Cloning, Expression, and Functional Characterization of Two Highly Efficient Flavonoid-di-O-glycosyltransferases ZmUGT84A1 and ZmUGT84A2 from Maize (Zea mays L.).

The maize (Zea mays L.) glycosyltransferase family 1 comprises many uridine diphosphate glycosyltransferase (UGT) members. However, UGT activities and biochemical functions have seldom been revealed. In this study, the genes of two flavonoid di-O-glycosyltransferases ZmUGT84A1 and ZmUGT84A2 were cloned from maize plant and expressed in Escherichia coli. Phylogenetic analysis showed that the two enzymes were homologous to AtUGT84A1 and AtUGT84A3. The two recombinant enzymes showed a high conversion rate of luteolin to its glucosides, mainly 4',7-di-O-glucoside and minorly 3',7-di-O-glucoside in two-step glycosylation reactions in vitro. Moreover, the recombinant ZmUGT84A1 and ZmUGT84A2 had a broad substrate spectrum, converting eriodictyol, naringenin, apigenin, quercetin, and kaempferol to monoglucosides and diglucosides. The highly efficient ZmUGT84A1 and ZmUGT84A2 may be used as a tool for the effective synthesis of various flavonoid O-glycosides and as markers for crop breeding to increase O-glycosyl flavonoid content in food.

Norcantharidin inhibits the malignant progression of cervical cancer by inducing endoplasmic reticulum stress.

The antitumor effect of norcantharidin (NCTD) has been widely reported. However, whether NCTD can inhibit cervical cancer remains unknown. In the present study, it was shown that NCTD inhibited the viability of cervical cancer cells and caused cell cycle arrest in a concentration­dependent manner. Further analysis revealed that the NCTD­induced reduction in cell viability could be reversed by the inhibitor of apoptosis z­VAD­FMK and by the inhibitor of endoplasmic reticulum (ER) stress, 4­phenylbutyric acid (4­PBA). Additionally, NCTD led to the accumulation of reactive oxygen species as well as a decrease in the mitochondrial membrane potential in cervical cancer cells, whereas 4­PBA pre­treatment attenuated these alterations. In addition, NCTD increased the expression of the apoptosis­related proteins Bip, activating transcription factor (ATF) 4 and C/EBP homologous protein in a concentration­dependent manner. Moreover, NCTD significantly increased the expression of the ER stress­related signaling molecules protein kinase R­like ER kinase, inositol­requiring enzyme 1 and ATF6, but 4­PBA abolished these effects. In vivo experiments showed that NCTD significantly inhibited the growth of subcutaneous tumors in mice. Additionally, the expression of ER stress­related molecules and apoptosis­related proteins increased significantly after NCTD treatment. In conclusion, NCTD induces apoptosis by activating ER stress and ultimately curtails the progression of cervical cancer.

Efficient and genotype independent maize transformation using pollen transfected by DNA-coated magnetic nanoparticles.

Current gene delivery methods for maize are limited to specific genotypes and depend on time-consuming and labor-intensive tissue culture techniques. Here, we report a new method to transfect maize that is culture-free and genotype independent. To enhance efficiency of DNA entry and maintain high pollen viability of 32%-55%, transfection was performed at cool temperature using pollen pretreated to open the germination aperture (40%-55%). Magnetic nanoparticles (MNPs) coated with DNA encoding either red fluorescent protein (RFP), ß-glucuronidase gene (GUS), enhanced green fluorescent protein (EGFP) or bialaphos resistance (bar) was delivered into pollen grains, and female florets of maize inbred lines were pollinated. Red fluorescence was detected in 22% transfected pollen grains, and GUS stained 55% embryos at 18 d after pollination. Green fluorescence was detected in both silk filaments and immature kernels. The T1 generation of six inbred lines showed considerable EGFP or GUS transcripts (29%-74%) quantitated by polymerase chain reaction, and 5%-16% of the T1 seedlings showed immunologically active EGFP or GUS protein. Moreover, 1.41% of the bar transfected T1 plants were glufosinate resistant, and heritable bar gene was integrated into the maize genome effectively as verified by DNA hybridization. These results demonstrate that exogenous DNA could be delivered efficiently into elite maize inbred lines recalcitrant to tissue culture-mediated transformation and expressed normally through our genotype-independent pollen transfection system.

