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Additional file 7:Primers used for stability evaluation of housekeeping gene expression. (DOC 31 KB)

Additional file 6:The relative expression levels of 31 ZmARFs in three tissues of maize inbred line B73. Leaves (8-day-old seedling), roots (8-day-old seedling) and embryos (15d after pollination) of maize inbred line B73 were used for real-time RT-PCR analysis. (DOC 78 KB)

Additional file 4:miR160, 167 and TAS3 target site prediction for ZmARF genes. (DOC 37 KB)

Additional file 9:Primers used for expression study of ZmARF genes. (DOC 50 KB)

Electrospun nanofibers have shown great potential for removal of contaminants from aqueous solutions owing to their large specific surface area, high porosity, easy modification, and good compatibility with other functional materials. To date, hundreds of polymers have been successfully fabricated into ultrafine nanofibers by electrospinning for adsorption purposes. In this review, the recent progress in electrospun adsorbents is summarized, and special attention is paid to the critical parameters for electrospinning of ultrafine fibers, commonly used polymers and fillers, as well as the adsorption capacities of electrospun nanocomposites for various water contaminants removal. First, various parameters including solution viscosity, polymer molecular weight, applied voltage, tip-to-collector distance, and flow rate for the fabrication of defectless and uniform nanofibers are illustrated. Electrospun fibrous adsorbents are then categorized according to their compositions, such as single polymer adsorbents, polymer composite adsorbents, and filler-added polymeric adsorbents, and their adsorption capacities towards specific contaminants are compared with discussions on the adsorption mechanism. Finally, a conclusion on recent achievements and current challenges is stated, and prospects associated with the applications of electrospun nanofibers in water treatment are provided.

Seed oils are important natural resources used in the processing and preparation of food. Histone modifications represent key epigenetic mechanisms that regulate gene expression, plant growth and development. However, histone modification events during fatty acid (FA) biosynthesis are not well understood. Here, we demonstrate that a mutation of the histone acetyltransferase GCN5 can decrease the ratio of α-linolenic acid (ALA) to linoleic acid (LA) in seed oil. Using RNA-Seq and ChIP assays, we identified FAD3, LACS2, LPP3 and PLAIIIβ as the targets of GCN5. Notably, the GCN5-dependent H3K9/14 acetylation of FAD3 determined the expression levels of FAD3 in Arabidopsis thaliana seeds, and the ratio of ALA/LA in the gcn5 mutant was rescued to the wild-type levels through the overexpression of FAD3. The results of this study indicated that GCN5 modulated FA biosynthesis by affecting the acetylation levels of FAD3. We provide evidence that histone acetylation is involved in FA biosynthesis in Arabidopsis seeds and might contribute to the optimization of the nutritional structure of edible oils through epigenetic engineering.

One QTL qLRI4 controlling leaf rolling index on chromosome 4 was finely mapped, and ZmOCL5, a member of the HD-Zip class IV genes, is likely a candidate. Leaf rolling is an important agronomic trait related to plant architecture that can change the light condition and photosynthetic efficiency of the population. Here, we isolated one EMS-induced mutant in Chang7-2 background with extreme abaxial rolling leaf, named abrl1. Histological analysis showed that the increased number and area of bulliform cells may contribute to abaxial rolling leaf in abrl1. The F2 and F2:3 populations derived from Wu9086 with flat leaves and abrl1 were developed to map abrl1. Non-Mendelian segregation of phenotypic variation was observed in these populations and five genomic regions controlling the leaf rolling index (LRI) were identified, which could be due to the phenotypic difference between Chang7-2 and Wu9086. Moreover, one major QTL qLRI4 on chromosome 4 was further validated and finely mapped to a genetic interval between InDel13 and InDel10, with a physical distance of approximately 277 kb using NIL populations, among which one 602-bp insertion was identified in the promoter region of HD-Zip class IV gene Zm00001d049443 (named as ZmOCL5) of abrl1 compared with wild-type Chang7-2. Remarkably, the 602-bp InDel was associated with LRI in an F2 population developed by crossing abrl1 mutant and its wild-type. In addition, the 602-bp insertion increased ZmOCL5 promoter activity and expression. Haplotype analysis demonstrated that the 602-bp insertion was a rare mutation event. Taken together, we propose that the rolled leaf in the abrl1 mutant may be partially attributed to the 602-bp insertion, which may be an attractive target for the genetic improvement of LRI in maize.

