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Sample records for adh gene cluster

  1. Extended genetic effects of ADH cluster genes on the risk of alcohol dependence: from GWAS to replication.

    PubMed

    Park, Byung Lae; Kim, Jee Wook; Cheong, Hyun Sub; Kim, Lyoung Hyo; Lee, Boung Chul; Seo, Cheong Hoon; Kang, Tae-Cheon; Nam, Young-Woo; Kim, Goon-Bo; Shin, Hyoung Doo; Choi, Ihn-Geun

    2013-06-01

    Alcohol dependence (AD) is a multifactorial and polygenic disorder involving complex gene-to-gene and gene-to-environment interactions. Several genome-wide association studies have reported numerous risk factors for AD, but replication results following these studies have been controversial. To identify new candidate genes, the present study used GWAS and replication studies in a Korean cohort with AD. Genome-wide association analysis revealed that two chromosome regions on Chr. 4q22-q23 (ADH gene cluster, including ADH5, ADH4, ADH6, ADH1A, ADH1B, and ADH7) and Chr. 12q24 (ALDH2) showed multiple association signals for the risk of AD. To investigate detailed genetic effects of these ADH genes on AD, a follow-up study of the ADH gene cluster on 4q22-q23 was performed. A total of 90 SNPs, including ADH1B rs1229984 (H47R), were genotyped in an additional 975 Korean subjects. In case-control analysis, ADH1B rs1229984 (H47R) showed the most significant association with the risk of AD (p = 2.63 × 10(-21), OR = 2.35). Moreover, subsequent conditional analyses revealed that all positive associations of other ADH genes in the cluster disappeared, which suggested that ADH1B rs1229984 (H47R) might be the sole functional genetic marker across the ADH gene cluster. Our findings could provide additional information on the ADH gene cluster regarding the risk of AD, as well as a new and important insight into the genetic factors associated with AD.

  2. Diplotype Trend Regression Analysis of the ADH Gene Cluster and the ALDH2 Gene: Multiple Significant Associations with Alcohol Dependence

    PubMed Central

    Luo, Xingguang; Kranzler, Henry R.; Zuo, Lingjun; Wang, Shuang; Schork, Nicholas J.; Gelernter, Joel

    2006-01-01

    The set of alcohol-metabolizing enzymes has considerable genetic and functional complexity. The relationships between some alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) genes and alcohol dependence (AD) have long been studied in many populations, but not comprehensively. In the present study, we genotyped 16 markers within the ADH gene cluster (including the ADH1A, ADH1B, ADH1C, ADH5, ADH6, and ADH7 genes), 4 markers within the ALDH2 gene, and 38 unlinked ancestry-informative markers in a case-control sample of 801 individuals. Associations between markers and disease were analyzed by a Hardy-Weinberg equilibrium (HWE) test, a conventional case-control comparison, a structured association analysis, and a novel diplotype trend regression (DTR) analysis. Finally, the disease alleles were fine mapped by a Hardy-Weinberg disequilibrium (HWD) measure (J). All markers were found to be in HWE in controls, but some markers showed HWD in cases. Genotypes of many markers were associated with AD. DTR analysis showed that ADH5 genotypes and diplotypes of ADH1A, ADH1B, ADH7, and ALDH2 were associated with AD in European Americans and/or African Americans. The risk-influencing alleles were fine mapped from among the markers studied and were found to coincide with some well-known functional variants. We demonstrated that DTR was more powerful than many other conventional association methods. We also found that several ADH genes and the ALDH2 gene were susceptibility loci for AD, and the associations were best explained by several independent risk genes. PMID:16685648

  3. Genome-Wide Significant Association between Alcohol Dependence and a Variant in the ADH Gene Cluster

    PubMed Central

    Frank, Josef; Cichon, Sven; Treutlein, Jens; Ridinger, Monika; Mattheisen, Manuel; Hoffmann, Per; Herms, Stefan; Wodarz, Norbert; Soyka, Michael; Zill, Peter; Maier, Wolfgang; Mössner, Rainald; Gaebel, Wolfgang; Dahmen, Norbert; Scherbaum, Norbert; Schmäl, Christine; Steffens, Michael; Lucae, Susanne; Ising, Marcus; Müller-Myhsok, Bertram; Nöthen, Markus M; Mann, Karl; Kiefer, Falk; Rietschel, Marcella

    2011-01-01

    Alcohol dependence (AD) is an important contributory factor to the global burden of disease. The etiology of AD involves both environmental and genetic factors, and the disorder has a heritability of around 50%. The aim of the present study was to identify susceptibility genes for AD by performing a genome-wide association study (GWAS). The sample comprised 1,333 male in-patients with severe DSM-IV AD and 2,168 controls. These included 487 patients and 1,358 controls from a previous GWAS study by our group. All individuals were of German descent. Single marker tests and a polygenic score based analysis to assess the combined contribution of multiple markers with small effects were performed. The SNP rs1789891, which is located between the ADH1B and ADH1C genes, achieved genome-wide significance (p=1.27E–8; OR=1.46). Other markers from this region were also associated with AD, and conditional analyses indicated that these made a partially independent contribution. The SNP rs1789891 is in complete linkage disequilibrium with the functional Arg272Gln variant (p=1.24E–7, OR=1.31) of the ADH1C gene, which has been reported to modify the rate of ethanol oxidation to acetaldehyde in vitro. A polygenic score based approach produced a significant result (p=9.66E–9). This is the first GWAS of AD to provide genome-wide significant support for the role of the ADH gene cluster and to suggest a polygenic component to the etiology of AD. The latter result suggests that many more AD susceptibility genes still await identification. PMID:22004471

  4. Genome-wide significant association between alcohol dependence and a variant in the ADH gene cluster.

    PubMed

    Frank, Josef; Cichon, Sven; Treutlein, Jens; Ridinger, Monika; Mattheisen, Manuel; Hoffmann, Per; Herms, Stefan; Wodarz, Norbert; Soyka, Michael; Zill, Peter; Maier, Wolfgang; Mössner, Rainald; Gaebel, Wolfgang; Dahmen, Norbert; Scherbaum, Norbert; Schmäl, Christine; Steffens, Michael; Lucae, Susanne; Ising, Marcus; Müller-Myhsok, Bertram; Nöthen, Markus M; Mann, Karl; Kiefer, Falk; Rietschel, Marcella

    2012-01-01

    Alcohol dependence (AD) is an important contributory factor to the global burden of disease. The etiology of AD involves both environmental and genetic factors, and the disorder has a heritability of around 50%. The aim of the present study was to identify susceptibility genes for AD by performing a genome-wide association study (GWAS). The sample comprised 1333 male in-patients with severe AD according to the Diagnostic and Statistical Manual of Mental Disorders, 4th edition, and 2168 controls. These included 487 patients and 1358 controls from a previous GWAS study by our group. All individuals were of German descent. Single-marker tests and a polygenic score-based analysis to assess the combined contribution of multiple markers with small effects were performed. The single nucleotide polymorphism (SNP) rs1789891, which is located between the ADH1B and ADH1C genes, achieved genome-wide significance [P = 1.27E-8, odds ratio (OR) = 1.46]. Other markers from this region were also associated with AD, and conditional analyses indicated that these made a partially independent contribution. The SNP rs1789891 is in complete linkage disequilibrium with the functional Arg272Gln variant (P = 1.24E-7, OR = 1.31) of the ADH1C gene, which has been reported to modify the rate of ethanol oxidation to acetaldehyde in vitro. A polygenic score-based approach produced a significant result (P = 9.66E-9). This is the first GWAS of AD to provide genome-wide significant support for the role of the ADH gene cluster and to suggest a polygenic component to the etiology of AD. The latter result may indicate that many more AD susceptibility genes still await identification.

  5. Opossum alcohol dehydrogenases: Sequences, structures, phylogeny and evolution: evidence for the tandem location of ADH genes on opossum chromosome 5.

    PubMed

    Holmes, Roger S

    2009-03-16

    BLAT (BLAST-Like Alignment Tool) analyses and interrogations of the recently published opossum genome were undertaken using previously reported rat ADH amino acid sequences. Evidence is presented for six opossum ADH genes localized on chromosome 5 and organized in a comparable ADH gene cluster to that reported for human and rat ADH genes. The predicted amino acid sequences and secondary structures for the opossum ADH subunits and the intron-exon boundaries for opossum ADH genes showed a high degree of similarity with other mammalian ADHs, and four opossum ADH classes were identified, namely ADH1, ADH3, ADH6 and ADH4 (for which three genes were observed: ADH4A, ADH4B and ADH4C). Previous biochemical analyses of opossum ADHs have reported the tissue distribution and properties for these enzymes: ADH1, the major liver enzyme; ADH3, widely distributed in opossum tissues with similar kinetic properties to mammalian class 3 ADHs; and ADH4, for which several forms were localized in extrahepatic tissues, especially in the digestive system and in the eye. These ADHs are likely to perform similar functions to those reported for other mammalian ADHs in the metabolism of ingested and endogenous alcohols and aldehydes. Phylogenetic analyses examined opossum, human, rat, chicken and cod ADHs, and supported the proposed designation of opossum ADHs as class I (ADH1), class III (ADH3), class IV (ADH4A, ADH4B and ADH4C) and class VI (ADH6). Percentage substitution rates were examined for ADHs during vertebrate evolution which indicated that ADH3 is evolving at a much slower rate to that of the other ADH classes.

  6. The plant ADH gene family.

    PubMed

    Strommer, Judith

    2011-04-01

    The structures, evolution and functions of alcohol dehydrogenase gene families and their products have been scrutinized for half a century. Our understanding of the enzyme structure and catalytic activity of plant alcohol dehydrogenase (ADH-P) is based on the vast amount of information available for its animal counterpart. The probable origins of the enzyme from a simple β-coil and eventual emergence from a glutathione-dependent formaldehyde dehydrogenase have been well described. There is compelling evidence that the small ADH gene families found in plants today are the survivors of multiple rounds of gene expansion and contraction. To the probable original function of their products in the terminal reaction of anaerobic fermentation have been added roles in yeast-like aerobic fermentation and the production of characteristic scents that act to attract animals that serve as pollinators or agents of seed dispersal and to protect against herbivores.

  7. Ascidian and amphioxus Adh genes correlate functional and molecular features of the ADH family expansion during vertebrate evolution.

    PubMed

    Cañestro, Cristian; Albalat, Ricard; Hjelmqvist, Lars; Godoy, Laura; Jörnvall, Hans; Gonzàlez-Duarte, Roser

    2002-01-01

    The alcohol dehydrogenase (ADH) family has evolved into at least eight ADH classes during vertebrate evolution. We have characterized three prevertebrate forms of the parent enzyme of this family, including one from an urochordate (Ciona intestinalis) and two from cephalochordates (Branchiostoma floridae and Branchiostoma lanceolatum). An evolutionary analysis of the family was performed gathering data from protein and gene structures, exon-intron distribution, and functional features through chordate lines. Our data strongly support that the ADH family expansion occurred 500 million years ago, after the cephalochordate/vertebrate split, probably in the gnathostome subphylum line of the vertebrates. Evolutionary rates differ between the ancestral, ADH3 (glutathione-dependent formaldehyde dehydrogenase), and the emerging forms, including the classical alcohol dehydrogenase, ADH1, which has an evolutionary rate 3.6-fold that of the ADH3 form. Phylogenetic analysis and chromosomal mapping of the vertebrate Adh gene cluster suggest that family expansion took place by tandem duplications, probably concurrent with the extensive isoform burst observed before the fish/tetrapode split, rather than through the large-scale genome duplications also postulated in early vertebrate evolution. The absence of multifunctionality in lower chordate ADHs and the structures compared argue in favor of the acquisition of new functions in vertebrate ADH classes. Finally, comparison between B. floridae and B. lanceolatum Adhs provides the first estimate for a cephalochordate speciation, 190 million years ago, probably concomitant with the beginning of the drifting of major land masses from the Pangea.

  8. Association between common alcohol dehydrogenase gene (ADH) variants and schizophrenia and autism

    PubMed Central

    Wang, Kesheng; Zhang, Xiang-Yang; Pan, Xinghua; Wang, Guilin; Tan, Yunlong; Zhong, Chunlong; Krystal, John H.; State, Matthew; Zhang, Heping

    2013-01-01

    Humans express at least seven alcohol dehydrogenase (ADH) isoforms that are encoded by ADH gene cluster (ADH7–ADH1C–ADH1B–ADH1A–ADH6–ADH4–ADH5) at chromosome 4. ADHs are key catabolic enzymes for retinol and ethanol. The functional ADH variants (mostly rare) have been implicated in alcoholism risk. In addition to catalyzing the oxidation of retinol and ethanol, ADHs may be involved in the metabolic pathways of several neurotransmitters that are implicated in the neurobiology of neuropsychiatric disorders. In the present study, we comprehensively examined the associations between common ADH variants [minor allele frequency (MAF) >0.05] and 11 neuropsychiatric and neurological disorders. A total of 50,063 subjects in 25 independent cohorts were analyzed. The entire ADH gene cluster was imputed across these 25 cohorts using the same reference panels. Association analyses were conducted, adjusting for multiple comparisons. We found 28 and 15 single nucleotide polymorphisms (SNPs), respectively, that were significantly associated with schizophrenia in African-Americans and autism in European-Americans after correction by false discovery rate (FDR) (q <0.05); and 19 and 6 SNPs, respectively, that were significantly associated with these two disorders after region-wide correction by SNPSpD (8.9 × 10−5 ≤ p ≤ 0.0003 and 2.4 × 10−5 ≤ p ≤ 0.0003, respectively). No variants were significantly associated with the other nine neuropsychiatric disorders, including alcohol dependence. We concluded that common ADH variants conferred risk for both schizophrenia in African-Americans and autism in European-Americans. PMID:23468174

  9. Association between common alcohol dehydrogenase gene (ADH) variants and schizophrenia and autism.

    PubMed

    Zuo, Lingjun; Wang, Kesheng; Zhang, Xiang-Yang; Pan, Xinghua; Wang, Guilin; Tan, Yunlong; Zhong, Chunlong; Krystal, John H; State, Matthew; Zhang, Heping; Luo, Xingguang

    2013-07-01

    Humans express at least seven alcohol dehydrogenase (ADH) isoforms that are encoded by ADH gene cluster (ADH7-ADH1C-ADH1B-ADH1A-ADH6-ADH4-ADH5) at chromosome 4. ADHs are key catabolic enzymes for retinol and ethanol. The functional ADH variants (mostly rare) have been implicated in alcoholism risk. In addition to catalyzing the oxidation of retinol and ethanol, ADHs may be involved in the metabolic pathways of several neurotransmitters that are implicated in the neurobiology of neuropsychiatric disorders. In the present study, we comprehensively examined the associations between common ADH variants [minor allele frequency (MAF) >0.05] and 11 neuropsychiatric and neurological disorders. A total of 50,063 subjects in 25 independent cohorts were analyzed. The entire ADH gene cluster was imputed across these 25 cohorts using the same reference panels. Association analyses were conducted, adjusting for multiple comparisons. We found 28 and 15 single nucleotide polymorphisms (SNPs), respectively, that were significantly associated with schizophrenia in African-Americans and autism in European-Americans after correction by false discovery rate (FDR) (q < 0.05); and 19 and 6 SNPs, respectively, that were significantly associated with these two disorders after region-wide correction by SNPSpD (8.9 × 10(-5) ≤ p ≤ 0.0003 and 2.4 × 10(-5) ≤ p ≤ 0.0003, respectively). No variants were significantly associated with the other nine neuropsychiatric disorders, including alcohol dependence. We concluded that common ADH variants conferred risk for both schizophrenia in African-Americans and autism in European-Americans.

