine species and showed that YABBY gene loss occurred during specific periods or in certain species during evolution. The putative YABBY family genes of Pak-choi were comprehensively analyzed by comparison with the corresponding orthologs in Arabidopsis and Chinese cabbage (Brassica rapa ssp. pekinensis) and classified into five subfamilies based on the specific protein domains and phylogenetic clades. A subcellular localization assay involving BcYABBY1b and BcYABBY2c confirmed that the BcYABBY proteins were localized in the nucleus. qRT-PCR data revealed that the BcYABBY genes are specifically expressed in distinct organs and developmental stages. Furthermore, the expression profiles of the BcYABBY genes were investigated under different hormone treatments and abiotic stress factors. In this study, we comprehensively identified and analyzed the YABBY gene family in the Pak-choi genome. Our data provide possible functional information regarding the involvement of BcYABBY genes in plant growth and development and in the response to abiotic stress and hormone treatments.

Background Cervical cancer caused by human papillomavirus (HPV) infections is one of the most common cancers affecting women worldwide. Current preventative HPV vaccines on the market are composed of HPV L1 protein produced either in the yeast such as Gardasil or in the insect cells such as Cervarix. The duration of efficacy and cross-protection remain highly desirable for the improvement of current prophylactic HPV vaccine. Given that HPV carries out infection and replicates in mammalian cells, L2 protein, which is not included in the current licensed vaccines, is included in the third generation of HPV vaccine in pursuing of providing broader prevention. We hypothesize that a virus-like particle (VLP) consisting of HPV L1 plus L2 proteins generated in mammalian cells will present conformations more closely to native HPV, thus it will provide more durable and broader efficacy of prevention. Methods We took advantage of 293TT cells to produce VLP containing L1 and L2 proteins of HPV16 and HPV18, respectively. Results VLP particles of uniformed size and morphology were observed, and potent and broadly neutralizing antibodies were induced in mice and rabbits. In addition, compared to bivalent HPV vaccine of Cervarix, our HPV L1-L2 VLPs elicited higher titer of anti-sera, and the anti-sera also presented comparable neutralization potency against HPV16 and HPV18 infections even a much less potent adjuvant was used in our case. Conclusion Our VLPs were capable of eliciting stronger and more broadly neutralizing activities against various HPV subtypes and were potential candidate HPV vaccines.