Objective To evaluate the efficacy of daily low-dose sildenafil for the treatment of nonulcer interstitial cystitis (IC) in women. Patients and Methods Forty-eight women with a clinical diagnosis of IC from 3 medical centers were randomly assigned to treatment with daily low-dose sildenafil (25 mg, n = 24) or placebo (n = 24) for 3 months. The O'Leary-Sant IC symptom and problem indices, visual analog scale scores, and a micturition diary with the interval of micturition, the frequency of nocturia, and urgency episodes were recorded before treatment, every 2 weeks after the treatment until 3 months. Patient Overall Rating of Improvement in Symptoms was assessed and regarded as effective when the value was above 50%. Results The IC symptom and problem indices scores and urodynamic index were significantly improved in sildenafil treatment group as compared with placebo group and baselines at week 4, 6, 8, 10, and 12, as well as 3 months after treatment (P

Cytokinin signaling is vital for plant growth and development which function via the two-component system (TCS). As one of the key component of TCS, transmembrane histidine kinases (HK) are encoded by a small gene family in plants. In this study, we focused on expression and functional analysis of cytokinin receptor-like HK genes (ZmHK) in maize. Firstly, bioinformatics analysis revealed that seven cloned ZmHK genes have different expression patterns during maize development. Secondly, ectopic expression by CaMV35S promoter in Arabidopsis further revealed that functional differentiation exists among these seven members. Among them, the ZmHK1a2-OX transgenic line has the lowest germination rate in the dark, ZmHK1-OX and ZmHK2a2-OX can delay leaf senescence, and seed size of ZmHK1-OX, ZmHK1a2-OX, ZmHK2-OX, ZmHK3b-OX and ZmHK2a2-OX was obviously reduced as compared to wild type. Additionally, ZmHK genes play opposite roles in shoot and root development; all ZmHK-OX transgenic lines display obvious shorter root length and reduced number of lateral roots, but enhanced shoot development compared with the wild type. Most notably, Arabidopsis response regulator ARR5 gene was up-regulated in ZmHK1-OX, ZmHK1a2-OX, ZmHK2-OX, ZmHK3b-OX and ZmHK2a2-OX as compared to wild type. Although the causal link between ZmHK genes and cytokinin signaling pathway is still an area to be further elucidated, these findings reflected that the diversification of ZmHK genes expression patterns and functions occurred in the course of maize evolution, indicating that some ZmHK genes might play different roles during maize development.

Purpose We investigated the prognostic potential and regulatory mechanism of microRNA-500 (miR-500), and human gene of tissue factor pathway inhibitor (TFPI) in prostate cancer. Methods MiR-500 expression was assessed by qRT-PCR in prostate cancer cell lines and primary tumors. Cancer patients’ clinicopathological factors and overall survival were analyzed according to endogenous miR-500 level. MiR-500 was downregulated in DU145 and VCaP cells. Its effect on prostate cancer proliferation, invasion in vitro, and tumorigenicity in vivo, were probed. Possible downstream target of miR-500, TFPI was assessed by luciferase assay and qRT-PCR in prostate cancer cells. In miR-500-downregulated DU145 and VCaP cells, TFPI was silenced to see whether it was directly involved in the regulation of miR-500 in prostate cancer. TFPI alone was either upregulated or downregulated in DU145 and VCaP cells. Their effect on prostate cancer development was further evaluated. Results MiR-500 is upregulated in both prostate cancer cells and primary tumors. In prostate cancer patients, high miR-500 expression is associated with poor prognosis and overall survival. In DU145 and VCaP cells, miR-500 downregulation inhibited cancer proliferation, invasion in vitro, and explant growth in vivo. TFPI was verified to be associated with miR-500 in prostate cancer. Downregulation of TFPI reversed anti-cancer effects of miR-500 downregulation in prostate cancer cells. However, neither TFPI upregulation nor downregulation alone had any functional impact on prostate cancer development. Conclusion MiR-500 may be a potential biomarker and molecular target in prostate cancer. TFPI may conditionally regulate prostate cancer in miR-500-downregualted prostate cancer cells.