  10. Associations between ADH gene variants and alcohol phenotypes in Dutch adults.

    PubMed

    van Beek, Jenny H D A; Willemsen, Gonneke; de Moor, Marleen H M; Hottenga, Jouke Jan; Boomsma, Dorret I

    2010-02-01

    Recently, Macgregor et al. (2009) demonstrated significant associations of ADH polymorphisms with reactions to alcohol and alcohol consumption measures in an Australian sample. The aim of the present study was to replicate these findings in a Dutch sample. Survey data on alcohol phenotypes came from 1,754 unrelated individuals registered with the Netherlands Twin Register. SNPs in the ADH gene cluster located on chromosome 4q (n = 491) were subdivided in seven gene sets: ADH5, ADH4, ADH6, ADH1A, ADH1B, ADH1C and ADH7. Within these sets associations of SNPs with alcohol consumption measures, age at onset variables, reactions to alcohol and problem drinking liability were examined. Of the original 38 SNPs studied by Macgregor et al. (2009), six SNPs were not available in our dataset, because one of them had a minor allele frequency < .01 (rs1229984) and five could not be imputed. The remaining SNP associations with alcohol phenotypes as identified by Macgregor et al. (2009) were not replicated in the Dutch sample, after correcting for multiple genotype and phenotype testing. Significant associations were found however, for reactions to alcohol with a SNP in ADH5 (rs6827292, p = .001) and a SNP just upstream of ADH5 (rs6819724, p = .0007) that is in strong LD with rs6827292. Furthermore, an association between age at onset of regular alcohol use and a SNP just upstream of ADH7 (rs2654849, p = .003) was observed. No significant associations were found for alcohol consumption and problem drinking liability. Although these findings do not replicate the earlier findings at the SNP level, the results confirm the role of the ADH gene cluster in alcohol phenotypes.

  11. The ADH gene cluster SNP rs1789891 and temperamental dimensions in patients with alcohol dependence and affective disorders.

    PubMed

    Oniszczenko, Włodzimierz; Rybakowski, Janusz K; Dragan, Wojciech Ł; Grzywacz, Anna; Samochowiec, Jerzy

    2015-08-01

    This study had three objectives: (1) to assess the relationship between the single nucleotide polymorphism (SNP) rs1789891 in the alcohol dehydrogenase gene cluster and alcohol dependence and affective disorders; (2) to assess the differences in the Regulative Theory of Temperament (RTT) traits between an alcohol dependent group, an affective disorders group, and a healthy group; and (3) to assess the relationship between rs1789891 and temperament traits in a healthy group, taking into account the interaction of genotype and sex. The SNP rs1789891 was genotyped in a group of 194 alcohol dependent men, aged 21 to 71 years; 137 patients with affective disorders, including 51 males and 86 females, aged 19 to 85 years; and a group of 207 healthy individuals, including 89 males and 118 females, aged 18 to 71 years. Temperament traits (briskness, perseveration, sensory sensitivity, emotional reactivity, endurance, and activity) were assessed in all groups using the Formal Characteristics of Behaviour-Temperament Inventory. The comparative analysis of genotypic frequencies showed no significant differences between patients with alcoholism or affective disorders and those in the control group. Alcohol dependent men and the affective disorder group were characterised by higher levels of emotional reactivity (p-value 1.4e-5 and 9.84e-7, respectively) and lower levels of briskness, sensory sensitivity, endurance, and activity (p-value from 3.76e-8 to 0.012) when compared to the healthy group. The rs1789891 polymorphism was associated with briskness (p = 0.02), sensory sensitivity (p = 0.036), and activity (p = 0.049). None of the results were statistically significant after Bonferroni correction.

  12. Multiple ADH genes modulate risk for drug dependence in both African- and European-Americans

    PubMed Central

    Luo, Xingguang; Kranzler, Henry R.; Zuo, Lingjun; Wang, Shuang; Schork, Nicholas J.; Gelernter, Joel

    2007-01-01

    Drug dependence (DD) is commonly co-morbid with alcohol dependence (AD). Many studies have also shown common genetic risk factors for these disorders. We previously reported associations of AD with seven alcohol dehydrogenase (ADH ) genes. The present study examines the relationship between these genes and DD. We genotyped 16 markers within the ADH gene cluster and 38 unlinked ancestry-informative markers in a case–control sample of 718 individuals. All markers were consistent with Hardy–Weinberg equilibrium in controls, but some markers showed Hardy–Weinberg disequilibrium in cases (minimal P = 0.002). Genotypes of many markers were associated with DD, both before and after controlling for admixture effects (minimal P < 1.0 × 10−6). Diplotype trend regression analysis showed that ADH5 and ADH6 genotypes, and diplotypes at ADH1A, ADH1B, ADH1C and ADH7 (minimal P = 0.002), were associated with DD in European-Americans and/or African-Americans. This first report of an allelic association of these loci with DD provides new insight into the mechanism of genetic risk for DD. These findings, obtained using a series of powerful and reliable analytic methods, may also help to explain the high rate of co-morbidity between AD and DD. PMID:17185388

  13. A Global Perspective on Genetic Variation at the ADH Genes Reveals Unusual Patterns of Linkage Disequilibrium and Diversity

    PubMed Central

    Osier, Michael V.; Pakstis, Andrew J.; Soodyall, Himla; Comas, David; Goldman, David; Odunsi, Adekunle; Okonofua, Friday; Parnas, Josef; Schulz, Leslie O.; Bertranpetit, Jaume; Bonne-Tamir, Batsheva; Lu, Ru-Band; Kidd, Judith R.; Kidd, Kenneth K.

    2002-01-01

    Variants of different Class I alcohol dehydrogenase (ADH) genes have been shown to be associated with an effect that is protective against alcoholism. Previous work from our laboratory has shown that the two sites showing the association are in linkage disequilibrium and has identified the ADH1B Arg47His site as causative, with the ADH1C Ile349Val site showing association only because of the disequilibrium. Here, we describe an initial study of the nature of linkage disequilibrium and genetic variation, in population samples from different regions of the world, in a larger segment of the ADH cluster (including the three Class I ADH genes and ADH7). Linkage disequilibrium across ∼40 kb of the Class I ADH cluster is moderate to strong in all population samples that we studied. We observed nominally significant pairwise linkage disequilibrium, in some populations, between the ADH7 site and some Class I ADH sites, at moderate values and at a molecular distance as great as 100 kb. Our data indicate (1) that most ADH-alcoholism association studies have failed to consider many sites in the ADH cluster that may harbor etiologically significant alleles and (2) that the relevance of the various ADH sites will be population dependent. Some individual sites in the Class I ADH cluster show Fst values that are among the highest seen among several dozen unlinked sites that were studied in the same subset of populations. The high Fst values can be attributed to the discrepant frequencies of specific alleles in eastern Asia relative to those in other regions of the world. These alleles are part of a single haplotype that exists at high (>65%) frequency only in the eastern-Asian samples. It seems unlikely that this haplotype, which is rare or unobserved in other populations, reached such high frequency because of random genetic drift alone. PMID:12050823

  14. Complementation of an Escherichia coli adhE mutant by the Entamoeba histolytica EhADH2 gene provides a method for the identification of new antiamebic drugs.

    PubMed Central

    Yong, T S; Li, E; Clark, D; Stanley, S L

    1996-01-01

    The pathogenic protozoan parasite Entamoeba histolytica, the cause of amebic dysentery and amebic liver abscess, is an obligate anaerobe, and derives energy from the fermentation of glucose to ethanol with pyruvate and acetyl coenzyme A as intermediates. We have isolated EhADH2, a key enzyme in this pathway, that is a NAD+- and Fe2+-dependent bifunctional enzyme with acetaldehyde dehydrogenase and alcohol dehydrogenase activities. EhADH2 is the only known eukaryotic member of a newly defined family of prokaryotic multifunctional enzymes, which includes the Escherichia coli AdhE enzyme, an enzyme required for anaerobic growth of E. coli. Because of the critical role of EhADH2 in the amebic fermentation pathway and the lack of known eukaryotic homologues of the EhADH2 enzyme, EhADH2 represents a potential target for antiamebic chemotherapy. However, screening of compounds for antiamebic activity is hampered by the cost of large scale growth of Ent. histolytica, and difficulties in quantitating drug efficacy in vitro. To approach this problem, we expressed the EhADH2 gene in a mutant strain of E. coli carrying a deletion of the adhE gene. Expression of EhADH2 restored the ability of the mutant E. coli strain to grow under anaerobic conditions. By screening compounds for the ability to inhibit the anaerobic growth of the E. coli/EhADH2 strain, we have developed a rapid assay for identifying compounds with anti-EhADH2 activity. Using bacteria to bypass the need for parasite culture in the initial screening process for anti-parasitic agents could greatly simplify and reduce the cost of identifying new therapeutic agents effective against parasitic diseases. Images Fig. 1 Fig. 2 Fig. 3 PMID:8692838

  15. Screening of Two ADH4 Variations in a Swedish Cluster Headache Case–Control Material

    PubMed Central

    Fourier, Carmen; Ran, Caroline; Steinberg, Anna; Sjöstrand, Christina; Waldenlind, Elisabet

    2016-01-01

    Background Cluster headache (CH) is a severe neurovascular disorder and an increasing amount of evidence points to a genetic contribution to this disease. When CH was first described, it was observed that alcohol may precipitate an attack during the active phase of the disease. The alcohol dehydrogenase 4 (ADH4) gene encodes an enzyme which contributes to the metabolization of alcohol and is, therefore, an interesting candidate gene for CH. Two Italian groups have reported association of the single nucleotide polymorphism (SNP) rs1126671 located in the ADH4 gene with an increased risk of CH in Italy. In addition, one of the groups found an association between the ADH4 SNP rs1800759 and CH. Objective To perform a replication study on the ADH4 SNPs rs1126671 and rs1800759 in a large homogeneous Swedish case–control cohort in order to further investigate the possible contribution of ADH4 to CH. Methods A total of 390 unrelated patients diagnosed with CH and 389 controls representing a general Swedish population were recruited to the study. DNA samples from patients and controls were genotyped for the two ADH4 SNPs rs1126671 and rs1800759 using quantitative real‐time polymerase chain reaction. Statistical analyses of genotype, allele and haplotype frequencies for the two SNPs were performed and compared between patients and controls. Results For rs1126671, the minor allele frequency (A allele) was 32.8% (n = 254) in controls compared with 31.9% (n = 249) in CH patients. The minor allele frequency (A allele) of rs1800759 was 42.3% (n = 324) in controls and 41.9% (n = 327) in CH patients. Statistical analysis showed no significant differences in allele as well as in genotype or haplotype frequencies between the patient and control group for either SNP. This was also seen after stratifying the patient group for experiencing alcohol as a trigger factor. Conclusions The data did not support an association of the ADH4 SNPs rs1126671 and rs1800759 with CH

  16. The Bifunctional Alcohol and Aldehyde Dehydrogenase Gene, adhE, Is Necessary for Ethanol Production in Clostridium thermocellum and Thermoanaerobacterium saccharolyticum

    PubMed Central

    Lo, Jonathan; Zheng, Tianyong; Hon, Shuen; Olson, Daniel G.

    2015-01-01

    ABSTRACT Thermoanaerobacterium saccharolyticum and Clostridium thermocellum are anaerobic thermophilic bacteria being investigated for their ability to produce biofuels from plant biomass. The bifunctional alcohol and aldehyde dehydrogenase gene, adhE, is present in these bacteria and has been known to be important for ethanol formation in other anaerobic alcohol producers. This study explores the inactivation of the adhE gene in C. thermocellum and T. saccharolyticum. Deletion of adhE reduced ethanol production by >95% in both T. saccharolyticum and C. thermocellum, confirming that adhE is necessary for ethanol formation in both organisms. In both adhE deletion strains, fermentation products shifted from ethanol to lactate production and resulted in lower cell density and longer time to reach maximal cell density. In T. saccharolyticum, the adhE deletion strain lost >85% of alcohol dehydrogenase (ADH) activity. Aldehyde dehydrogenase (ALDH) activity did not appear to be affected, although ALDH activity was low in cell extracts. Adding ubiquinone-0 to the ALDH assay increased activity in the T. saccharolyticum parent strain but did not increase activity in the adhE deletion strain, suggesting that ALDH activity was inhibited. In C. thermocellum, the adhE deletion strain lost >90% of ALDH and ADH activity in cell extracts. The C. thermocellum adhE deletion strain contained a point mutation in the lactate dehydrogenase gene, which appears to deregulate its activation by fructose 1,6-bisphosphate, leading to constitutive activation of lactate dehydrogenase. IMPORTANCE Thermoanaerobacterium saccharolyticum and Clostridium thermocellum are bacteria that have been investigated for their ability to produce biofuels from plant biomass. They have been engineered to produce higher yields of ethanol, yet questions remain about the enzymes responsible for ethanol formation in these bacteria. The genomes of these bacteria encode multiple predicted aldehyde and alcohol

  17. Nucleotide sequence of the genomic region encompassing Adh and Adh-dup genes of D. lebanonensis (Scaptodrosophila): gene expression and evolutionary relationships.

    PubMed

    Juan, E; Papaceit, M; Quintana, A

    1994-05-01

    The region of the genome of D. lebanonensis that contains the Adh gene and the downstream Adh-dup gene was sequenced. The structure of the two genes is the same as has been described for D. melanogaster. Adh has two promoters and Adh-dup has only one putative promoter. The levels of expression of the two genes in this species are dramatically different. Hybridizing the same Northern blots with a specific probe for Adh-dup, we did not find transcripts for this gene in D. lebanonensis. The level of Adh distal transcript in adults of D. lebanonensis is five times greater than that of D. melanogaster adults. The maximum levels of proximal transcript are attained at different larval stages in the two species, being three times higher in D. melanogaster late-second-instar larvae than in D. lebanonensis first-instar larvae. The level of Adh transcripts allowed us to determine distal and proximal initiation transcription sites, the position of the first intron, the use of two polyadenylation signals, and the heterogeneity of polyadenylation sites. Temporal and spatial expression profiles of the Adh gene of D. lebanonensis show qualitative differences compared with D. melanogaster. Adh and Adh-dup evolve differently as shown by the synonymous and nonsynonymous substitution rates for the coding region of both genes when compared across two species of the melanogaster group, two of the obscura group of the subgenus Sophophora and D. lebanonensis of the victoria group of the subgenus Scaptodrsophila. Synonymous rates for Adh are approximately half those for Adh-dup, while nonsynonymous rates for Adh are generally higher than those for Adh-dup. Adh shows 76.8% identities at the protein level and 70.2% identities at the nucleotide level while Adh-dup shows 83.7% identities at the protein level and 67.5% identities at the nucleotide level. Codon usage for Adh-dup is shown to be less biased than for Adh, which could explain the higher synonymous rates and the generally lower

  18. Transcriptional control of ADH genes in the xylose-fermenting yeast Pichia stipitis

    SciTech Connect

    Cho, J.Y.; Jeffries, T.W. |

    1999-06-01

    The authors studied the expression of the genes encoding group 1 alcohol dehydrogenases (PsADH1 and PsADH2) in the xylose-fermenting yeast Pichia stipitis CBS 6054. The cells expressed PsADH1 approximately 10 times higher under oxygen-limited conditions than under fully aerobic conditions when cultivated on xylose. Transcripts of PsADH2 were not detectable under either aeration condition. The authors used a PsADH1::lacZ fusion to monitor PsADH1 expression and found that expression increased as oxygen decreased. The level of PsADH1 transcript was expressed about 10-fold in cells grown in the presence of heme under oxygen-limited conditions. Concomitantly with the induction of PsADH1, PsCYC1 expression was regressed. These results indicate that oxygen availability regulates PsADH1 expression and that regulation may be mediated by heme. The regulation of PsADH2 expression was also examined in other genetic backgrounds. Disruption of PsADH1 dramatically increased PsADH2 expression on nonfermentable carbon sources under fully aerobic conditions, indicating that the expression of PsADH2 is subject to feedback regulation under these conditions.