Natural variation of ZmCGT1 is responsible for isoorientin accumulation in maize silk.

Maize (Zea mays L.) silk contains high levels of flavonoids and is widely used to promote human health. Isoorientin, a natural C-glycoside flavone abundant in maize silk, has attracted considerable attention due to its potential value. Although different classes of flavonoid have been well characterized in plants, the genes involved in the biosynthesis of isoorientin in maize are largely unknown. Here, we used targeted metabolic profiling of isoorientin on the silks in an association panel consisting of 294 maize inbred lines. We identified the gene ZmCGT1 by genome-wide association analysis. The ZmCGT1 protein was characterized as a 2-hydroxyflavanone C-glycosyltransferase that can C-glycosylate 2-hydroxyflavanone to form flavone-C-glycoside after dehydration. Moreover, ZmCGT1 overexpression increased isoorientin levels and RNA interference-mediated ZmCGT1 knockdown decreased accumulation of isoorientin in maize silk. Further, two nucleotide polymorphisms, A502C and A1022G, which led to amino acid changes I168L and E341G, respectively, were identified to be functional polymorphisms responsible for the natural variation in isoorientin levels. In summary, we identified the gene ZmCGT1, which plays an important role in isoorientin biosynthesis, providing insights into the genetic basis of the natural variation in isoorientin levels in maize silk. The identified favorable CG allele of ZmCGT1 may be further used for genetic improvement of nutritional quality in maize.

Isoorientin Affects Markers of Alzheimer's Disease via Effects on the Oral and Gut Microbiota in APP/PS1 Mice.

BACKGROUND: There is growing evidence of strong associations between the pathogenesis of Alzheimer's disease (AD) and dysbiotic oral and gut microbiota. Recent studies demonstrated that isoorientin (ISO) is anti-inflammatory and alleviates markers of AD, which were hypothesized to be mediated by the oral and gut microbiota. OBJECTIVES: We studied the effects of oral administration of ISO on AD-related markers and the oral and gut microbiota in mice. METHODS: Eight-month-old amyloid precursor protein/presenilin-1 (AP) transgenic male mice were randomly allocated to 3 groups of 15 mice each: vehicle (AP) alone or with a low dose of ISO (AP + ISO-L; 25 mg/kg) or a high dose of ISO (AP + ISO-H; 50 mg/kg). Age-matched wild-type (WT) C57BL/6 male littermates were used as controls. The 4 groups were treated intragastrically with ISO or sterilized ultrapure water for 2 months. AD-related markers in the brain, serum, colon, and liver were analyzed with immunohistochemical and histochemical staining, Western blotting, and ELISA. Oral and gut microbiotas were analyzed using 16S ribosomal RNA gene sequencing. RESULTS: The high-dose ISO treatment significantly decreased amyloid beta 42-positive deposition by 38.1% and 45.2% in the cortex and hippocampus, respectively, of AP mice (P < 0.05). Compared with the AP group, both ISO treatments reduced brain phospho-Tau, phosphor-p65, phosphor-inhibitor of NF-κB, and brain and serum LPS and TNF-α by 17.9%-72.5% and increased brain and serum IL-4 and IL-10 by 130%-210% in the AP + ISO-L and AP + ISO-H groups (P < 0.05). Abundances of 26, 25, and 23 microbial taxa in oral, fecal and cecal samples, respectively, were increased in both the AP + ISO-L and AP + ISO-H groups relative to the AP group [linear discriminant analysis (LDA) >3.0; P < 0.05]. Gram-negative bacteria, Alteromonas, Campylobacterales, and uncultured Bacteroidales bacterium were positively correlated (rho = 0.28-0.59; P < 0.05) with the LPS levels and responses of inflammatory cytokines. CONCLUSIONS: The microbiota-gut-brain axis is a potential mechanism by which ISO reduces AD-related markers in AP mice.