Detailed information of the five environments in which the materials were evaluated. (DOC 630 kb)

Comparison of QTLs for PH (plant height) and EH (ear height) with QTLs for ear-weight-related traits in our previous study. (DOC 3075 kb)

Heterosis plays a decisive role in maize production worldwide, but its genetic basis remains unclear. In this study, we explored heterosis for ear-weight (EW)-related traits using a North Carolina Experiment III design (Design III) population derived from the elite maize hybrid Zhengdan 958. Quantitative trait loci (QTL) analysis was conducted based on phenotypic data collected from five environments using a high-density linkage map that consisted of 905 single nucleotide polymorphisms (SNP). A total of 38 environmentally stable QTLs were detected, and the numbers for the Z1 and Z2 populations were 18 and 20, respectively. All environmentally stable QTLs for Z2 were characterized by the overdominance effect (OD), which indicated that overdominance was one of the most important contributors to the heterosis of EW-related traits. Consistent with the significant positive correlations between EW-related traits, nine genomic regions with overlapped QTLs for different traits were found and were located on chromosomes 1(1), 3 (2), 4 (3), 7 (1), 8 (1) and 9 (1). Compared to previous reports, we found that the genomic regions for heterosis were not always congruent between different hybrids, which suggested that the combination of heterotic loci in different hybrids was genotype-dependent. Collectively, these data provided further evidence that the potential utilization of QTLs for heterosis may be feasible by pyramiding if we treat the QTLs as inherited units.

Objective To evaluate the efficacy of daily low-dose tadalafil therapy for overactive bladder (OAB) in women. Patients and Methods A total of 96 women with idiopathic OAB from 3 medical centers in the south of Zhejiang Province of China were randomly assigned to treatment with daily low-dose tadalafil (5 mg, n = 48) or placebo (n = 48) for 3 months. The Indevus Urgency Severity Scale, overactive bladder symptom score (Homma et al, 2006), and a 3-day micturition diary with frequency, incontinence, and urgency episodes were recorded and compared before the treatment, every 2 weeks following the treatment, and 3 months after the treatment. Uroflowmetry and transabdominal ultrasound were also conducted following the treatment to determine the maximum flow rate, voided volume, postvoid residual volume, total bladder capacity, and voiding efficiency. The patient's overall rating of improvement in symptoms was assessed as well. Results The overactive bladder symptom score significantly decreased, and the frequency, incontinence, and urgency episodes significantly improved in the tadalafil treatment group as compared with the placebo group and baselines at weeks 4, 6, 8, 10, and 12, as well as 3 months posttreatment ( P .05). In addition, voided volume and total bladder capacity obviously increased in the treatment group ( P .05). The Indevus Urgency Severity Scale decreased from week 4 to 3 months posttreatment in the treatment group ( P .05). No changes were found in the maximum flow rate, postvoid residual volume, and voiding efficiency. All adverse symptoms were mild to moderate. Conclusion Daily low-dose tadalafil is a considerable, well-tolerated, and effective treatment for OAB in women.

Highlight ZmABA8ox1b-mediated abscisic acid inactivation increases seed germination rate by promoting cell expansion in the maize hybrid B73/Mo17 compared with its parental inbred lines., Heterosis has been widely used in agriculture, but the underlying molecular principles are still largely unknown. During seed germination, we observed that maize (Zea mays) hybrid B73/Mo17 was less sensitive than its parental inbred lines to exogenous abscisic acid (ABA), and endogenous ABA content in hybrid embryos decreased more rapidly than in the parental inbred lines. ZmABA8ox1b, an ABA inactivation gene, was consistently more highly up-regulated in hybrid B73/Mo17 than in its parental inbred lines at early stages of seed germination. Moreover, ectopic expression of ZmABA8ox1b obviously promoted seed germination in Arabidopsis. Remarkably, microscopic observation revealed that cell expansion played a major role in the ABA-mediated maize seed germination heterosis, which could be attributed to the altered expression of cell wall-related genes.