  19. Transcriptional Control of ADH Genes in the Xylose-Fermenting Yeast Pichia stipitis

    PubMed Central

    Cho, Jae-Yong; Jeffries, Thomas W.

    1999-01-01

    We studied the expression of the genes encoding group I alcohol dehydrogenases (PsADH1 and PsADH2) in the xylose-fermenting yeast Pichia stipitis CBS 6054. The cells expressed PsADH1 approximately 10 times higher under oxygen-limited conditions than under fully aerobic conditions when cultivated on xylose. Transcripts of PsADH2 were not detectable under either aeration condition. We used a PsADH1::lacZ fusion to monitor PsADH1 expression and found that expression increased as oxygen decreased. The level of PsADH1 transcript was repressed about 10-fold in cells grown in the presence of heme under oxygen-limited conditions. Concomitantly with the induction of PsADH1, PsCYC1 expression was repressed. These results indicate that oxygen availability regulates PsADH1 expression and that regulation may be mediated by heme. The regulation of PsADH2 expression was also examined in other genetic backgrounds. Disruption of PsADH1 dramatically increased PsADH2 expression on nonfermentable carbon sources under fully aerobic conditions, indicating that the expression of PsADH2 is subject to feedback regulation under these conditions. PMID:10347014

  20. Temperature and water loss affect ADH activity and gene expression in grape berry during postharvest dehydration.

    PubMed

    Cirilli, Marco; Bellincontro, Andrea; De Santis, Diana; Botondi, Rinaldo; Colao, Maria Chiara; Muleo, Rosario; Mencarelli, Fabio

    2012-05-01

    Clusters of Aleatico wine grape were picked at 18°Brix and placed at 10, 20, or 30°C, 45% relative humidity (RH) and 1.5m/s of air flow to dehydrate the berries up to 40% of loss of initial fresh weight. Sampling was done at 0%, 10%, 20%, 30%, and 40% weight loss (wl). ADH (alcohol dehydrogenase) gene expression, enzyme activity, and related metabolites were analysed. At 10°C, acetaldehyde increased rapidly and then declined, while ethanol continued to rise. At 20°C, acetaldehyde and ethanol increased significantly with the same pattern and declined at 40%wl. At 30°C, acetaldehyde did not increase but ethanol increased rapidly already at 10%wl. At the latter temperature, a significant increase in acetic acid and ethyl acetate occurred, while at 10°C their values were low. At 30°C, the ADH activity (ethanol to acetaldehyde direction), increased rapidly but acetaldehyde did not rise because of its oxidation to acetic acid, which increased together with ethyl acetate. At 10°C, the ADH activity increased at 20%wl and continued to rise even at 40%wl, meaning that ethanol oxidation was delayed. At 20°C, the behaviour was intermediate to the other temperatures. The relative expression of the VvAdh2 gene was the highest at 10°C already at 10%wl in a synchrony with the ADH activity, indicating a rapid response likely due to low temperature. The expression subsequently declined. At 20 and 30°C, the expression was lower and increased slightly during dehydration in combination with the ADH activity. This imbalance between gene expression and ADH activity at 10°C, as well as the unexpected expression of the carotenoid cleavage dioxygenase 1 (CCD1) gene, opens the discussion on the stress sensitivity and transcription event during postharvest dehydration, and the importance of carefully monitoring temperature during dehydration.

  1. Evolution of the adhE gene product of Escherichia coli from a functional reductase to a dehydrogenase. Genetic and biochemical studies of the mutant proteins.

    PubMed

    Membrillo-Hernandez, J; Echave, P; Cabiscol, E; Tamarit, J; Ros, J; Lin, E C

    2000-10-27

    The multifunctional AdhE protein of Escherichia coli (encoded by the adhE gene) physiologically catalyzes the sequential reduction of acetyl-CoA to acetaldehyde and then to ethanol under fermentative conditions. The NH(2)-terminal region of the AdhE protein is highly homologous to aldehyde:NAD(+) oxidoreductases, whereas the COOH-terminal region is homologous to a family of Fe(2+)-dependent ethanol:NAD(+) oxidoreductases. This fusion protein also functions as a pyruvate formate lyase deactivase. E. coli cannot grow aerobically on ethanol as the sole carbon and energy source because of inadequate rate of adhE transcription and the vulnerability of the AdhE protein to metal-catalyzed oxidation. In this study, we characterized 16 independent two-step mutants with acquired and improved aerobic growth ability on ethanol. The AdhE proteins in these mutants catalyzed the sequential oxidation of ethanol to acetaldehyde and to acetyl-CoA. All first stage mutants grew on ethanol with a doubling time of about 240 min. Sequence analysis of a randomly chosen mutant revealed an Ala-267 --> Thr substitution in the acetaldehyde:NAD(+) oxidoreductase domain of AdhE. All second stage mutants grew on ethanol with a doubling time of about 90 min, and all of them produced an AdhE(A267T/E568K). Purified AdhE(A267T) and AdhE(A267T/E568K) showed highly elevated acetaldehyde dehydrogenase activities. It therefore appears that when AdhE catalyzes the two sequential reactions in the counter-physiological direction, acetaldehyde dehydrogenation is the rate-limiting step. Both mutant proteins were more thermosensitive than the wild-type protein, but AdhE(A267T/E568K) was more thermal stable than AdhE(A267T). Since both mutant enzymes exhibited similar kinetic properties, the second mutation probably conferred an increased growth rate on ethanol by stabilizing AdhE(A267T). PMID:10922373

  2. [Molecular phylogenetic relationships among species in the genus Sorghum based on partial Adh1 gene].

    PubMed

    Liao, Fang; Liu, Yong; Yang, Xiu-Li; Huang, Guo-Ming; Niu, Chun-Jing

    2009-05-01

    The genus Sorghum contains some important grain crops and economically important forage grasses as well as agricultural weeds. The goals of this study were to elucidate the phylogenetic relationships between Sorghum species and ascertain the taxonomy status of the quarantine weeds which provide bases for effective utilization of its germ resources on molecular breeding and improvement of crop qualities and thus provide important guidance for port detection. In the present study, total DNA from the seeds of 8 Sorghum species (four S. almum, two S. halepense, one S. silk and one S. sudanense)were extracted, and the partial Adh1 gene of about 2,000 bp in length were amplified by PCR using specific primers designed from conserved regions of Adh1 gene reported in the GenBank (AF050456) and sequenced. Based on these sequences and other 24 Adh1 sequences registered in the GenBank, the phylogenetic trees constructed by multiple methods (MP, ML, and NJ) with the corresponding Adh1 sequence of Cleistachne sorghoides as the outgroup shared almost the same topology. The results showed that: (1) there were obviously three lineages for the genus Sorghum. One included two subgenera Chaetosorghum and Heterosorghum, and another included subgenus Eusorghum, both of which consisted of 2n=20 and 2n=40 species with small chromosomes. Still another lineage contained the two subgenera Parasorghum and Stiposorghum with 2n=10 species and their polyploid relatives with relatively large chromosomes; (2) geographical divergence clearly existed in the S. almum; (3) S. purpureosericeum, S. versicolor, S. nitidum and S. leiocladum of subgenus Parasorghum clustered together, but S. matarankense, S. grande, and S. timorense of the same subgenus clustered with the species of subgenus Stiposorghum, showing closer relationship with subgenus Stiposorghum; (4) S. macrospermum was more closely related to S. laxiflorum than the other species of the genus Sorghum.

  3. Multiway real-time PCR gene expression profiling in yeast Saccharomyces cerevisiae reveals altered transcriptional response of ADH-genes to glucose stimuli

    PubMed Central

    Ståhlberg, Anders; Elbing, Karin; Andrade-Garda, José Manuel; Sjögreen, Björn; Forootan, Amin; Kubista, Mikael

    2008-01-01

    Background The large sensitivity, high reproducibility and essentially unlimited dynamic range of real-time PCR to measure gene expression in complex samples provides the opportunity for powerful multivariate and multiway studies of biological phenomena. In multiway studies samples are characterized by their expression profiles to monitor changes over time, effect of treatment, drug dosage etc. Here we perform a multiway study of the temporal response of four yeast Saccharomyces cerevisiae strains with different glucose uptake rates upon altered metabolic conditions. Results We measured the expression of 18 genes as function of time after addition of glucose to four strains of yeast grown in ethanol. The data are analyzed by matrix-augmented PCA, which is a generalization of PCA for 3-way data, and the results are confirmed by hierarchical clustering and clustering by Kohonen self-organizing map. Our approach identifies gene groups that respond similarly to the change of nutrient, and genes that behave differently in mutant strains. Of particular interest is our finding that ADH4 and ADH6 show a behavior typical of glucose-induced genes, while ADH3 and ADH5 are repressed after glucose addition. Conclusion Multiway real-time PCR gene expression profiling is a powerful technique which can be utilized to characterize functions of new genes by, for example, comparing their temporal response after perturbation in different genetic variants of the studied subject. The technique also identifies genes that show perturbed expression in specific strains. PMID:18412983

  4. Effects of glucose, ethanol and acetic acid on regulation of ADH2 gene from Lachancea fermentati.

    PubMed

    Yaacob, Norhayati; Mohamad Ali, Mohd Shukuri; Salleh, Abu Bakar; Abdul Rahman, Nor Aini

    2016-01-01

    Background. Not all yeast alcohol dehydrogenase 2 (ADH2) are repressed by glucose, as reported in Saccharomyces cerevisiae. Pichia stipitis ADH2 is regulated by oxygen instead of glucose, whereas Kluyveromyces marxianus ADH2 is regulated by neither glucose nor ethanol. For this reason, ADH2 regulation of yeasts may be species dependent, leading to a different type of expression and fermentation efficiency. Lachancea fermentati is a highly efficient ethanol producer, fast-growing cells and adapted to fermentation-related stresses such as ethanol and organic acid, but the metabolic information regarding the regulation of glucose and ethanol production is still lacking. Methods. Our investigation started with the stimulation of ADH2 activity from S. cerevisiae and L. fermentati by glucose and ethanol induction in a glucose-repressed medium. The study also embarked on the retrospective analysis of ADH2 genomic and protein level through direct sequencing and sites identification. Based on the sequence generated, we demonstrated ADH2 gene expression highlighting the conserved NAD(P)-binding domain in the context of glucose fermentation and ethanol production. Results. An increase of ADH2 activity was observed in starved L. fermentati (LfeADH2) and S. cerevisiae (SceADH2) in response to 2% (w/v) glucose induction. These suggest that in the presence of glucose, ADH2 activity was activated instead of being repressed. An induction of 0.5% (v/v) ethanol also increased LfeADH2 activity, promoting ethanol resistance, whereas accumulating acetic acid at a later stage of fermentation stimulated ADH2 activity and enhanced glucose consumption rates. The lack in upper stream activating sequence (UAS) and TATA elements hindered the possibility of Adr1 binding to LfeADH2. Transcription factors such as SP1 and RAP1 observed in LfeADH2 sequence have been implicated in the regulation of many genes including ADH2. In glucose fermentation, L. fermentati exhibited a bell-shaped ADH2

  5. Effects of glucose, ethanol and acetic acid on regulation of ADH2 gene from Lachancea fermentati

    PubMed Central

    Yaacob, Norhayati; Salleh, Abu Bakar; Abdul Rahman, Nor Aini

    2016-01-01

    Background. Not all yeast alcohol dehydrogenase 2 (ADH2) are repressed by glucose, as reported in Saccharomyces cerevisiae. Pichia stipitis ADH2 is regulated by oxygen instead of glucose, whereas Kluyveromyces marxianus ADH2 is regulated by neither glucose nor ethanol. For this reason, ADH2 regulation of yeasts may be species dependent, leading to a different type of expression and fermentation efficiency. Lachancea fermentati is a highly efficient ethanol producer, fast-growing cells and adapted to fermentation-related stresses such as ethanol and organic acid, but the metabolic information regarding the regulation of glucose and ethanol production is still lacking. Methods. Our investigation started with the stimulation of ADH2 activity from S. cerevisiae and L. fermentati by glucose and ethanol induction in a glucose-repressed medium. The study also embarked on the retrospective analysis of ADH2 genomic and protein level through direct sequencing and sites identification. Based on the sequence generated, we demonstrated ADH2 gene expression highlighting the conserved NAD(P)-binding domain in the context of glucose fermentation and ethanol production. Results. An increase of ADH2 activity was observed in starved L. fermentati (LfeADH2) and S. cerevisiae (SceADH2) in response to 2% (w/v) glucose induction. These suggest that in the presence of glucose, ADH2 activity was activated instead of being repressed. An induction of 0.5% (v/v) ethanol also increased LfeADH2 activity, promoting ethanol resistance, whereas accumulating acetic acid at a later stage of fermentation stimulated ADH2 activity and enhanced glucose consumption rates. The lack in upper stream activating sequence (UAS) and TATA elements hindered the possibility of Adr1 binding to LfeADH2. Transcription factors such as SP1 and RAP1 observed in LfeADH2 sequence have been implicated in the regulation of many genes including ADH2. In glucose fermentation, L. fermentati exhibited a bell-shaped ADH2

  6. A highly conserved sequence in the 3'-untranslated region of the drosophila Adh gene plays a functional role in Adh expression.