Jianqu fermentation with the isolated fungi significantly improves the immune response in immunosuppressed mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Jianqu, a classical formula of traditional Chinese medicine, is used clinically to treat symptoms like chill and fever headache, diarrhea and loss of appetite and act on patients with low immunity. However, the quality control of Jianqu fermentation is not well established, and its function in regulating the body's immunity still remains unclear. AIM OF THE STUDY: The present study firstly assesses the structure and diversity of fungal community during Jianqu fermentation and then investigates the immune regulating function of Jianqu extract in mouse model. MATERIALS AND METHOD: The high-throughput sequencing is conducted to analyze the diversity and distribution of fungal community during the fermentation process of Jianqu, and then fungi with a high frequency and relative abundance are isolated. The immunosuppressed mice are induced by using cyclophosphamide (CTX) and used to evaluate the immune regulating function of Jianqu extract from natural fermentation or directed fermentation, respectively. RESULTS: With the fermentation, the diversity and distribution of fungal community significantly changed. The number of OTU (operational taxonomic unit) was gradually decreased from 223 ± 1 in the early phase to 201 ± 11 in the middle phase and to 175 ± 32 in the later phase of Jianqu fermentation. Generally, in genus level, Millerozyma, Debaryomyces and Rhizomucor showed a significant increase and became dominant in the mid or later phase of fermentation, while the Aspergillus displayed a decrease following the fermentation. However, Saccharomycopsis is a dominate species in surveyed samples. Next, six fungi strains with a high frequency and relative abundance, including Saccharomycopsis fibuligera, Millerozyma farinose, Hyphopichia burtonii, Rhizomucor pusillus, Lichtheimia ramosa, and Monascus purpureus, are isolated successfully. Interestingly, directed fermentation for Jianqu with the six isolated fungi strains could achieve similar morphological characteristics with the natural fermentation. Consistently, Jianqu extract from directed fermentation demonstrated a similar therapeutic effect on immune response as that of naturally fermented Jianqu. CONCLUSIONS: We firstly showed the significant change of structural profiles of fungal communities during Jianqu fermentation, and successfully isolated six dominate fungi strains in Jianqu. Interestingly, directed fermentation for Jianqu with these isolated strains could achieve a similar morphological characteristics and immune-modulating function as natural fermentation. It was suggested that Jianqu fermentation with functional fungi instead of natural microbes provide a new approach for the improvement of the production and quality control of the traditional Chinese medicine of Jianqu.

Identifying key regulatory genes of maize root growth and development by RNA sequencing.

Root system architecture (RSA), the spatio-temporal configuration of roots, plays vital roles in maize (Zea mays L.) development and productivity. We sequenced the maize root transcriptome of four key growth and development stages: the 6th leaf stage, the 12th leaf stage, the tasseling stage and the milk-ripe stage. Differentially expressed genes (DEGs) were detected. 81 DEGs involved in plant hormone signal transduction pathway and 26 transcription factor (TF) genes were screened. These DEGs and TFs were predicted to be potential candidate genes during maize root growth and development. Several of these genes are homologous to well-known genes regulating root architecture or development in Arabidopsis or rice, such as, Zm00001d005892 (AtERF109), Zm00001d027925 (AtERF73/HRE1), Zm00001d047017 (AtMYC2, OsMYC2), Zm00001d039245 (AtWRKY6). Identification of these key genes will provide a further understanding of the molecular mechanisms responsible for maize root growth and development, it will be beneficial to increase maize production and improve stress resistance by altering RSA traits in modern breeding.

Effects of aspirin on hepatocellular carcinoma and its potential molecular mechanism.

PURPOSE: To explore the effects of aspirin (ASP) on the proliferation and apoptosis of HepG2 hepatocellular carcinoma (HCC) cells via the Wnt/ß-catenin signaling pathway. METHODS: Human HCC cells were cultured and treated with ASP at different concentrations. Cell proliferation was determined with cell counting kit-8 (CCK-8) and colony formation, and the rate of apoptosis was measured by flow cytometry. Western blotting (WB) and quantitative polymerase chain reaction (qRT-PCR) assays were used to assess the changes in the expression levels of related proteins. RESULTS: ASP showed a time-and concentration-depented inhibitory effect on HepG2 cell proliferation. The number of colonies formed in ASP-treated HCC cells was significantly lower than in control cells. For HCC cells treated with ASP, the apoptosis rate enhanced with the increase of ASP concentration. The expression levels of TCF4 and LEF1, key molecules of the Wnt/ß-catenin signaling pathway, were lowered in HCC cells treated with 4 mM ASP, and the nuclear translocation of ß-catenin was weakened. The ß-catenin activator exerted a negative influence on the anticancer effect of ASP. CONCLUSIONS: ASP inhibits the proliferation and promotes the apoptosis of HCC cells through the Wnt/ß-catenin signaling pathway.