Plants have developed complex signaling networks to regulate biochemical and physiological acclimation, environmental signals were perceived and transmitted to cellular machinery to activate adaptive responses. Here, a novel drought responsive histidine kinase gene was identified and designated as ZmHK9. Under normal conditions, ZmHK9 was predominantly expressed in roots, and the roots of ZmHK9-OX transgenic lines are markedly hypersensitive to ABA and ethylene, as compare to wild type. Consistent with its expression induced by PEG and exogenous ABA treatment, promoter sequence of this gene possessed drought and ABA responsive element. Moreover, the transgenic plants were much less affected by drought stress and recovered quickly after rewatering, stomatal complex size and stomatal density in the transgenic plants are significantly smaller and lower than those of the wild-type plants. In addition, ABA induced stomatal closure and the stomatal aperture of ZmHK9-OX lines was smaller than that of wild type. Collectively, it can be concluded that ZmHK9 regulates root elongation, stomatal development and drought tolerance through ABA dependent signaling pathway in Arabidopsis.

Validamycin A is a member of microbial-derived C(7)N-aminocyclitol family of natural products that is widely used as crop protectant and the precursor of the antidiabetic drug voglibose. Its biosynthetic gene clusters have been identified in several Streptomyces hygroscopicus strains, and a number of genes within the clusters have been functionally analyzed. Of these genes, valB, which encodes a sugar nucleotidyltransferase, was found through inactivation study to be essential for validamycin biosynthesis, but its role was unclear. To characterize the role of ValB in validamycin biosynthesis, four carbasugar phosphate analogues were synthesized and tested as substrate for ValB. The results showed that ValB efficiently catalyzes the conversion of valienol 1-phosphate to its nucleotidyl diphosphate derivatives, whereas other unsaturated carbasugar phosphates were found to be not the preferred substrate. ValB requires Mg(2+), Mn(2+), or Co(2+) for its optimal activity and uses the purine-based nucleotidyltriphosphates (ATP and GTP) more efficiently than the pyrimidine-based NTPs (CTP, dTTP, and UTP) as nucleotidyl donor. ValB represents the first member of unsaturated carbasugar nucleotidyltransferases involved in natural products biosynthesis. Its characterization not only expands our understanding of aminocyclitol-derived natural products biosynthesis, but may also facilitate the development of new tools for chemoenzymatic synthesis of carbohydrate mimetics.

Summary Gene valD , encodes a large vicinal oxygen chelate (VOC) superfamily protein, has been identified in the validamycin biosynthetic gene cluster. Inactivation of valD significantly reduced validamycin A production, which was fully restored with the full-length valD and partially restored with either N-terminal or C-terminal half by complementation. Heterologously expressed ValD catalyzed the epimerization of 2- epi -5- epi -valiolone to 5- epi -valiolone. This metalloenzyme is a homodimer with a metal ion-binding ratio of 0.73 mol/mole protein toward Fe 2+ , Mn 2+ , Ni 2+ , and Zn 2+ . Individual and combined site-directed mutations of eight putative active site residues revealed that the N-terminal H44/E107 and the C-terminal H315/E366 are more critical for the activity than the internal H130, E183, H229, and E291. Our data have established ValD as one of the largest proteins of the VOC superfamily, catalyzing an alternative epimerization for C 7 N-aminocyclitol biosynthesis.