    PubMed Central

    Parsch, J; Stephan, W; Tanda, S

    1999-01-01

    Phylogenetic analysis identified a highly conserved eight-base sequence (AAGGCTGA) within the 3'-untranslated region (UTR) of the Drosophila alcohol dehydrogenase gene, Adh. To examine the functional significance of this conserved motif, we performed in vitro deletion mutagenesis on the D. melanogaster Adh gene followed by P-element-mediated germline transformation. Deletion of all or part of the eight-base sequence leads to a twofold increase in in vivo ADH enzymatic activity. The increase in activity is temporally and spatially general and is the result of an underlying increase in Adh transcript. These results indicate that the conserved 3'-UTR motif plays a functional role in the negative regulation of Adh gene expression. The evolutionary significance of our results may be understood in the context of the amino acid change that produces the ADH-F allele and also leads to a twofold increase in ADH activity. While there is compelling evidence that the amino acid replacement has been a target of positive selection, the conservation of the 3'-UTR sequence suggests that it is under strong purifying selection. The selective difference between these two sequence changes, which have similar effects on ADH activity, may be explained by different metabolic costs associated with the increase in activity. PMID:9927459

  7. Molecular phylogeny and evolution of alcohol dehydrogenase (Adh) genes in legumes

    PubMed Central

    Fukuda, Tatsuya; Yokoyama, Jun; Nakamura, Toru; Song, In-Ja; Ito, Takuro; Ochiai, Toshinori; Kanno, Akira; Kameya, Toshiaki; Maki, Masayuki

    2005-01-01

    Background Nuclear genes determine the vast range of phenotypes that are responsible for the adaptive abilities of organisms in nature. Nevertheless, the evolutionary processes that generate the structures and functions of nuclear genes are only now be coming understood. The aim of our study is to isolate the alcohol dehydrogenase (Adh) genes in two distantly related legumes, and use these sequences to examine the molecular evolutionary history of this nuclear gene. Results We isolated the expressed Adh genes from two species of legumes, Sophora flavescens Ait. and Wisteria floribunda DC., by a RT-PCR based approach and found a new Adh locus in addition to homologues of the Adh genes found previously in legumes. To examine the evolution of these genes, we compared the species and gene trees and found gene duplication of the Adh loci in the legumes occurred as an ancient event. Conclusion This is the first report revealing that some legume species have at least two Adh gene loci belonging to separate clades. Phylogenetic analyses suggest that these genes resulted from relatively ancient duplication events. PMID:15836788

  8. Tissue-Specific Expression Phenotypes of Hawaiian Drosophila Adh Genes in Drosophila Melanogaster Transformants

    PubMed Central

    Wu, C. Y.; Mote-Jr., J.; Brennan, M. D.

    1990-01-01

    Interspecific differences in the tissue-specific patterns of expression displayed by the alcohol dehydrogenase (Adh) genes within the Hawaiian picture-winged Drosophila represent a rich source of evolutionary variation in gene regulation. Study of the cis-acting elements responsible for regulatory differences between Adh genes from various species is greatly facilitated by analyzing the behavior of the different Adh genes in a homogeneous background. Accordingly, the Adh gene from Drosophila grimshawi was introduced into the germ line of Drosophila melanogaster by means of P element-mediated transformation, and transformants carrying this gene were compared to transformants carrying the Adh genes from Drosophila affinidisjuncta and Drosophila hawaiiensis. The results indicate that the D. affinidisjuncta and D. grimshawi genes have relatively higher levels of expression and broader tissue distribution of expression than the D. hawaiiensis gene in larvae. All three genes are expressed at similar overall levels in adults, with differences in tissue distribution of enzyme activity corresponding to the pattern in the donor species. However, certain systematic differences between Adh gene expression in transformants and in the Hawaiian Drosophila are noted along with tissue-specific position effects in some cases. The implications of these findings for the understanding of evolved regulatory variation are discussed. PMID:2165967

  9. A Drosophila Adh gene can be activated in trans by an enhancer.

    PubMed Central

    Rothberg, I; Hotaling, E; Sofer, W

    1991-01-01

    The ability of a segment of the Drosophila Adh gene to produce ADH activity in larvae is dependent upon the presence of a 53 bp sequence (called NS1) located between 289 and 341 bp upstream of the larval transcription start site. This sequence behaves like an enhancer in that it can stimulate gene activity when it is placed at various distances from, or on either side of, an Adh gene. Like a typical enhancer, NS1 does not ordinarily function in trans. However, when an Adh gene lacking NS1 is placed on one plasmid, and a second gene carrying NS1 is placed on another, and the two plasmids are interlocked in a catenane, both genes are active. This finding supports the mechanism of loop-mediated enhancer action. Images PMID:1945848

  10. In vivo introduction of unpreferred synonymous codons into the Drosophila Adh gene results in reduced levels of ADH protein.

    PubMed Central

    Carlini, David B; Stephan, Wolfgang

    2003-01-01

    The evolution of codon bias, the unequal usage of synonymous codons, is thought to be due to natural selection for the use of preferred codons that match the most abundant species of isoaccepting tRNA, resulting in increased translational efficiency and accuracy. We examined this hypothesis by introducing 1, 6, and 10 unpreferred codons into the Drosophila alcohol dehydrogenase gene (Adh). We observed a significant decrease in ADH protein production with number of unpreferred codons, confirming the importance of natural selection as a mechanism leading to codon bias. We then used this empirical relationship to estimate the selection coefficient (s) against unpreferred synonymous mutations and found the value (s >or= 10(-5)) to be approximately one order of magnitude greater than previous estimates from population genetics theory. The observed differences in protein production appear to be too large to be consistent with current estimates of the strength of selection on synonymous sites in D. melanogaster. PMID:12586711

  11. The Adh1 gene of the fungus Metarhizium anisopliae is expressed during insect colonization and required for full virulence.

    PubMed

    Callejas-Negrete, Olga Alicia; Torres-Guzmán, Juan Carlos; Padilla-Guerrero, Israel Enrique; Esquivel-Naranjo, Ulises; Padilla-Ballesteros, Maria Fernanda; García-Tapia, Adriana; Schrank, Augusto; Salazar-Solís, Eduardo; Gutiérrez-Corona, Félix; González-Hernández, Gloria Angélica

    2015-03-01

    Zymography of alcohol dehydrogenase (ADH) activity in the entomopathogenic fungus Metarhizium anisopliae grown under various conditions revealed that micro-aerobic growth was associated with increased ADH activity. The major ADH protein, AdhIp, was purified to homogeneity by affinity chromatography and has an estimated molecular weight of 41kDa and an isoelectric point (pI) of 6.4. Peptide mass fingerprint analysis allowed the identification and cloning of the gene that encodes this protein, Adh1, as annotated in the M. anisopliae genome database. AdhIp is related to the medium-chain dehydrogenase/reductase (MDR)/zinc-dependent alcohol dehydrogenase-like family and contains conserved ADH sequence motifs, such as the zinc-containing ADH signature, the FAD/NAD binding domain and amino acid residues that are conserved in most microbial ADHs. Semi-quantitative RT-PCR analysis revealed that Adh1 gene expression occurs at low levels during early Plutella xylostella infection and that the Adh1 gene was primarily expressed at larval death and as mycelia emerge from the insect cuticle before conidiation. Antisense-RNA experiments indicated that NAD(+)-dependent ADH activity was diminished by 20-75% in the transformants, and the transformants that had lower ADH activity showed allyl alcohol resistance, which indicates that reduction in ADH activity also occurs in vivo. Bioassays performed using antisense adh1 transformants, which have lower ADH activity, showed that LC50 values were two to five times higher than the wild-type, indicating that AdhIp is required for full capability of the fungus to penetrate and/or colonize the insect.

  12. ADH2 gene polymorphisms are determinants of alcohol pharmacokinetics.

    PubMed

    Thomasson, H R; Beard, J D; Li, T K

    1995-12-01

    The class I hepatic alcohol dehydrogenases (ADHs) are primarily responsible for ethanol metabolism in humans. Genetic polymorphism at the ADH2 locus results in the inheritance of isozymes of strikingly different catalytic properties. The most common ADH2 allele, ADH2*1, encodes the low K(m) isozyme subunit beta 1. The ADH2*3 allele encodes a high-activity isozyme subunit of alcohol dehydrogenase, beta 3, identified in approximately 25% of African-Americans. The Vmax of beta 3 beta 3-ADH is 30 times greater than that of the beta 1 beta 1-ADH. Therefore, we hypothesized that the rate of ethanol metabolism, an important factor in the toxicity of ethanol, in persons with beta 3-containing ADH, either beta 3 beta 3- or beta 1 beta 3-ADH, would be faster than that of persons with only beta 1 beta 1-ADH. We tested this hypothesis with ethanol administered orally to healthy, young African-Americans. Three hundred and twenty-six African-American men and women were genotyped using polymerase chain reaction amplification of their leukocyte DNA followed by hybridization with allele-specific probes. One hundred twelve volunteers, selected by genotype, received an oral dose of ethanol designed to produce a blood ethanol concentration of 80 mg/dl (0.080 g/dl), when the blood alcohol concentration-time curve was extrapolated back to time 0. Ethanol metabolic rates (beta 60s) were determined in the 112 subjects from the slope of the pseudolinear portion of the blood ethanol concentration-time curves. The mean beta 60 of African-Americans having beta 3-containing ADH isozymes had significantly faster ethanol elimination rates than those with only beta 1 beta 1-ADH isozymes. There were no significant differences in body weight, ethanol intake in the week before testing, peak breath ethanol concentration, time to peak, or volume of distribution between the genotype groups. Within each of these groups, men had lower ethanol disappearance rates than women. These results demonstrate in

  13. Retinoic acid response element in the human alcohol dehydrogenase gene ADH3: implications for regulation of retinoic acid synthesis.

    PubMed Central

    Duester, G; Shean, M L; McBride, M S; Stewart, M J

    1991-01-01

    Retinoic acid regulation of one member of the human class I alcohol dehydrogenase (ADH) gene family was demonstrated, suggesting that the retinol dehydrogenase function of ADH may play a regulatory role in the biosynthetic pathway for retinoic acid. Promoter activity of human ADH3, but not ADH1 or ADH2, was shown to be activated by retinoic acid in transient transfection assays of Hep3B human hepatoma cells. Deletion mapping experiments identified a region in the ADH3 promoter located between -328 and -272 bp which confers retinoic acid activation. This region was also demonstrated to confer retinoic acid responsiveness on the ADH1 and ADH2 genes in heterologous promoter fusions. Within a 34-bp stretch, the ADH3 retinoic acid response element (RARE) contains two TGACC motifs and one TGAAC motif, both of which exist in RAREs controlling other genes. A block mutation of the TGACC sequence located at -289 to -285 bp eliminated the retinoic acid response. As assayed by gel shift DNA binding studies, the RARE region (-328 to -272 bp) of ADH3 bound the human retinoic acid receptor beta (RAR beta) and was competed for by DNA containing a RARE present in the gene encoding RAR beta. Since ADH catalyzes the conversion of retinol to retinal, which can be further converted to retinoic acid by aldehyde dehydrogenase, these results suggest that retinoic acid activation of ADH3 constitutes a positive feedback loop regulating retinoic acid synthesis. Images PMID:1996113

  14. Increased Variation in Adh Enzyme Activity in Drosophila Mutation-Accumulation Experiment Is Not Due to Transposable Elements at the Adh Structural Gene

    PubMed Central

    Aquadro, C. F.; Tachida, H.; Langley, C. H.; Harada, K.; Mukai, T.

    1990-01-01

    We present here a molecular analysis of the region surrounding the structural gene encoding alcohol dehydrogenase (Adh) in 47 lines of Drosophila melanogaster that have each accumulated mutations for 300 generations. While these lines show a significant increase in variation of alcohol dehydrogenase enzyme activity compared to control lines, we found no restriction map variation in a 13-kb region including the complete Adh structural gene and roughly 5 kb of both 5' and 3' sequences. Thus, the rapid accumulation of ADH activity variation after 28,200 allele generations does not appear to have been due to the mobilization of transposable elements into or out of the Adh structural gene region. PMID:1963870

  15. Molecular cloning and characterization of two inducible NAD⁺-adh genes encoding NAD⁺-dependent alcohol dehydrogenases from Acetobacter pasteurianus SKU1108.

    PubMed

    Masud, Uraiwan; Matsushita, Kazunobu; Theeragool, Gunjana

    2011-11-01

    The cytosolic NAD⁺-dependent alcohol dehydrogenases (NAD⁺-ADHs) are induced in the quinoprotein ADH-(PQQ-ADH) defective Acetobacter pasteurianus SKU1108 mutant during growth in an ethanol medium. The adhI and adhII genes, which encode NAD⁺-ADH I and ADH II, respectively, of this strain have been cloned and characterized. Sequence analyses have revealed that the adhI gene consists of 1029 bp coding for 342 amino acids, which share 99.71% identity with the same protein from A. pasteurianus IFO 3283. Conversely, the adhII gene is composed of 762 bp encoding for a polypeptide of 253 amino acids, which exhibit 99.60% identity with the A. pasteurianus IFO 3283 protein. ADH I is a member of the group I Zn-dependent long-chain ADHs, while the ADH II belongs to the group II short-chain dehydrogenase/reductase NAD⁺-ADHs. The NAD⁺-adh gene disruptants exhibited a growth reduction when grown in an ethanol medium. In Escherichia coli, ethanol induced adhI and adhII promoter activities by approximately 1.5 and 2.0 times, respectively, and the promoter activity of the adhII gene exceeded that of the adhI gene by approximately 3.5 times. The possible promoter regions of the adhI and adhII genes are located at approximately 81-105 bp and 74-92 bp, respectively, from their respective ATG start codons. Their repressor regions might be located in proximity to these promoters and may repress gene expression in the wild-type, where the membrane-bound ADH effectively functions.

  16. Rare ADH Variant Constellations are Specific for Alcohol Dependence

    PubMed Central

    Zuo, Lingjun; Zhang, Heping; Malison, Robert T.; Li, Chiang-Shan R.; Zhang, Xiang-Yang; Wang, Fei; Lu, Lingeng; Lu, Lin; Wang, Xiaoping; Krystal, John H.; Zhang, Fengyu; Deng, Hong-Wen; Luo, Xingguang

    2013-01-01

    Aims: Some of the well-known functional alcohol dehydrogenase (ADH) gene variants (e.g. ADH1B*2, ADH1B*3 and ADH1C*2) that significantly affect the risk of alcohol dependence are rare variants in most populations. In the present study, we comprehensively examined the associations between rare ADH variants [minor allele frequency (MAF) <0.05] and alcohol dependence, with several other neuropsychiatric and neurological disorders as reference. Methods: A total of 49,358 subjects in 22 independent cohorts with 11 different neuropsychiatric and neurological disorders were analyzed, including 3 cohorts with alcohol dependence. The entire ADH gene cluster (ADH7–ADH1C–ADH1B–ADH1A–ADH6–ADH4–ADH5 at Chr4) was imputed in all samples using the same reference panels that included whole-genome sequencing data. We stringently cleaned the phenotype and genotype data to obtain a total of 870 single nucleotide polymorphisms with 0< MAF <0.05 for association analysis. Results: We found that a rare variant constellation across the entire ADH gene cluster was significantly associated with alcohol dependence in European-Americans (Fp1: simulated global P = 0.045), European-Australians (Fp5: global P = 0.027; collapsing: P = 0.038) and African-Americans (Fp5: global P = 0.050; collapsing: P = 0.038), but not with any other neuropsychiatric disease. Association signals in this region came principally from ADH6, ADH7, ADH1B and ADH1C. In particular, a rare ADH6 variant constellation showed a replicable association with alcohol dependence across these three independent cohorts. No individual rare variants were statistically significantly associated with any disease examined after group- and region-wide correction for multiple comparisons. Conclusion: We conclude that rare ADH variants are specific for alcohol dependence. The ADH gene cluster may harbor a causal variant(s) for alcohol dependence. PMID:23019235

  17. Identification of Elymus (Triticeae, Poaceae) and its related genera genomes by RFLP analysis of PCR-amplified Adh genes.