Analyses of the Complete Genome Sequence of the Strain Bacillus pumilus ZB201701 Isolated from Rhizosphere Soil of Maize under Drought and Salt Stress.

Bacillus pumilus ZB201701 is a rhizobacterium with the potential to promote plant growth and tolerance to drought and salinity stress. We herein present the complete genome sequence of the Gram-positive bacterium B. pumilus ZB201701, which consists of a linear chromosome with 3,640,542 base pairs, 3,608 protein-coding sequences, 24 ribosomal RNAs, and 80 transfer RNAs. Genome analyses using bioinformatics revealed some of the putative gene clusters involved in defense mechanisms. In addition, activity analyses of the strain under salt and simulated drought stress suggested its potential tolerance to abiotic stress. Plant growth-promoting bacteria-based experiments indicated that the strain promotes the salt tolerance of maize. The complete genome of B. pumilus ZB201701 provides valuable insights into rhizobacteria-mediated salt and drought tolerance and rhizobacteria-based solutions for abiotic stress in agriculture.

Structural and functional profiles of the gut microbial community in polycystic ovary syndrome with insulin resistance (IR-PCOS): a pilot study.

Polycystic ovary syndrome (PCOS) is a complex endocrine and metabolic disorder that affects 9-21% of reproductive-aged women. Affected women frequently display obesity, insulin resistance, and inflammation. Altered gut microbial community has been reported in PCOS and obese PCOS patients. However, the profile of the gut microbial community in insulin resistant PCOS (IR-PCOS) patients still remains unknown. In this study, next-generation sequencing based on the 16S rRNA gene was used to compare the gut microbial composition of women with IR-PCOS (n = 9, PCOS with insulin resistance), NIR-PCOS (n = 8, PCOS alone) and healthy controls (n = 8, HC). We assessed that the composition of the gut microbial communities in NIR-PCOS and IR-PCOS patients were significantly altered. The family Bacteroidaceae was prolific in the NIR-PCOS group and reached its highest level in the IR-PCOS group, while the Prevotellaceae dramatically decreased in PCOS patients, especially in the IR-PCOS group. Subsequent correlation analysis revealed that the increased clinical parameter levels, including insulin resistance, sex-hormones and inflammation, were positively associated with the abundance of Bacteroidaceae, but negatively associated with that of Prevotellaceae. In addition, IR-PCOS patients also displayed a significant difference in their amounts of Ruminococcaceae and Lachnospiraceae when compared to the NIR-PCOS group. Moreover, the functional prediction from PICRUSt revealed that 73 pathways are significantly changed in the gut microbial communities of PCOS patients. Specifically, 21 metabolism-associated pathways, including the steroid hormone biosynthesis and lipopolysaccharide biosynthesis pathways, are obviously changed in IR-PCOS when compared to NIR-PCOS and HC groups. Taking this into consideration, our present study suggests that the dysbiosis of gut microbial communities occurred most notably in IR-PCOS patients, and the difference in gut dysbiosis profile between the IR-PCOS and NIR-PCOS should be considered in clinical treatment for PCOS patients and future drugs development.

Effective Biodegradation of Aflatoxin B1 Using the Bacillus licheniformis (BL010) Strain.