Polyploidy, or whole-genome duplication, is an evolutionary innovation common in plants. Several major crops, including wheat, cotton and canola, are allopolyploids, containing two or more divergent genomes and some plants exist as intra- and interspecific hybrids. Such plants can display vigorous growth. A study of hybrids of Arabidopsis thaliana and A. arenosa sheds light on the mechanistic basis of this increased vigour. Epigenetic modulation of circadian clock regulators is shown to mediate the expression of genes in photosynthetic and starch metabolic pathways in allopolyploids. As a result, they accumulate more chlorophyll, produce more starch, and grow larger than the parents. Thus the hybrid and allopolyploid plants appear to gain advantages from the control of physiological and metabolic pathways by reversibly adjusting circadian clock regulators to make better use of the daylight hours. Polploidy is a common feature of many plants, and in addition some plants exist as intra- and interspecific hybrinds. Such plants display growth vigour, and genes involved in metabolism and energy, photosynthesis and starch accumulation are upregulated compared to the parents. This study examines the mechanistic basis of increased growth, and reports that epigenetic modifications of circadian clock regulators mediates the expression of genes in photosynthetic and metabolic pathways. Segregating hybrids and stable allopolyploids display morphological vigour1,2,3, and Arabidopsis allotetraploids are larger than the parents Arabidopsis thaliana and Arabidopsis arenosa1,4—the mechanisms for this are unknown. Circadian clocks mediate metabolic pathways and increase fitness in animals and plants5,6,7,8. Here we report that epigenetic modifications of the circadian clock genes CIRCADIAN CLOCK ASSOCIATED 1 (CCA1) and LATE ELONGATED HYPOCOTYL (LHY)9,10,11 and their reciprocal regulators TIMING OF CAB EXPRESSION 1 (TOC1) and GIGANTEA (GI)10,12,13 mediate expression changes in downstream genes and pathways. During the day, epigenetic repression of CCA1 and LHY induced the expression of TOC1, GI and downstream genes containing evening elements14 in chlorophyll and starch metabolic pathways in allotetraploids and F1 hybrids, which produced more chlorophyll and starch than the parents in the same environment. Mutations in cca1 and cca1 lhy and the daily repression of cca1 by RNA interference (RNAi) in TOC1::cca1(RNAi) transgenic plants increased the expression of downstream genes and increased chlorophyll and starch content, whereas constitutively expressing CCA1 or ectopically expressing TOC1::CCA1 had the opposite effect. The causal effects of CCA1 on output traits suggest that hybrids and allopolyploids gain advantages from the control of circadian-mediated physiological and metabolic pathways, leading to growth vigour and increased biomass.

SummaryAnsamitocins are potent antitumor maytansinoids produced by Actinosynnema pretiosum. Their biosynthesis involves the initial assembly of a macrolactam polyketide, followed by a series of postpolyketide synthase (PKS) modifications. Three ansamitocin glycosides were isolated from A. pretiosum and fully characterized structurally as novel ansamitocin derivatives, carrying a β-D-glucosyl group attached to the macrolactam amide nitrogen in place of the N-methyl group. By gene inactivation and complementation, asm25 was identified as the N-glycosyltransferase gene responsible for the macrolactam amide N-glycosylation of ansamitocins. Soluble, enzymatically active Asm25 protein was obtained from asm25-expressing E. coli by solubilization from inclusion bodies. Its optimal reaction conditions, including temperature, pH, metal ion requirement, and Km/Kcat, were determined. Asm25 also showed broad substrate specificity toward other ansamycins and synthetic indolin-2-ones. To the best of our knowledge, this represents the first in vitro characterization of a purified antibiotic N-glycosyltransferase.

The alteration of gene expression in hybrids may be an important factor promoting phenotypic variation and plasticity. To provide insight into the underlying molecular basis of maize heterosis in terms of the kernel number per ear, we established DGE profiles for the immature ears of maize hybrid Zong3/87-1 and its parental lines at the floral organ differentiation stage. Among 4,337 identified differentially expressed genes, 4,021 (92%) exhibited nonadditive expression patterns in the hybrid. Notably, the maize homolog of Arabidopsis EBP1, designated ZmEBP1, displayed an overdominant expression pattern in the Zong3/87-1 hybrid. Moreover, the results of qRT-PCR revealed that the ZmEBP1 gene was upregulated in the immature ears of the reciprocal hybrids Zong3/87-1 and 87-1/Zong3 at different developmental stages. Additionally, ectopic expression of ZmEBP1 in Arabidopsis increased organ size, which was mainly attributed to an increase in cell numbers, rather than cell size. Considering all of these findings together, we speculate that upregulation of ZmEBP1 in maize hybrids may accelerate cell proliferation and promote ear development.

SummaryA 45 kb DNA sequencing analysis from Streptomyces hygroscopicus 5008 involved in validamycin A (VAL-A) biosynthesis revealed 16 structural genes, 2 regulatory genes, 5 genes related transport, transposition/integration or tellurium resistance; another 4 genes had no obvious identity. The VAL-A biosynthetic pathway was proposed, with assignment of the required genetic functions confined to the sequenced region. A cluster of eight reassembled genes was found to support VAL-A synthesis in a heterologous host, S. lividans 1326. In vivo inactivation of the putative glycosyltransferase gene (valG) abolished the final attachment of glucose for VAL production and resulted in accumulation of the VAL-A precursor, validoxylamine, while the normal production of VAL-A could be restored by complementation with valG. The role of valG in the glycosylation of validoxylamine to VAL-A was demonstrated in vitro by enzymatic assay.