    PubMed

    Liu, QuanLan; Zhang, NingNing; Li, Lei; Liu, Jie

    2010-10-01

    Elymus L. is the largest genus in Triticeae, containing about 150 species with four recognized genome donors (St, H, P, and W). Traditionally, the genome compound of this genus is identified based on cytological data. Recently, molecular phylogenetic analysis was used to investigate its genomic combination. Here we describe a restriction fragment length polymorphism (RFLP) assay based on digesting alcohol dehydrogenase (Adh) amplicons with two restriction enzyme combinations, EcoRI-HindIII and EcoRI-PstI, which easily can be used to distinguish Elymus and its closely related genera genomes. The method includes only four steps: (1) amplifying nuclear Adh genes with universal primers; (2) purifying and cloning PCR products; (3) digesting plasmids with restriction enzymes that identify a given genome; (4) running the digested products on an agarose gel and identify the sample based on the restriction profiles. Results showed that: (1) PCR products ranged from 1,200 to 2,000 bp; (2) Adh2 gene was amplified from all the tested genomes; Adh1 gene was amplified from almost all of the tested genomes except the W genome; Adh3 gene was amplified only from the St genome; (3) the EcoRI-HindIII combination was effective to distinguish different Adh gene types (Adh1, Adh2, and Adh3); (4) the Adh2-EcoRI-PstI fragments could be used to distinguish Elymus and its closely related genera genomes. Therefore, This RFLP assay provides an inexpensive and simple means of identifying Elymus genomes.

  18. Patterns of Amino Acid Evolution in the Drosophila ananassae Chimeric Gene, siren, Parallel Those of Other Adh-Derived Chimeras

    PubMed Central

    Shih, Hung-Jui; Jones, Corbin D.

    2008-01-01

    siren1 and siren2 are novel alcohol dehydrogenase (Adh)-derived chimeric genes in the Drosophila bipectinata complex. D. ananassae, however, harbors a single homolog of these genes. Like other Adh-derived chimeric genes, siren evolved adaptively shortly after it was formed. These changes likely shifted the catalytic activity of siren. PMID:18780749

  19. Isolation and Identification of Genes Activating Uas2-Dependent Adh2 Expression in Saccharomyces Cerevisiae

    PubMed Central

    Donoviel, M. S.; Young, E. T.

    1996-01-01

    Two cis-acting elements have been identified that act synergistically to regulate expression of the glucose-repressed alcohol dehydrogenase 2 (ADH2) gene. UAS1 is bound by the trans-activator Adr1p. UAS2 is thought to be the binding site for an unidentified regulatory protein. A genetic selection based on a UAS2-dependent ADH2 reporter was devised to isolate genes capable of activating UAS2-dependent transcription. One set of UAS2-dependent genes contained SPT6/CRE2/SSN20. Multicopy SPT6 caused improper expression of chromosomal ADH2. A second set of UAS2-dependent clones contained a previously uncharacterized open reading frame designated MEU1 (Multicopy Enhancer of UAS2). A frame shift mutation in MEU1 abolished its ability to activate UAS2-dependent gene expression. Multicopy MEU1 expression suppressed the constitutive ADH2 expression caused by cre2-1. Disruption of MEU1 reduced endogenous ADH2 expression about twofold but had no effect on cell viability or growth. No homologues of MEU1 were identified by low-stringency Southern hybridization of yeast genomic DNA, and no significant homologues were found in the sequence data bases. A MEU1/β-gal fusion protein was not localized to a particular region of the cell. MEU1 is linked to PPR1 on chromosome XII. PMID:8807288

  20. Primary structure and functional analysis of the lysis genes of Lactobacillus gasseri bacteriophage phi adh.

    PubMed Central

    Henrich, B; Binishofer, B; Bläsi, U

    1995-01-01

    The lysis genes of the Lactobacillus gasseri bacteriophage phi adh were isolated by complementation of a lambda Sam mutation in Escherichia coli. Nucleotide sequencing of a 1,735-bp DNA fragment revealed two adjacent coding regions of 342 bp (hol) and 951 bp (lys) in the same reading frame which appear to belong to a common transcriptional unit. Proteins corresponding to the predicted gene products, holin (12.9 kDa) and lysin (34.7 kDa), were identified by in vitro and in vivo expression of the cloned genes. The phi adh holin is a membrane-bound protein with structural similarity to lysis proteins of other phage, known to be required for the transit of murein hydrolases through the cytoplasmic membrane. The phi adh lysin shows homology with mureinolytic enzymes encoded by the Lactobacillus bulgaricus phage mv4, the Streptococcus pneumoniae phage Cp-1, Cp-7, and Cp-9, and the Lactococcus lactis phage phi LC3. Significant homology with the N termini of known muramidases suggests that phi adh lysin acts by a similar catalytic mechanism. In E. coli, the phi adh lysin seems to be associated with the total membrane fraction, from which it can be extracted with lauryl sarcosinate. Either one of the phi adh lysis proteins provoked lysis of E. coli when expressed along with holins or lysins of phage lambda or Bacillus subtilis phage phi 29. Concomitant expression of the combined holin and lysin functions of phi adh in E. coli, however, did not result in efficient cell lysis. PMID:7836307

  1. Rearrangement of Upstream Regulatory Elements Leads to Ectopic Expression of the Drosophila Mulleri Adh-2 Gene

    PubMed Central

    Falb, D.; Fischer, J.; Maniatis, T.

    1992-01-01

    The Adh-2 gene of Drosophila mulleri is expressed in the larval fat body and the adult fat body and hindgut, and a 1500-bp element located 2-3 kb upstream of the Adh-2 promoter is necessary for maximal levels of transcription. Previous work demonstrated that deletion of sequences between this upstream element and the Adh-2 promoter results in Adh-2 gene expression in a novel larval tissue, the middle midgut. In this study we show that the upstream element possesses all of the characteristics of a transcriptional enhancer: its activity is independent of orientation, it acts on a heterologous promoter, and it functions at various positions both 5' and 3' to the Adh-2 gene. Full enhancer function can be localized to a 750-bp element, although other regions possess some redundant activity. The ectopic expression pattern is dependent on the proximity of at least two sequence elements. Thus, tissue-specific transcription can involve complex proximity-dependent interactions among combinations of regulatory elements. PMID:1459428

  2. Compensatory evolution of a precursor messenger RNA secondary structure in the Drosophila melanogaster Adh gene

    PubMed Central

    Chen, Ying; Stephan, Wolfgang

    2003-01-01

    Evidence for the evolutionary maintenance of a hairpin structure possibly involved in intron processing had been found in intron 1 of the alcohol dehydrogenase gene (Adh) in diverse Drosophila species. In this study, the putative hairpin structure was evaluated systematically in Drosophila melanogaster by elimination of either side of the stem using site-directed mutagenesis. The effects of these mutations and the compensatory double mutant on intron splicing efficiency and ADH protein production were assayed in Drosophila melanogaster Schneider L2 cells and germ-line transformed adult flies. Mutations that disrupt the putative hairpin structure right upstream of the intron branch point were found to cause a significant reduction in both splicing efficiency and ADH protein production. In contrast, the compensatory double mutant that restores the putative hairpin structure was indistinguishable from the WT in both splicing efficiency and ADH level. It was also observed by mutational analysis that a more stable secondary structure (with a longer stem) in this intron decreases both splicing efficiency and ADH protein production. Implications for RNA secondary structure and intron evolution are discussed. PMID:12972637

  3. The two small introns of the Drosophila affinidisjuncta Adh gene are required for normal transcription.

    PubMed Central

    McKenzie, R W; Brennan, M D

    1996-01-01

    All Drosophila alcohol dehydrogenase (Adh) genes sequenced to date contain two small introns within the coding region. These are conserved in location and, to some extent, in sequence between the various species analyzed. To determine if these introns play a role in Adh gene expression, derivatives of the Drosophila affinidisjuncta Adh gene lacking one or both introns were constructed and analyzed by germline and transient transformation of Drosophila melanogaster. Removal of both introns lowered expression, whether measured by enzyme activity or by RNA levels. The decrease was seen in both germline transformed and transiently transformed larvae, with the effect being larger for germline transformants. Similar decreases (averaging 5-fold) were also seen at the embryonic and adult stages for germline transformants. Nuclear run-off transcription with nuclei from germline transformed embryos indicated that the reduction in RNA levels is due to decreased transcription. However, LacZ fusion constructs designed to test for the presence of a classical enhancer in the introns provided no evidence for such a mechanism. Removal of each intron individually resulted in more complex phenotypes. The introns have smaller, additive effects on expression in adults. In larvae, removal of the upstream intron significantly increases RNA levels but modestly decreases enzyme activity. Removal of the downstream intron lowers expression in both germline and transiently transformed larvae, but also increases position effects in germline transformants. Therefore, the small introns are clearly needed for optimal transcription of this Adh gene, but multiple mechanisms are involved. PMID:8836194

  4. Characterization of polymorphisms of genes ADH2, ADH3, ALDH2 and CYP2E1 and relationship to the alcoholism in a Colombian population

    PubMed Central

    Méndez, Claudia

    2015-01-01

    Objective: Identify and characterize polymorphisms of genes ADH2, ADH3, ALDH2 and CYP2E1 in a Colombian population residing in the city of Bogotá and determine its possible relationship to the alcoholism. Methods: ADH2, ADH3, ALDH2, and CYP2E1 genotypes a population of 148 individuals with non-problematic alcohol and 65 individuals with alcoholism were determined with TaqMan probes and PCR-RFLP. DNA was obtained from peripheral blood white cells. Results: Significant difference was found in family history of alcoholism and use of other psychoactive substances to compare alcoholics with controls. When allelic frequencies for each category (gender) were considered, frequency of A2 allele carriers in ADH2 was found higher in male patients than controls. In women, the relative frequency for c1 allele in CYP2E1 was lower in controls than alcoholics. The ALDH2 locus is monomorphic. No significant differences in allele distributions of the loci examined to compare two populations were observed, however when stratifying the same trend was found that these differences tended to be significant. Conclusions: This study allows us to conclude the positive association between family history of alcoholism and alcoholism suggesting that there is a favourable hereditary predisposition. Since substance dependence requires interaction of multiple genes, the combination of genotypes ADH2 * 2, CYP2E1 * 1 combined with genotype homozygous ALDH2 * 1 found in this study could be leading to the population to a potential risk to alcoholism. PMID:26848198

  5. Regulation of Expression of the adhE Gene, Encoding Ethanol Oxidoreductase in Escherichia coli: Transcription from a Downstream Promoter and Regulation by Fnr and RpoS

    PubMed Central

    Membrillo-Hernández, Jorge; Lin, E. C. C.

    1999-01-01

    The adhE gene of Escherichia coli, located at min 27 on the chromosome, encodes the bifunctional NAD-linked oxidoreductase responsible for the conversion of acetyl-coenzyme A to ethanol during fermentative growth. The expression of adhE is dependent on both transcriptional and posttranscriptional controls and is about 10-fold higher during anaerobic than during aerobic growth. Two putative transcriptional start sites have been reported: one at position −292 and the other at −188 from the translational start codon ATG. In this study we show, by using several different transcriptional and translational fusions to the lacZ gene, that both putative transcriptional start sites can be functional and each site can be redox regulated. Although both start sites are NarL repressible in the presence of nitrate, Fnr activates only the −188 start site and Fis is required for the transcription of only the −292 start site. In addition, it was discovered that RpoS activates adhE transcription at both start sites. Under all experimental conditions tested, however, only the upstream start site is active. Available evidence indicates that under those conditions, the upstream promoter region acts as a silencer of the downstream transcriptional start site. Translation of the mRNA starting at −292, but not the one starting at −188, requires RNase III. The results support the previously postulated ribosomal binding site (RBS) occlusion model, according to which RNase III cleavage is required to release the RBS from a stem-loop structure in the long transcript. PMID:10601216

  6. Copy number lability and evolutionary dynamics of the Adh gene family in diploid and tetraploid cotton (Gossypium).

    PubMed Central

    Small, R L; Wendel, J F

    2000-01-01

    Nuclear-encoded genes exist in families of various sizes. To further our understanding of the evolutionary dynamics of nuclear gene families we present a characterization of the structure and evolution of the alcohol dehydrogenase (Adh) gene family in diploid and tetraploid members of the cotton genus (Gossypium, Malvaceae). A PCR-based approach was employed to isolate and sequence multiple Adh gene family members, and Southern hybridization analyses were used to document variation in gene copy number. Adh gene copy number varies among Gossypium species, with diploids containing at least seven Adh loci in two primary gene lineages. Allotetraploid Gossypium species are inferred to contain at least 14 loci. Intron lengths vary markedly between loci, and one locus has lost two introns usually found in other plant Adh genes. Multiple examples of apparent gene duplication events were observed and at least one case of pseudogenization and one case of gene elimination were also found. Thus, Adh gene family structure is dynamic within this single plant genus. Evolutionary rate estimates differ between loci and in some cases between organismal lineages at the same locus. We suggest that dynamic fluctuation in copy number will prove common for nuclear genes, and we discuss the implications of this perspective for inferences of orthology and functional evolution. PMID:10924485

  7. An experimental test for lineage-specific position effects on alcohol dehydrogenase (Adh) genes in Drosophila

    PubMed Central

    Siegal, Mark L.; Hartl, Daniel L.