Aflatoxin B1 (AFB1), a pollutant of agricultural products, has attracted considerable attention in recent years, due to its potential impact on health. In the present study, Bacillus licheniformis (BL010) was demonstrated to efficiently degrade AFB1, reducing over 89.1% of the toxin content within 120 h. A crude enzyme solution of BL010 exhibited the highest degradation level (97.3%) after three induction periods. However, uninduced BL010 bacteria was not capable of reducing AFB1. Furthermore, high performance liquid chromatography (HPLC) analysis showed that while a cell-free extract caused a significant decrease in AFB1 content (93.6%, p < 0.05), cell culture fluid treatment did not significantly degrade AFB1. The biotransformation products of AFB1 were detected and further identified by quadrupole time-of-flight liquid chromatography⁻mass spectrometry (LC-Q-TOF/MS); these corresponded to a molecular formula of C12H14O4. A sequence analysis of whole BL010 genes with a bioinformatics approach identified the secondary structures of two degrading enzymes (Chia010 and Lac010), providing an important basis for subsequent homology modeling and functional predictions.

[Preparation of multivalent egg yolk IgY antibodies against Helicobacter pylori].

Objective To prepare a specific multivalent immunoglobulin of yolk (IgY) against multi-virulence factors of Helicobacter pylori (Hp). Methods The purified recombinant BabA2, UerB and FlaA of Hp were mixed equally with adjuvant, and then used to immunize egg-laying hens to produce specific multivalent IgY. Next, the biological characteristics of purified IgY were analyzed by SDS-PAGE, Western blot and ELISA. To analyze the inhibition effects of multivalent IgY on the growth of Hp in vitro, Hp was co-cultured with different final concentration of IgY (1, 5, 10) mg/mL in the presence or absence of antibiotics. Results After immunization, the titer of egg yolk IgY reached up to 1:150 000. Furthermore, the bacteriostasis assay showed that the multivalent IgY had a significant inhibitory effect on the growth of Hp at 5 mg/mL, and the combination with amoxicillin could further inhibited the growth of Hp. Conclusion The specific multivalent IgY against Hp is successfully prepared.

Distribution of Four Bioactive Flavonoids in Maize Tissues of Five Varieties and Correlation with Expression of the Biosynthetic Genes.

Flavonoids are characteristic in maize and have diverse biological functions. C-Glycosylflavones are neuroprotective against ß-amyloid-induced tau hyperphosphorylation and neurotoxicity in SH-SY5Y cells, which is relevant to Alzheimer's disease prevention and treatment. The content of the flavonoids eriodictyol, luteolin, isoorientin, and maysin varied in pollens, silks, tassels, and seeds among five maize varieties. Eriodictyol content was high (51-322 ng/g dw) in pollens, while luteolin content was low (0.2-106 ng/g dw) in all four tissues. The isoorientin content was approximately 3- to 10-fold greater than eriodictyol in pollens and tassels, particularly in the hybrid M1 and sweet corn M5 varieties. Maysin content was high in most silks and tassels. The differential expression of five genes involved in the maysin biosynthesis correlated well with the profiles of the four flavonoids among tissues and varieties. The present study offers valuable data for maize breeding and the use of maize flavonoids as functional food components.

The complete genome sequence of Bacillus halotolerans ZB201702 isolated from a drought- and salt-stressed rhizosphere soil.

Bacillus halotolerans is a rhizobacterium with the potential to promote plant growth and tolerance to drought and salinity stress. Here, we present the complete genome sequence of B. halotolerans ZB201702, which consists of 4,150,000 bp in a linear chromosome, including 3074 protein-coding sequences, 30 rRNAs, and 85 tRNAs. Genome analysis revealed many putative gene clusters involved in defense mechanisms. Activity analysis of the strain under salt and simulated drought stress suggests tolerance to abiotic stresses. The complete genome information of B. halotolerans ZB201702 could provide valuable insights into rhizobacteria-mediated plant salt and drought tolerance and rhizobacteria-based solutions for abiotic stress agriculture.

Complete genome sequence of Bacillus licheniformis BL-010.

The biodegradation of Aflatoxin B1 (AFB1) is an industry of increasing importance. Bacillus licheniformis BL-010 was isolated from the aflatoxin contaminated corn feed storage, and was shown to degrade AFB1 efficiently. Here we present the complete genome sequence of BL-010, the genome comprises 4,287,714 bp in a circular chromosome with a GC content of 46.12% and contains genes encoding AFB1 degrading enzymes. The genome sequence displayed that this strain contains genes involved in production of laccase, aromatic ring-opening dioxygenase which could detoxify AFB1. Complete genome sequence of the strain BL-010 can further provide the genomic basis for the biotechnological application of strain BL-010 as an effective way to degrade AFB1.