Based on the research of the standard twist drill geometry parameters, the cutting part of the structure of the geometric parameters are expressed as a fuction of radius, the chisel edge and the main cutting edge geometric parameters of the mathematical model are established. Drilling force is one of the most important parameters in the process of drilling, it directly affects the size of the drilling quality, but the geometry of a drill make the drilling process complicated. Under certain spindle speed and feed rate, apply MATLAB software simulation to drilling force model, achieve the prediction of the dynamic of drilling force and torque of the forecast. Keywords-Twist drill; geometrical parameter; Mechanistic model; MATLAB; Prediction.

In spite of commercial use of heterosis in agriculture, the molecular basis of heterosis is poorly understood. It was observed that maize hybrid Zong3/87-1 exhibited an earlier onset or heterosis in radicle emergence. To get insights into the underlying mechanism of heterosis in radicle emergence, differential proteomic analysis between hybrid and its parental lines was performed. In total, the number of differentially expressed protein spots between hybrid and its parental lines in dry and 24 h imbibed seed embryos were 134 and 191, respectively, among which 47.01% (63/134) and 34.55% (66/191) protein spots displayed nonadditively expressed pattern. Remarkably, 54.55% of nonadditively accumulated proteins in 24 h imbibed seed embryos displayed above or equal to the level of the higher parent patterns. Moreover, 155 differentially expressed protein spots were identified, which were grouped into eight functional classes, including transcription & translation, energy & metabolism, signal transduction, disease & defense, storage protein, transposable element, cell growth & division and unclassified proteins. In addition, one of the upregulated proteins in F1 hybrids was ZmACT2, a homolog of Arabidopsis thaliana ACT7 (AtACT7). Expressing ZmACT2 driven by the AtACT7 promoter partially complemented the low germination phenotype in the Atact7 mutant. These results indicated that hybridization between two parental lines can cause changes in the expression of a variety of proteins, and it is concluded that the altered pattern of gene expression at translational level in the hybrid may be responsible for the observed heterosis.

Background Auxin signaling is vital for plant growth and development, and plays important role in apical dominance, tropic response, lateral root formation, vascular differentiation, embryo patterning and shoot elongation. Auxin Response Factors (ARFs) are the transcription factors that regulate the expression of auxin responsive genes. The ARF genes are represented by a large multigene family in plants. The first draft of full maize genome assembly has recently been released, however, to our knowledge, the ARF gene family from maize (ZmARF genes) has not been characterized in detail. Results In this study, 31 maize (Zea mays L.) genes that encode ARF proteins were identified in maize genome. It was shown that maize ARF genes fall into related sister pairs and chromosomal mapping revealed that duplication of ZmARFs was associated with the chromosomal block duplications. As expected, duplication of some ZmARFs showed a conserved intron/exon structure, whereas some others were more divergent, suggesting the possibility of functional diversification for these genes. Out of these 31 ZmARF genes, 14 possess auxin-responsive element in their promoter region, among which 7 appear to show small or negligible response to exogenous auxin. The 18 ZmARF genes were predicted to be the potential targets of small RNAs. Transgenic analysis revealed that increased miR167 level could cause degradation of transcripts of six potential targets (ZmARF3, 9, 16, 18, 22 and 30). The expressions of maize ARF genes are responsive to exogenous auxin treatment. Dynamic expression patterns of ZmARF genes were observed in different stages of embryo development. Conclusions Maize ARF gene family is expanded (31 genes) as compared to Arabidopsis (23 genes) and rice (25 genes). The expression of these genes in maize is regulated by auxin and small RNAs. Dynamic expression patterns of ZmARF genes in embryo at different stages were detected which suggest that maize ARF genes may be involved in seed development and germination.