    1998-01-01

    Independent transgene insertions differ in expression based on their location in the genome; these position effects are of interest because they reflect the influence of genome organization on gene regulation. Position effects also represent potentially insurmountable obstacles to the rigorous functional comparison of homologous genes from different species because (i) quantitative variation in expression of each gene across genomic positions (generalized position effects, or GPEs) may overwhelm differences between the genes of interest, or (ii) divergent genes may be differentially sensitive to position effects, reflecting unique interactions between each gene and its genomic milieu (lineage-specific position effects, or LSPEs). We have investigated both types of position-effect variation by applying our method of transgene coplacement, which allows comparisons of transgenes in the same position in the genome of Drosophila melanogaster. Here we report an experimental test for LSPE in Drosophila. The alcohol dehydrogenase (Adh) genes of D. melanogaster and Drosophila affinidisjuncta differ in both tissue distribution and amounts of ADH activity. Despite this striking regulatory divergence, we found a very high correlation in overall ADH activity between the genes of the two species when placed in the same genomic position as assayed in otherwise Adh-null adults and larvae. These results argue against the influence of LSPE for these sequences, although the effects of GPE are significant. Our new findings validate the coplacement approach and show that it greatly magnifies the power to detect differences in expression between transgenes. Transgene coplacement thus dramatically extends the range of functional and evolutionary questions that can be addressed by transgenic technology. PMID:9861000

  8. Redundant cis-acting elements control expression of the Drosophila affinidisjuncta Adh gene in the larval fat body.

    PubMed Central

    McKenzie, R W; Hu, J; Brennan, M D

    1994-01-01

    The alcohol dehydrogenase (Adh) gene in the Hawaiian species of fruit fly, Drosophila affinidisjuncta, like the Adh genes from all Drosophila species analyzed, is expressed at high levels in the larval fat body via a larval-specific promoter. To identify the cis-acting elements involved in this highly conserved aspect of Adh gene expression, deleted D. affinidisjuncta genes were introduced into D. melanogaster by somatic transformation. Unlike previously described methods, this transformation system allows analysis of Adh gene expression specifically in the larval fat body. The arrangement of sequences influencing expression of the proximal promoter of this gene in the larval fat body differs markedly from that described for the Adh gene from the distant relative, D. melanogaster. Multiple redundant elements dispersed 5' and 3' to the gene, only some of which map to regions carrying evolutionarily conserved sequences, affect expression in the fat body. D. affinidisjuncta employs a novel mode of Adh gene regulation in which the proximal promoter is influenced by sequences having roles in expression of the distal promoter. This gene is also unique in that far upstream sequences can compensate for loss of sequences within 200 bp of the proximal RNA start site. Furthermore, expression is influenced in an unusual, context-dependent manner by a naturally-occurring 3' duplication of the proximal promoter--a feature found only in Hawaiian species. Images PMID:8165141

  9. A phylogenetic analysis of the genus Fragaria (strawberry) using intron-containing sequence from the ADH-1 gene.

    PubMed

    DiMeglio, Laura M; Staudt, Günter; Yu, Hongrun; Davis, Thomas M

    2014-01-01

    The genus Fragaria encompasses species at ploidy levels ranging from diploid to decaploid. The cultivated strawberry, Fragaria×ananassa, and its two immediate progenitors, F. chiloensis and F. virginiana, are octoploids. To elucidate the ancestries of these octoploid species, we performed a phylogenetic analysis using intron-containing sequences of the nuclear ADH-1 gene from 39 germplasm accessions representing nineteen Fragaria species and one outgroup species, Dasiphora fruticosa. All trees from Maximum Parsimony and Maximum Likelihood analyses showed two major clades, Clade A and Clade B. Each of the sampled octoploids contributed alleles to both major clades. All octoploid-derived alleles in Clade A clustered with alleles of diploid F. vesca, with the exception of one octoploid allele that clustered with the alleles of diploid F. mandshurica. All octoploid-derived alleles in clade B clustered with the alleles of only one diploid species, F. iinumae. When gaps encoded as binary characters were included in the Maximum Parsimony analysis, tree resolution was improved with the addition of six nodes, and the bootstrap support was generally higher, rising above the 50% threshold for an additional nine branches. These results, coupled with the congruence of the sequence data and the coded gap data, validate and encourage the employment of sequence sets containing gaps for phylogenetic analysis. Our phylogenetic conclusions, based upon sequence data from the ADH-1 gene located on F. vesca linkage group II, complement and generally agree with those obtained from analyses of protein-encoding genes GBSSI-2 and DHAR located on F. vesca linkage groups V and VII, respectively, but differ from a previous study that utilized rDNA sequences and did not detect the ancestral role of F. iinumae.

  10. A Phylogenetic Analysis of the Genus Fragaria (Strawberry) Using Intron-Containing Sequence from the ADH-1 Gene

    PubMed Central

    DiMeglio, Laura M.; Yu, Hongrun; Davis, Thomas M.

    2014-01-01

    The genus Fragaria encompasses species at ploidy levels ranging from diploid to decaploid. The cultivated strawberry, Fragaria×ananassa, and its two immediate progenitors, F. chiloensis and F. virginiana, are octoploids. To elucidate the ancestries of these octoploid species, we performed a phylogenetic analysis using intron-containing sequences of the nuclear ADH-1 gene from 39 germplasm accessions representing nineteen Fragaria species and one outgroup species, Dasiphora fruticosa. All trees from Maximum Parsimony and Maximum Likelihood analyses showed two major clades, Clade A and Clade B. Each of the sampled octoploids contributed alleles to both major clades. All octoploid-derived alleles in Clade A clustered with alleles of diploid F. vesca, with the exception of one octoploid allele that clustered with the alleles of diploid F. mandshurica. All octoploid-derived alleles in clade B clustered with the alleles of only one diploid species, F. iinumae. When gaps encoded as binary characters were included in the Maximum Parsimony analysis, tree resolution was improved with the addition of six nodes, and the bootstrap support was generally higher, rising above the 50% threshold for an additional nine branches. These results, coupled with the congruence of the sequence data and the coded gap data, validate and encourage the employment of sequence sets containing gaps for phylogenetic analysis. Our phylogenetic conclusions, based upon sequence data from the ADH-1 gene located on F. vesca linkage group II, complement and generally agree with those obtained from analyses of protein-encoding genes GBSSI-2 and DHAR located on F. vesca linkage groups V and VII, respectively, but differ from a previous study that utilized rDNA sequences and did not detect the ancestral role of F. iinumae. PMID:25078607

  11. Polymorphisms in Alcohol Metabolism Genes ADH1B and ALDH2, Alcohol Consumption and Colorectal Cancer

    PubMed Central

    Crous-Bou, Marta; Rennert, Gad; Cuadras, Daniel; Salazar, Ramon; Cordero, David; Saltz Rennert, Hedy; Lejbkowicz, Flavio; Kopelovich, Levy; Monroe Lipkin, Steven; Bernard Gruber, Stephen; Moreno, Victor

    2013-01-01

    Background Colorectal cancer (CRC) is a leading cause of cancer death worldwide. Epidemiological risk factors for CRC included alcohol intake, which is mainly metabolized to acetaldehyde by alcohol dehydrogenase and further oxidized to acetate by aldehyde dehydrogenase; consequently, the role of genes in the alcohol metabolism pathways is of particular interest. The aim of this study is to analyze the association between SNPs in ADH1B and ALDH2 genes and CRC risk, and also the main effect of alcohol consumption on CRC risk in the study population. Methodology/Principal Findings SNPs from ADH1B and ALDH2 genes, included in alcohol metabolism pathway, were genotyped in 1694 CRC cases and 1851 matched controls from the Molecular Epidemiology of Colorectal Cancer study. Information on clinicopathological characteristics, lifestyle and dietary habits were also obtained. Logistic regression and association analysis were conducted. A positive association between alcohol consumption and CRC risk was observed in male participants from the Molecular Epidemiology of Colorectal Cancer study (MECC) study (OR = 1.47; 95%CI = 1.18-1.81). Moreover, the SNPs rs1229984 in ADH1B gene was found to be associated with CRC risk: under the recessive model, the OR was 1.75 for A/A genotype (95%CI = 1.21-2.52; p-value = 0.0025). A path analysis based on structural equation modeling showed a direct effect of ADH1B gene polymorphisms on colorectal carcinogenesis and also an indirect effect mediated through alcohol consumption. Conclusions/Significance Genetic polymorphisms in the alcohol metabolism pathways have a potential role in colorectal carcinogenesis, probably due to the differences in the ethanol metabolism and acetaldehyde oxidation of these enzyme variants. PMID:24282520

  12. Genetic Association and Gene-Gene Interaction Reveal Genetic Variations in ADH1B, GSTM1 and MnSOD Independently Confer Risk to Alcoholic Liver Diseases in India

    PubMed Central

    Mukhopadhyay, Indranil; Chatterjee, Ankita; Das, Kausik; Bhowmik, Pradip; Das, Soumyajit; Basu, Priyadarshi; Santra, Amal K.; Datta, Simanti; Dhali, Gopal Krishna; Chowdhury, Abhijit; Banerjee, Soma

    2016-01-01

    Genetic susceptibility is an important modifier of clinical outcome and natural history of progression in Alcoholic liver disease (ALD). While the significance of ethnicity in this evolution is very clear, subtle inter-individual genetic variant(s) might be important and thus we investigated those in an Indian population. Fourteen markers were genotyped within two alcohol metabolism genes [Alcohol dehydrogenase (ADH) gene clusters (ADH1B and ADH1C) and Aldehyde dehydrogenase (ALDH2)], one microsomal ethanol oxidizing enzyme cytochrome p450 (CYP2E1) and three oxidative stress response (OSR) genes (MnSOD, GSTT1 and GSTM1) among 490 Bengali individuals (322 ALD and 168 control) from Eastern and North-Eastern India and validation was performed in a new cohort of 150 Bengali patients including 100 ALD and 50 advanced non-alcoholic steatohepatitis (NASH). Out of 14 genetic variants, carriage of 5 genotypes (rs2066701CC in ADH1B, rs1693425TT in ADH1C, rs4880TT in MnSOD and GSTT1/GSTM1 null, p-value <0.05) were noted significantly higher among ALD patients while inter or intra group gene-gene interaction analysis revealed that addition of risk genotype of any OSR gene enhanced the possibility of ALD synergistically. Multiple logistic regression analysis showed independent association of rs2066701CC, rs4880TT and GSTM1 null genotype with ALD while lower frequencies of those genotypes in advanced NASH patients further confirmed their causal relation to ALD. Thus these findings suggest that the three variants of ADH1C, MnSOD and GSTM1 can be used to identify individuals who are at high risk to develop ALD and may be helpful in proper management of Indian alcoholics. PMID:26937962

  13. Genetic Association and Gene-Gene Interaction Reveal Genetic Variations in ADH1B, GSTM1 and MnSOD Independently Confer Risk to Alcoholic Liver Diseases in India.

    PubMed

    Roy, Neelanjana; Dasgupta, Debanjali; Mukhopadhyay, Indranil; Chatterjee, Ankita; Das, Kausik; Bhowmik, Pradip; Das, Soumyajit; Basu, Priyadarshi; Santra, Amal K; Datta, Simanti; Dhali, Gopal Krishna; Chowdhury, Abhijit; Banerjee, Soma

    2016-01-01

    Genetic susceptibility is an important modifier of clinical outcome and natural history of progression in Alcoholic liver disease (ALD). While the significance of ethnicity in this evolution is very clear, subtle inter-individual genetic variant(s) might be important and thus we investigated those in an Indian population. Fourteen markers were genotyped within two alcohol metabolism genes [Alcohol dehydrogenase (ADH) gene clusters (ADH1B and ADH1C) and Aldehyde dehydrogenase (ALDH2)], one microsomal ethanol oxidizing enzyme cytochrome p450 (CYP2E1) and three oxidative stress response (OSR) genes (MnSOD, GSTT1 and GSTM1) among 490 Bengali individuals (322 ALD and 168 control) from Eastern and North-Eastern India and validation was performed in a new cohort of 150 Bengali patients including 100 ALD and 50 advanced non-alcoholic steatohepatitis (NASH). Out of 14 genetic variants, carriage of 5 genotypes (rs2066701CC in ADH1B, rs1693425TT in ADH1C, rs4880TT in MnSOD and GSTT1/GSTM1 null, p-value <0.05) were noted significantly higher among ALD patients while inter or intra group gene-gene interaction analysis revealed that addition of risk genotype of any OSR gene enhanced the possibility of ALD synergistically. Multiple logistic regression analysis showed independent association of rs2066701CC, rs4880TT and GSTM1 null genotype with ALD while lower frequencies of those genotypes in advanced NASH patients further confirmed their causal relation to ALD. Thus these findings suggest that the three variants of ADH1C, MnSOD and GSTM1 can be used to identify individuals who are at high risk to develop ALD and may be helpful in proper management of Indian alcoholics.

  14. Association of ADH and ALDH Genes With Alcohol Dependence in the Irish Affected Sib Pair Study of Alcohol Dependence (IASPSAD) Sample

    PubMed Central

    Kuo, Po-Hsiu; Kalsi, Gursharan; Prescott, Carol A.; Hodgkinson, Colin A.; Goldman, David; van den Oord, Edwin J.; Alexander, Jeffry; Jiang, Cizhong; Sullivan, Patrick F.; Patterson, Diana G.; Walsh, Dermot; Kendler, Kenneth S.; Riley, Brien P.

    2008-01-01

    Background: The genes coding for ethanol metabolism enzymes [alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH)] have been widely studied for their influence on the risk to develop alcohol dependence (AD). However, the relation between polymorphisms of these metabolism genes and AD in Caucasian subjects has not been clearly established. The present study examined evidence for the association of alcohol metabolism genes with AD in the Irish Affected Sib Pair Study of alcohol dependence. Methods: We conducted a case–control association study with 575 independent subjects who met Diagnostic and Statistical Manual of Mental Disorders, 4th Edition, AD diagnosis and 530 controls. A total of 77 single nucleotide polymorphisms (SNPs) in the seven ADH (ADH1-7) and two ALDH genes (ALDH1A1 and ALDH2) were genotyped using the Illumina GoldenGate protocols. Several statistical procedures were implemented to control for false discoveries. Results: All markers with minor allele frequency greater than 0.01 were in Hardy–Weinberg equilibrium. Numerous SNPs in ADH genes showed association with AD, including one marker in the coding region of ADH1C (rs1693482 in exon6, Ile271Gln). Haplotypic association was observed in the ADH5 and ADH1C genes, and in a long haplotype block formed by the ADH1A and ADH1B loci. We detected two significant interactions between pairs of markers in intron 6 of ADH6 and intron 12 of ALDH2 (p = 5 × 10−5), and 5′ of both ADH4 and ADH1A (p = 2 × 10−4). Conclusion: We found evidence for the association of several ADH genes with AD in a sample of Western European origin. The significant interaction effects between markers in ADH and ALDH genes suggest possible epistatic roles between alcohol metabolic enzymes in the risk for AD. PMID:18331377

  15. The alcohol dehydrogenase gene adh1 is induced in Aspergillus flavus grown on medium conducive to aflatoxin biosynthesis.

    PubMed Central

    Woloshuk, C P; Payne, G A

    1994-01-01

    An Aspergillus flavus cDNA library was screened by differential hybridization to isolate clones corresponding to genes that are actively transcribed under culture conditions conducive to aflatoxin biosynthesis. One clone with a 1.28-kb insert was isolated, and its nucleotide sequence was determined. The nucleotide sequence of this clone had 75% DNA identity to those of the alcohol dehydrogenase genes from Aspergillus nidulans, and the putative polypeptide translated from the cDNA sequence had 82% similarity with the amino acid sequences of the A. nidulans proteins. Thus, this gene has been designated adh1. Southern hybridization analysis of genomic DNA from A. flavus indicated that there was one copy of the adh1 gene. Northern (RNA) hybridization analysis indicated that the adh1 transcript accumulated in culture medium conducive to aflatoxin production and the timing of accumulation of adh1 transcripts was similar to that for aflatoxin. Fusion of the promoter region of adh1 to a beta-glucuronidase reporter gene indicated that accumulation of the adh1 transcript was the result of transcriptional activation. These molecular data support previous physiological evidence that showed the importance of carbohydrate metabolism during aflatoxin biosynthesis. Images PMID:8135521

  16. Differential interactions of promoter elements in stress responses of the Arabidopsis Adh gene.

    PubMed Central

    Dolferus, R; Jacobs, M; Peacock, W J; Dennis, E S

    1994-01-01

    The Adh (alcohol dehydrogenase, EC 1.1.1.1.) gene from Arabidopsis thaliana (L.) Heynh. can be induced by dehydration and cold, as well as by hypoxia. A 1-kb promoter fragment (CADH: -964 to +53) is sufficient to confer the stress induction and tissue-specific developmental expression characteristics of the Adh gene to a beta-glucuronidase reporter gene. Deletion mapping of the 5' end and site-specific mutagenesis identified four regions of the promoter essential for expression under the three stress conditions. Some sequence elements are important for response to all three stress treatments, whereas others are stress specific. The most critical region essential for expression of the Arabidopsis Adh promoter under all three environmental stresses (region IV: -172 to -141) contains sequences homologous to the GT motif (-160 to -152) and the GC motif (-147 to -144) of the maize Adh1 anaerobic responsive element. Region III (-235 to -172) contains two regions shown by R.J. Ferl and B.H. Laughner ([1989] Plant Mol Biol 12: 357-366) to bind regulatory proteins; mutation of the G-box-1 region (5'-CCACGTGG-3', -216 to -209) does not affect expression under uninduced or hypoxic conditions, but significantly reduces induction by cold stress and, to a lesser extent, by dehydration stress. Mutation of the other G-box-like sequence (G-box-2: 5'-CCAAGTGG-3', -193 to -182) does not change hypoxic response and affects cold and dehydration stress only slightly. G-box-2 mutations also promote high levels of expression under uninduced conditions. Deletion of region I (-964 to -510) results in increased expression under uninduced and all stress conditions, suggesting that this region contains a repressor binding site. Region II (-510 to -384) contains a positive regulatory element and is necessary for high expression levels under all treatments. PMID:7972489

  17. Multiple Cis-Acting Sequences Contribute to Evolved Regulatory Variation for Drosophila Adh Genes

    PubMed Central

    Fang, X. M.; Brennan, M. D.