Isolation, structural analysis, and expression characteristics of the maize nuclear factor Y gene families.

NUCLEAR FACTOR-Y (NF-Y) has been shown to play an important role in growth, development, and response to environmental stress. A NF-Y complex, which consists of three subunits, NF-YA, NF-YB, and, NF-YC, binds to CCAAT sequences in a promoter to control the expression of target genes. Although NF-Y proteins have been reported in Arabidopsis and rice, a comprehensive and systematic analysis of ZmNF-Y genes has not yet been performed. To examine the functions of ZmNF-Y genes in this family, we isolated and characterized 50 ZmNF-Y (14 ZmNF-YA, 18 ZmNF-YB, and 18 ZmNF-YC) genes in an analysis of the maize genome. The 50 ZmNF-Y genes were distributed on all 10 maize chromosomes, and 12 paralogs were identified. Multiple alignments showed that maize ZmNF-Y family proteins had conserved regions and relatively variable N-terminal or C-terminal domains. The comparative syntenic map illustrated 40 paralogous NF-Y gene pairs among the 10 maize chromosomes. Microarray data showed that the ZmNF-Y genes had tissue-specific expression patterns in various maize developmental stages and in response to biotic and abiotic stresses. The results suggested that ZmNF-YB2, 4, 8, 10, 13, and 16 and ZmNF-YC6, 8, and 15 were induced, while ZmNF-YA1, 3, 4, 6, 7, 10, 12, and 13, ZmNF-YB15, and ZmNF-YC3 and 9 were suppressed by drought stress. ZmNF-YA3, ZmNF-YA8 and ZmNF-YA12 were upregulated after infection by the three pathogens, while ZmNF-YA1 and ZmNF-YB2 were suppressed. These results indicate that the ZmNF-Ys may have significant roles in the response to abiotic and biotic stresses.

LncRNA HOTAIR controls the expression of Rab22a by sponging miR-373 in ovarian cancer.

Increasing evidence suggests that the long non-coding RNA, HOX transcript antisense intergenic RNA (HOTAIR) is widely involved in the progression and metastasis of cancer. However, the specific role of HOTAIR in ovarian carcinogenesis remains to be fully elucidated. In the present study, the levels of HOTAIR were detected in 30 paired cancer and noncancer tissues using reverse transcription-quantitative polymerase chain reaction analysis. The effect of HOTAIR on the ovarian cancer cells was examined by overexpression or small interfering RNA interference experiments. To examine the competitive endogenous RNA (ceRNAs) mechanism, a luciferase reporter assay was used. In patients with ovarian cancer, HOTAIR was significantly upregulated. Furthermore, the upregulation of HOTAIR increased the proliferation, migration and invasion of ovarian cancer cells. By contrast, the knockdown of HOTAIR repressed cell invasion and viability. HOTAIR functioned as a ceRNA, and acted as a sink for microRNA (miR)­373, thereby regulating the expression of Rab22a. The upregulation of HOTAIR contributed to the malignant progression of ovarian cancer cells. Therefore, the positive regulation between HOTAIR and Rab22a can be partially attributed to the ceRNA regulatory network through miR-373.

The prognostic role and reduced expression of FOXJ2 in human hepatocellular carcinoma.