Gibberellin (GA) is an essential phytohormone that controls many aspects of plant development. To enhance our understanding of GA metabolism in maize, we intensively screened and identified 27 candidate genes encoding the seven GA metabolic enzymes including ent-copalyl diphosphate synthase (CPS), ent-kaurene synthase (KS), ent-kaurene oxidase (KO), ent-kaurenoic acid oxidase (KAO), GA 20-oxidase (GA20ox), GA 3-oxidase (GA3ox), and GA 2-oxidase (GA2ox), using all available public maize databases. The results indicate that maize genome contains three CPS, four KS, two KO and one KAO genes, and most of them are arranged separately on the maize genome, which differs from that in rice. In addition, the enzymes catalyzing the later steps (ZmGA20ox, ZmGA3ox and ZmGA2ox) are also encoded by gene families in maize, but GA3ox enzyme is likely to be encoded by single gene. Expression profiling analysis exhibited that transcripts of 15 GA metabolic genes could be detected during maize seed germination, which provides further evidence for the notion that increased synthesis of active GA in the embryo is required for triggering germination events. Moreover, a variety of temporal genes expression patterns of GA metabolic genes were detected, which revealed the complexity of underlying mechanism for GA regulated seed germination.

Cenp-F (also named mitosin) is a 350-kDa human kinetochore protein important for the mitotic progression. It is also a nuclear matrix protein in interphase cells. Here, we showed that overexpression of N-terminal deletion mutants of Cenp-F containing the C-terminal 112 residues induced chromatin condensation into numerous aggregates of varying sizes in interphase nucleus, colocalizing with the exogenous proteins. In situ hybridization using whole chromosome painting probes indicated that the chromatin aggregates were not prematurely condensed individual chromosomes. Neither were they due to apoptosis. We provided evidence showing association of Cenp-F with certain regions of interphase chromatin fibers. Cenp-F associated with the DNA-dependent protein kinase (DNA-PK), a trimeric protein complex critical for genome homeostasis. Moreover, the DNA-PK association activity of Cenp-F mutants correlated with their ability to induce chromatin aggregation. These results imply a role of Cenp-F in organization of interphase chromatin through association and possibly regulation of DNA-PK.

In some antibiotic producers,p-aminobenzoic acid (PABA) or its immediate precursor, 4-amino-4-deoxychorismate (ADC), is involved in primary metabolism and antibiotic biosynthesis. InStreptomycessp. FR-008, a genepabC-1putatively encoding a fold-type IV pyridoxal 5′-phosphate (PLP)-dependent enzyme was found within the antibiotic FR-008/candicidin biosynthetic gene cluster, whose inactivation significantly reduced the productivity of antibiotic FR-008 to about 20 % of the wild-type level. Its specific role in PABA formation was further demonstrated by the successful complementation of anEscherichia coli pabCmutant. Moreover, a free-standing genepabC-2, probably encoding another fold-type IV PLP-dependent enzyme, was cloned from the same strain. Inactivation ofpabC-2reduced antibiotic FR-008 yield to about 57 % of the wild-type level in the mutant, and the complementation of theE. coli pabCmutant established its involvement in PABA biosynthesis. Furthermore, apabC-1/pabC-2double mutant only retained about 4 % of the wild-type antibiotic FR-008 productivity, clearly indicating thatpabC-2also contributed to biosynthesis of this antibiotic. Surprisingly, apparently retarded growth of the double mutant was observed on minimal medium, which suggested that bothpabC-1andpabC-2are involved in PABA biosynthesis for primary metabolism. Finally, both PabC-1 and PabC-2 were shown to be functional ADC lyases byin vitroenzymic lysis with the release of pyruvate.pabC-1andpabC-2appear to represent the first two functional ADC lyase genes identified in actinomycetes. The involvement of these two ADC lyase genes in both cell growth and antibiotic FR-008 biosynthesis sets an example for the interplay between primary and secondary metabolisms in bacteria.

The gene valC, which encodes an enzyme homologous to the 2-epi-5-epi-valiolone kinase (AcbM) of the acarbose biosynthetic pathway, was identified in the validamycin A biosynthetic gene cluster. Inactivation of valC resulted in mutants that lack the ability to produce validamycin A. Complementation experiments with a replicating plasmid harboring full-length valC restored the production of validamycin A, thus suggesting a critical function of valC in validamycin biosynthesis. In vitro characterization of ValC revealed a new type of C7-cyclitol kinase, which phosphorylates valienone and validone--but not 2-epi-5-epi-valiolone, 5-epi-valiolone, or glucose--to afford their 7-phosphate derivatives. The results provide new insights into the activity of this enzyme and also confirm the existence of two different pathways leading to the same end-product: the valienamine moiety common to acarbose and validamycin A.