    1992-01-01

    Drosophila affinidisjuncta and Drosophila hawaiiensis are closely related species that display distinct tissue-specific expression patterns for their homologous alcohol dehydrogenase genes (Adh genes). In Drosophila melanogaster transformants, both genes are expressed at high levels in the larval and adult fat bodies, but the D. affinidisjuncta gene is expressed 10-50-fold more strongly in the larval and adult midguts and Malpighian tubules. The present study reports the mapping of cis-acting sequences contributing to the regulatory differences between these two genes in transformants. Chimeric genes were constructed and introduced into the germ line of D. melanogaster. Stage- and tissue-specific expression patterns were determined by measuring steady-state RNA levels in larvae and adults. Three portions of the promoter region make distinct contributions to the tissue-specific regulatory differences between the native genes. Sequences immediately upstream of the distal promoter have a strong effect in the adult Malpighian tubules, while sequences between the two promoters are relatively important in the larval Malpighian tubules. A third gene segment, immediately upstream of the proximal promoter, influences levels of the proximal Adh transcript in all tissues and developmental stages examined, and largely accounts for the regulatory difference in the larval and adult midguts. However, these as well as other sequences make smaller contributions to various aspects of the tissue-specific regulatory differences. In addition, some chimeric genes display aberrant RNA levels for the whole organism, suggesting close physical association between sequences involved in tissue-specific regulatory differences and those important for Adh expression in the larval and adult fat bodies. PMID:1644276

  18. The Adh-related gene of Drosophila melanogaster is expressed as a functional dicistronic messenger RNA: multigenic transcription in higher organisms.

    PubMed Central

    Brogna, S; Ashburner, M

    1997-01-01

    Essentially all eukaryotic cellular mRNAs are monocistronic, and are usually transcribed individually. Two tandemly arranged Drosophila genes, alcohol dehydrogenase (Adh) and Adh-related (Adhr), are transcribed as a dicistronic transcript. From transcripts initiated from the Adh promoter, two classes of mRNA are accumulated, one is monocistronic and encodes Adh alone, the other is dicistronic and includes the open reading frames of both Adh and Adhr. The dicistronic transcript is found in polysomes and the Adhr protein product is detected by antibody staining. We present evidence that the accumulation of the dicistronic mRNA is controlled at the level of the 3' end processing. PMID:9155028

  19. Associations between DNA Sequence Variation and Variation in Expression of the Adh Gene in Natural Populations of Drosophila Melanogaster

    PubMed Central

    Laurie, C. C.; Bridgham, J. T.; Choudhary, M.

    1991-01-01

    A large part of the genetic variation in alcohol dehydrogenase (ADH) activity level in natural populations of Drosophila melanogaster is associated with segregation of an amino acid replacement polymorphism at nucleotide 1490, which generates a difference in electrophoretic mobility. Part of the allozymic difference in activity level is due to a catalytic efficiency difference, which is also caused by the amino acid replacement, and part is due to a difference in the concentration of ADH protein. A previous site-directed in vitro mutagenesis experiment clearly demonstrated that the amino acid replacement has no effect on the concentration of ADH protein, nor does a strongly associated silent polymorphism at nucleotide 1443. Here we analyze associations between polymorphisms within the Adh gene and variation in ADH protein level for a number of chromosomes derived from natural populations. A sequence length polymorphism within the first intron of the distal (adult) transcript, &1, is in strong linkage disequilibrium with the amino acid replacement. Among a sample of 46 isochromosomal lines analyzed, all but one of the 14 Fast lines have &1 and all but one of the 32 Slow lines lack &1. The exceptional Fast line has an unusually low level of ADH protein (typical of Slow lines) and the exceptional Slow line has an unusually high level (typical of Fast lines). These results suggest that the &1 polymorphism may be responsible for the average difference in ADH protein between the allozymic classes. A previous experiment localized the effect on ADH protein to a 2.3-kb restriction fragment. DNA sequences of this fragment from several alleles of each allozymic type indicate that no other polymorphisms within this region are as closely associated with the ADH protein level difference as the &1 polymorphism. PMID:1683848

  20. The Arabidopsis Adh gene exhibits diverse nucleosome arrangements within a small DNase I-sensitive domain.

    PubMed Central

    Vega-Palas, M A; Ferl, R J

    1995-01-01

    The alcohol dehydrogenase (Adh) gene from Arabidopsis shows enhanced sensitivity to DNase I in cells that express the gene. This generalized sensitivity to DNase I is demarcated by position -500 on the 5' side and the end of the mRNA on the 3' side. Thus, the gene defined as the promoter and mRNA coding region corresponds very closely in size with the gene defined as a nuclease-sensitive domain. This is a remarkably close correspondence between a sensitive domain and a eukaryotic transcriptional unit, because previously reported DNase I-sensitive domains include large regions of DNA that are not transcribed. Nucleosomes are present in the coding region of the Adh gene when it is expressed, indicating that the transcriptional elongation process causes nucleosome disruption rather than release of nucleosomes from the coding region. In addition, the regulatory region contains a loosely positioned nucleosome that is separated from adjacent nucleosomes by internucleosomic DNA segments longer than the average linker DNA in bulk chromatin. This specific array of nucleosomes coexists with bound transcription factors that could contribute to the organization of the nucleosome arrangement. These results enhance our understanding of the complex interactions among DNA, nucleosomes, and transcription factors during gene expression in plants. PMID:8535143

  1. Pleiotropic effect of disrupting a conserved sequence involved in a long-range compensatory interaction in the Drosophila Adh gene.

    PubMed Central

    Baines, John F; Parsch, John; Stephan, Wolfgang

    2004-01-01

    Recent advances in experimental analyses of the evolution of RNA secondary structures suggest a more complex scenario than that typically considered by Kimura's classical model of compensatory evolution. In this study, we examine one such case in more detail. Previous experimental analysis of long-range compensatory interactions between the two ends of Drosophila Adh mRNA failed to fit the classical model of compensatory evolution. To further investigate and verify long-range pairing in Drosophila Adh with respect to models of compensatory evolution and its potential functional role, we introduced site-directed mutations in the Drosophila melanogaster Adh gene. We explore two alternative hypotheses for why previous analysis of long-range compensatory interactions failed to fit the classical model. Specifically, we investigate whether the disruption of a conserved short-range pairing within Adh exon 2 has an effect on Adh expression or if there is a dual functional role of a conserved sequence in the 3'-UTR in both long-range pairing and the negative regulation of Adh expression. We find that a classical result was not observed due to the pleiotropic effect of changing a nucleotide involved in both long-range base pairing and the negative regulation of gene expression. PMID:15020421

  2. Cloning of the Arabidopsis and Rice Formaldehyde Dehydrogenase Genes: Implications for the Origin of Plant Adh Enzymes

    PubMed Central

    Dolferus, R.; Osterman, J. C.; Peacock, W. J.; Dennis, E. S.

    1997-01-01

    This article reports the cloning of the genes encoding the Arabidopsis and rice class III ADH enzymes, members of the alcohol dehydrogenase or medium chain reductase/dehydrogenase superfamily of proteins with glutathione-dependent formaldehyde dehydrogenase activity (GSH-FDH). Both genes contain eight introns in exactly the same positions, and these positions are conserved in plant ethanol-active Adh genes (class P). These data provide further evidence that plant class P genes have evolved from class III genes by gene duplication and acquisition of new substrate specificities. The position of introns and similarities in the nucleic acid and amino acid sequences of the different classes of ADH enzymes in plants and humans suggest that plant and animal class III enzymes diverged before they duplicated to give rise to plant and animal ethanol-active ADH enzymes. Plant class P ADH enzymes have gained substrate specificities and evolved promoters with different expression properties, in keeping with their metabolic function as part of the alcohol fermentation pathway. PMID:9215914

  3. Regulation of adhE (Encoding Ethanol Oxidoreductase) by the Fis Protein in Escherichia coli

    PubMed Central

    Membrillo-Hernández, Jorge; Kwon, Ohsuk; De Wulf, Peter; Finkel, Steven E.; Lin, E. C. C.

    1999-01-01

    The adhE gene of Escherichia coli encodes a multifunctional ethanol oxidoreductase whose expression is 10-fold higher under anaerobic than aerobic conditions. Transcription of the gene is under the negative control of the Cra (catabolite repressor-activator) protein, whereas translation of the adhE mRNA requires processing by RNase III. In this report, we show that the expression of adhE also depends on the Fis (factor for inversion stimulation) protein. A strain bearing a fis::kan null allele failed to grow anaerobically on glucose solely because of inadequate adhE transcription. However, fis expression itself is not under redox control. Sequence inspection of the adhE promoter revealed three potential Fis binding sites. Electrophoretic mobility shift analysis, using purified Fis protein and adhE promoter DNA, showed three different complexes. PMID:10572146

  4. Sequences upstream of the homologous cis-elements of the Adh adult enhancer of Drosophila are required for maximal levels of Adh gene transcription in adults of Scaptodrosophila lebanonensis.

    PubMed Central

    Papaceit, Montserrat; Orengo, Dorcas; Juan, Elvira

    2004-01-01

    The evolution of cis-regulatory elements is of particular interest for our understanding of the evolution of gene regulation. The Adh gene of Drosophilidae shows interspecific differences in tissue-specific expression and transcript levels during development. In Scaptodrosophila lebanonensis adults, the level of distal transcripts is maximal between the fourth and eighth day after eclosion and is around five times higher than that in D. melanogaster Adh(S). To examine whether these quantitative differences are regulated by sequences lying upstream of the distal promoter, we performed in vitro deletion mutagenesis of the Adh gene of S. lebanonensis, followed by P-element-mediated germ-line transformation. All constructs included, as a cotransgene, a modified Adh gene of D. melanogaster (dAdh) in a fixed position and orientation that acted as a chromosomal position control. Using this approach, we have identified a fragment of 1.5 kb in the 5' region, 830 bp upstream of the distal start site, which is required to achieve maximal levels of distal transcript in S. lebanonensis. The presence of this fragment produces a 3.5-fold higher level of distal mRNA (as determined by real time quantitative PCR) compared with the D. melanogaster dAdh cotransgene. This region contains the degenerated end of a minisatellite sequence expanding farther upstream and does not correspond to the Adh adult enhancer (AAE) of D. melanogaster. Indeed, the cis-regulatory elements of the AAE have been identified by phylogenetic footprinting within the region 830 bp upstream of the distal start site of S. lebanonensis. Furthermore, the deletions Delta-830 and Delta-2358 yield the same pattern of tissue-specific expression, indicating that all tissue-specific elements are contained within the region 830 bp upstream of the distal start site. PMID:15166155

  5. Analysis of Sequences Regulating Larval Expression of the Adh Gene of Drosophila Melanogaster

    PubMed Central

    Shen, NLL.; Hotaling, E. C.; Subrahmanyam, G.; Martin, P. F.; Sofer, W.

    1991-01-01

    The effects of a series of eight, 50 base pair internal deletions in the 5' region upstream of the proximal transcription start site of the Adh gene of Drosophila melanogaster were examined in a quantitative assay. Mixtures of two plasmids, one bearing a deleted gene, the other with an intact reference gene, were injected into alcohol dehydrogenase-negative embryos. Third instar larvae of the injected generation were assayed for relative alcohol dehydrogenase enzyme activity. Quantitative analysis of the eight deletions indicated that two regions were required for any detectable enzyme activity and one region was required for appropriate tissue specificity. The remaining five deletions significantly decreased, but did not eliminate activity. When the deleted genes were placed on a plasmid with an intact reference gene, activities of all but one deletion were restored to levels equivalent to that of the intact reference gene (regardless of orientation). This restoration of activity did not occur when the regulatory region of the intact gene was replaced with the Hsp70 heat shock promoter nor when the 50-base pair deletion encompassed the region that includes the TATA sequence. The fact that seven of the eight deleted genes express activity in the presence of a reference gene on the same plasmid suggests that the deleted gene is controlled by regulatory elements in the reference gene. Further, these regulatory elements exhibit no preference for their own, more proximate, promoter. PMID:1752419

  6. Alcohol dehydrogenase gene ADH3 activates glucose alcoholic fermentation in genetically engineered Dekkera bruxellensis yeast.

    PubMed

    Schifferdecker, Anna Judith; Siurkus, Juozas; Andersen, Mikael Rørdam; Joerck-Ramberg, Dorte; Ling, Zhihao; Zhou, Nerve; Blevins, James E; Sibirny, Andriy A; Piškur, Jure; Ishchuk, Olena P

    2016-04-01

    Dekkera bruxellensis is a non-conventional Crabtree-positive yeast with a good ethanol production capability. Compared to Saccharomyces cerevisiae, its tolerance to acidic pH and its utilization of alternative carbon sources make it a promising organism for producing biofuel. In this study, we developed an auxotrophic transformation system and an expression vector, which enabled the manipulation of D. bruxellensis, thereby improving its fermentative performance. Its gene ADH3, coding for alcohol dehydrogenase, was cloned and overexpressed under the control of the strong and constitutive promoter TEF1. Our recombinant D. bruxellensis strain displayed 1.4 and 1.7 times faster specific glucose consumption rate during aerobic and anaerobic glucose fermentations, respectively; it yielded 1.2 times and 1.5 times more ethanol than did the parental strain under aerobic and anaerobic conditions, respectively. The overexpression of ADH3 in D. bruxellensis also reduced the inhibition of fermentation by anaerobiosis, the "Custer effect". Thus, the fermentative capacity of D. bruxellensis could be further improved by metabolic engineering. PMID:26743658

  7. Further clarification of the contribution of the ADH1C gene to vulnerability of alcoholism and selected liver diseases.