The current study aimed to investigate the potential role of the FOXJ2 (forkhead box J2) protein in the pathology of hepatocellular carcinoma (HCC). Western blotting was performed to determine the expression levels of FOXJ2 in HCC tissues and HCC cells. Specimens from 110 patients with HCC undergoing hepatic resection were evaluated for FOXJ2 expression using an immunohistochemical assay. The correlation between FOXJ2 expression and clinicopathological factors of the patients was determined by statistical analysis to determine the prognostic merit of FOXJ2 expression in HCC. The detailed involvement of FOXJ2 in the regulation of HCC proliferation was further investigated using FOXJ2­targeting small interfering RNA (siRNA). FOXJ2 protein was identified to be significantly downregulated in HCC tissues compared with adjacent normal liver tissues. Immunohistochemical analysis demonstrated that the expression of FOXJ2 was negatively correlated with Ki­67 levels in HCC specimens (r=­0.679, P<0.001). Furthermore, statistical analysis indicated FOXJ2 expression was significantly associated with histological differentiation (P=0.005), the size of largest tumor (P=0.002) and metastasis (P=0.036). Using Kaplan­Meier analysis, it was demonstrated that high FOXJ2 expression levels predicted significantly improved patient survival rates compared with low FOXJ2 expression levels (P<0.001). In addition, it was observed that interference of FOXJ2 expression using siRNA oligos led to the promotion of proliferation of HepG2 cells. FOXJ2 was markedly downregulated in HCC tissues. The expression of FOXJ2 was correlated with tumor size, histological differentiation and metastasis. Low expression levels of FOXJ2 predicted poor prognosis for patients with HCC, suggesting that FOXJ2 may be a candidate prognostic marker of HCC. Depletion of FOXJ2 caused the promotion of HCC cell proliferation, implicating that FOXJ2 may serve an inhibitory role in the regulation of HCC cell proliferation.

Isolation, structural analysis, and expression characteristics of the maize TIFY gene family.

TIFY, previously known as ZIM, comprises a plant-specific family annotated as transcription factors that might play important roles in stress response. Despite TIFY proteins have been reported in Arabidopsis and rice, a comprehensive and systematic survey of ZmTIFY genes has not yet been conducted. To investigate the functions of ZmTIFY genes in this family, we isolated and characterized 30 ZmTIFY (1 TIFY, 3 ZML, and 26 JAZ) genes in an analysis of the maize (Zea mays L.) genome in this study. The 30 ZmTIFY genes were distributed over eight chromosomes. Multiple alignment and motif display results indicated that all ZmTIFY proteins share two conserved TIFY and Jas domains. Phylogenetic analysis revealed that the ZmTIFY family could be divided into two groups. Putative cis-elements, involved in abiotic stress response, phytohormones, pollen grain, and seed development, were detected in the promoters of maize TIFY genes. Microarray data showed that the ZmTIFY genes had tissue-specific expression patterns in various maize developmental stages and in response to biotic and abiotic stresses. The results indicated that ZmTIFY4, 5, 8, 26, and 28 were induced, while ZmTIFY16, 13, 24, 27, 18, and 30 were suppressed, by drought stress in the maize inbred lines Han21 and Ye478. ZmTIFY1, 19, and 28 were upregulated after infection by three pathogens, whereas ZmTIFY4, 13, 21, 23, 24, and 26 were suppressed. These results indicate that the ZmTIFY family may have vital roles in response to abiotic and biotic stresses. The data presented in this work provide vital clues for further investigating the functions of the genes in the ZmTIFY family.

Isolation, structural analysis, and expression characteristics of the maize (Zea mays L.) hexokinase gene family.

Hexokinases (HXKs, EC 2.7.1.1) play important roles in metabolism, glucose (Glc) signaling, and phosphorylation of Glc and fructose and are ubiquitous in all organisms. Despite their physiological importance, the maize HXK (ZmHXK) genes have not been analyzed systematically. We isolated and characterized nine members of the ZmHXK gene family which were distributed on 3 of the 10 maize chromosomes. A multiple sequence alignment and motif analysis revealed that the maize ZmHXK proteins share three conserved domains. Phylogenetic analysis revealed that the ZmHXK family can be divided into four subfamilies. We identified putative cis-elements in the ZmHXK promoter sequences potentially involved in phytohormone and abiotic stress responses, sugar repression, light and circadian rhythm regulation, Ca(2+) responses, seed development and germination, and CO2-responsive transcriptional activation. To study the functions of maize HXK isoforms, we characterized the expression of the ZmHXK5 and ZmHXK6 genes, which are evolutionarily related to the OsHXK5 and OsHXK6 genes from rice. Analysis of tissue-specific expression patterns using quantitative real time-PCR showed that ZmHXK5 was highly expressed in tassels, while ZmHXK6 was expressed in both tassels and leaves. ZmHXK5 and ZmHXK6 expression levels were upregulated by phytohormones and by abiotic stress.