    PubMed

    Li, Dawei; Zhao, Hongyu; Gelernter, Joel

    2012-08-01

    The alcohol dehydrogenase 1C (ADH1C) subunit is an important member of the alcohol dehydrogenase family, a set of genes that plays a major role in the catabolism of ethanol. Numerous association studies have provided compelling evidence that ADH1C gene variation (formerly ADH3) is associated with altered genetic susceptibility to alcoholism and alcohol-related liver disease, cirrhosis, or pancreatitis. However, the results have been inconsistent, partially, because each study involved a limited number of subjects, and some were underpowered. Using cumulative data over the past two decades, this meta-analysis (6,796 cases and 6,938 controls) considered samples of Asian, European, African, and Native American origins to examine whether the aggregate genotype provide statistically significant evidence of association. The results showed strong evidence of association between ADH1C Ile350Val (rs698, formerly ADH1C *1/*2) and alcohol dependence (AD) and abuse in the combined studies. The overall allelic (Val vs. Ile or *2 vs. *1) P value was 1 × 10(-8) and odds ratio (OR) was 1.51 (1.31, 1.73). The Asian populations produced stronger evidence of association with an allelic P value of 4 × 10(-33) [OR 2.14 (1.89, 2.43)] with no evidence of heterogeneity, and the dominant and recessive models revealed even stronger effect sizes. The strong evidence remained when stricter criteria and sub-group analyses were applied, while Asians always showed stronger associations than other populations. Our findings support that ADH1C Ile may lower the risk of AD and alcohol abuse as well as alcohol-related cirrhosis in pooled populations, with the strongest and most consistent effects in Asians.

  8. Supervised clustering of genes

    PubMed Central

    Dettling, Marcel; Bühlmann, Peter

    2002-01-01

    Background We focus on microarray data where experiments monitor gene expression in different tissues and where each experiment is equipped with an additional response variable such as a cancer type. Although the number of measured genes is in the thousands, it is assumed that only a few marker components of gene subsets determine the type of a tissue. Here we present a new method for finding such groups of genes by directly incorporating the response variables into the grouping process, yielding a supervised clustering algorithm for genes. Results An empirical study on eight publicly available microarray datasets shows that our algorithm identifies gene clusters with excellent predictive potential, often superior to classification with state-of-the-art methods based on single genes. Permutation tests and bootstrapping provide evidence that the output is reasonably stable and more than a noise artifact. Conclusions In contrast to other methods such as hierarchical clustering, our algorithm identifies several gene clusters whose expression levels clearly distinguish the different tissue types. The identification of such gene clusters is potentially useful for medical diagnostics and may at the same time reveal insights into functional genomics. PMID:12537558

  9. Substitution rate comparisons between grasses and palms: synonymous rate differences at the nuclear gene Adh parallel rate differences at the plastid gene rbcL.

    PubMed Central

    Gaut, B S; Morton, B R; McCaig, B C; Clegg, M T

    1996-01-01

    A number of studies have noted that nucleotide substitution rates at the chloroplast-encoded rbcL locus violate the molecular clock principle. Substitution rate variation at this plastid gene is particularly pronounced between palms and grasses; for example, a previous study estimated that substitution rates in rbcL sequences are approximately 5-fold faster in grasses than in palms. To determine whether a proportionate change in substitution rates also occurs in plant nuclear genes, we characterized nucleotide substitution rates in palm and grass sequences for the nuclear gene Adh. In this article, we report that palm sequences evolve at a rate of 2.61 x 10(-9) substitution per synonymous site per year, a rate which is slower than most plant nuclear genes. Grass Adh sequences evolve approximately 2.5-fold faster than palms at synonymous sites. Thus, synonymous rates in nuclear Adh genes show a marked decrease in palms relative to grasses, paralleling the pattern found at the plastid rbcL locus. This shared pattern indicates that synonymous rates are correlated between a nuclear and a plastid gene. Remarkably, nonsynonymous rates do not show this correlation. Nonsynonymous rates vary between two duplicated grass Adh loci, and nonsynonymous rates at the palm Adh locus are not markedly reduced relative to grasses. Images Fig. 3 PMID:8816790

  10. Evidence of Positive Selection on a Class I ADH Locus

    PubMed Central

    Han, Yi; Gu, Sheng; Oota, Hiroki; Osier, Michael V.; Pakstis, Andrew J.; Speed, William C.; Kidd, Judith R.; Kidd, Kenneth K.

    2007-01-01

    The alcohol dehydrogenase (ADH) family of enzymes catalyzes the reversible oxidation of alcohol to acetaldehyde. Seven ADH genes exist in a segment of ∼370 kb on 4q21. Products of the three class I ADH genes that share 95% sequence identity are believed to play the major role in the first step of ethanol metabolism. Because the common belief that selection has operated at the ADH1B*47His allele in East Asian populations lacks direct biological or statistical evidence, we used genomic data to test the hypothesis. Data consisted of 54 single-nucleotide polymorphisms (SNPs) across the ADH clusters in a global sampling of 42 populations. Both the Fst statistic and the long-range haplotype (LRH) test provided positive evidence of selection in several East Asian populations. The ADH1B Arg47His functional polymorphism has the highest Fst of the 54 SNPs in the ADH cluster, and it is significantly above the mean Fst of 382 presumably neutral sites tested on the same 42 population samples. The LRH test that uses cores including that site and extending on both sides also gives significant evidence of positive selection in some East Asian populations for a specific haplotype carrying the ADH1B*47His allele. Interestingly, this haplotype is present at a high frequency in only some East Asian populations, whereas the specific allele also exists in other East Asian populations and in the Near East and Europe but does not show evidence of selection with use of the LRH test. Although the ADH1B*47His allele conveys a well-confirmed protection against alcoholism, that modern phenotypic manifestation does not easily translate into a positive selective force, and the nature of that selective force, in the past and/or currently, remains speculative. PMID:17273965

  11. Associations of ADH and ALDH2 gene variation with self report alcohol reactions, consumption and dependence: an integrated analysis

    PubMed Central

    Macgregor, Stuart; Lind, Penelope A.; Bucholz, Kathleen K.; Hansell, Narelle K.; Madden, Pamela A.F.; Richter, Melinda M.; Montgomery, Grant W.; Martin, Nicholas G.; Heath, Andrew C.; Whitfield, John B.

    2009-01-01

    Alcohol dependence (AD) is a complex disorder with environmental and genetic origins. The role of two genetic variants in ALDH2 and ADH1B in AD risk has been extensively investigated. This study tested for associations between nine polymorphisms in ALDH2 and 41 in the seven ADH genes, and alcohol-related flushing, alcohol use and dependence symptom scores in 4597 Australian twins. The vast majority (4296) had consumed alcohol in the previous year, with 547 meeting DSM-IIIR criteria for AD. There were study-wide significant associations (P < 2.3 × 10−4) between ADH1B-Arg48His (rs1229984) and flushing and consumption, but only nominally significant associations (P < 0.01) with dependence. Individuals carrying the rs1229984 G-allele (48Arg) reported a lower prevalence of flushing after alcohol (P = 8.2 × 10−7), consumed alcohol on more occasions (P = 2.7 × 10−6), had a higher maximum number of alcoholic drinks in a single day (P = 2.7 × 10−6) and a higher overall alcohol consumption (P = 8.9 × 10−8) in the previous year than those with the less common A-allele (48His). After controlling for rs1229984, an independent association was observed between rs1042026 (ADH1B) and alcohol intake (P = 4.7 × 10−5) and suggestive associations (P < 0.001) between alcohol consumption phenotypes and rs1693482 (ADH1C), rs1230165 (ADH5) and rs3762894 (ADH4). ALDH2 variation was not associated with flushing or alcohol consumption, but was weakly associated with AD measures. These results bridge the gap between DNA sequence variation and alcohol-related behavior, confirming that the ADH1B-Arg48His polymorphism affects both alcohol-related flushing in Europeans and alcohol intake. The absence of study-wide significant effects on AD results from the low P-value required when testing multiple single nucleotide polymorphisms and phenotypes. PMID:18996923

  12. Genetic variants in or near ADH1B and ADH1C affect susceptibility to alcohol dependence in a British and Irish population.

    PubMed

    Way, Michael; McQuillin, Andrew; Saini, Jit; Ruparelia, Kush; Lydall, Gregory J; Guerrini, Irene; Ball, David; Smith, Iain; Quadri, Giorgia; Thomson, Allan D; Kasiakogia-Worlley, Katherine; Cherian, Raquin; Gunwardena, Priyanthi; Rao, Harish; Kottalgi, Girija; Patel, Shamir; Hillman, Audrey; Douglas, Ewen; Qureshi, Sherhzad Y; Reynolds, Gerry; Jauhar, Sameer; O'Kane, Aideen; Dedman, Alex; Sharp, Sally; Kandaswamy, Radhika; Dar, Karim; Curtis, David; Morgan, Marsha Y; Gurling, Hugh M D

    2015-05-01

    Certain single nucleotide polymorphisms (SNPs) in genes encoding alcohol dehydrogenase (ADH) enzymes confer a significant protective effect against alcohol dependence syndrome (ADS) in East Asian populations. Recently, attention has focused on the role of these SNPs in determining ADS risk in European populations. To further elucidate these associations, SNPs of interest in ADH1B, ADH1C and the ADH1B/1C intergenic region were genotyped in a British and Irish population (ADS cases n = 1076: controls n = 1027) to assess their relative contribution to ADS risk. A highly significant, protective association was observed between the minor allele of rs1229984 in ADH1B and ADS risk [allelic P = 8.4 × 10(-6) , odds ratio (OR) = 0.26, 95 percent confidence interval, 0.14, 0.49]. Significant associations were also observed between ADS risk and the ADH1B/1C intergenic variant, rs1789891 [allelic P = 7.2 × 10(-5) , OR = 1.4 (1.2, 1.6)] and three non-synonymous SNPs rs698, rs1693482 and rs283413 in ADH1C. However, these associations were not completely independent; thus, while the ADH1B rs1229984 minor allele association was independent of those of the intergenic variant rs1789891 and the three ADH1C variants, the three ADH1C variants were not individually independent. In conclusion, the rare ADH1B rs1229984 mutation provides significant protection against ADS in this British and Irish population; other variants in the ADH gene cluster also alter ADS risk, although the strong linkage disequilibrium between SNPs at this location precluded clear identification of the variant(s) driving the associations.

  13. Improvement of Ethanol Production in Saccharomyces cerevisiae by High-Efficient Disruption of the ADH2 Gene Using a Novel Recombinant TALEN Vector

    PubMed Central

    Ye, Wei; Zhang, Weimin; Liu, Taomei; Tan, Guohui; Li, Haohua; Huang, Zilei

    2016-01-01

    Bioethanol is becoming increasingly important in energy supply and economic development. However, the low yield of bioethanol and the insufficiency of high-efficient genetic manipulation approaches limit its application. In this study, a novel transcription activator-like effector nuclease (TALEN) vector containing the left and right arms of TALEN was electroporated into Saccharomyces cerevisiae strain As2.4 to sequence the alcohol dehydrogenase gene ADH2 and the hygromycin-resistant gene hyg. Western blot analysis using anti-FLAG monoclonal antibody proved the successful expression of TALE proteins in As2.4 strains. qPCR and sequencing demonstrated the accurate knockout of the 17 bp target gene with 80% efficiency. The TALEN vector and ADH2 PCR product were electroporated into ΔADH2 to complement the ADH2 gene (ADH2+ As2.4). LC–MS and GC were employed to detect ethanol yields in the native As2.4, ΔADH2 As2.4, and ADH2+ As2.4 strains. Results showed that ethanol production was improved by 52.4 ± 5.3% through the disruption of ADH2 in As2.4. The bioethanol yield of ADH2+ As2.4 was nearly the same as that of native As2.4. This study is the first to report on the disruption of a target gene in S. cerevisiae by employing Fast TALEN technology to improve bioethanol yield. This work provides a novel approach for the disruption of a target gene in S. cerevisiae with high efficiency and specificity, thereby promoting the improvement of bioethanol production in S. cerevisiae by metabolic engineering. PMID:27462304

  14. Improvement of Ethanol Production in Saccharomyces cerevisiae by High-Efficient Disruption of the ADH2 Gene Using a Novel Recombinant TALEN Vector.

    PubMed

    Ye, Wei; Zhang, Weimin; Liu, Taomei; Tan, Guohui; Li, Haohua; Huang, Zilei

    2016-01-01

    Bioethanol is becoming increasingly important in energy supply and economic development. However, the low yield of bioethanol and the insufficiency of high-efficient genetic manipulation approaches limit its application. In this study, a novel transcription activator-like effector nuclease (TALEN) vector containing the left and right arms of TALEN was electroporated into Saccharomyces cerevisiae strain As2.4 to sequence the alcohol dehydrogenase gene ADH2 and the hygromycin-resistant gene hyg. Western blot analysis using anti-FLAG monoclonal antibody proved the successful expression of TALE proteins in As2.4 strains. qPCR and sequencing demonstrated the accurate knockout of the 17 bp target gene with 80% efficiency. The TALEN vector and ADH2 PCR product were electroporated into ΔADH2 to complement the ADH2 gene (ADH2 (+) As2.4). LC-MS and GC were employed to detect ethanol yields in the native As2.4, ΔADH2 As2.4, and ADH2 (+) As2.4 strains. Results showed that ethanol production was improved by 52.4 ± 5.3% through the disruption of ADH2 in As2.4. The bioethanol yield of ADH2 (+) As2.4 was nearly the same as that of native As2.4. This study is the first to report on the disruption of a target gene in S. cerevisiae by employing Fast TALEN technology to improve bioethanol yield. This work provides a novel approach for the disruption of a target gene in S. cerevisiae with high efficiency and specificity, thereby promoting the improvement of bioethanol production in S. cerevisiae by metabolic engineering. PMID:27462304

  15. Experimentally Increased Codon Bias in the Drosophila Adh Gene Leads to an Increase in Larval, But Not Adult, Alcohol Dehydrogenase Activity

    PubMed Central

    Hense, Winfried; Anderson, Nathan; Hutter, Stephan; Stephan, Wolfgang; Parsch, John; Carlini, David B.

    2010-01-01

    Although most amino acids can be encoded by more than one codon, the synonymous codons are not used with equal frequency. This phenomenon is known as codon bias and appears to be a universal feature of genomes. The translational selection hypothesis posits that the use of optimal codons, which match the most abundant species of isoaccepting tRNAs, results in increased translational efficiency and accuracy. Previous work demonstrated that the experimental reduction of codon bias in the Drosophila alcohol dehydrogenase (Adh) gene led to a significant decrease in ADH protein expression. In this study we performed the converse experiment: we replaced seven suboptimal leucine codons that occur naturally in the Drosophila melanogaster Adh gene with the optimal codon. We then compared the in vivo ADH activities imparted by the wild-type and mutant alleles. The introduction of optimal leucine codons led to an increase in ADH activity in third-instar larvae. In adult flies, however, the introduction of optimal codons led to a decrease in ADH activity. There is no evidence that other selectively constrained features of the Adh gene, or its rate of transcription, were altered by the synonymous replacements. These results are consistent with translational selection for codon bias being stronger in the larval stage and suggest that there may be a selective conflict over optimal codon usage between different developmental stages. PMID:19966063

  16. Experimentally increased codon bias in the Drosophila Adh gene leads to an increase in larval, but not adult, alcohol dehydrogenase activity.

    PubMed

    Hense, Winfried; Anderson, Nathan; Hutter, Stephan; Stephan, Wolfgang; Parsch, John; Carlini, David B

    2010-02-01

    Although most amino acids can be encoded by more than one codon, the synonymous codons are not used with equal frequency. This phenomenon is known as codon bias and appears to be a universal feature of genomes. The translational selection hypothesis posits that the use of optimal codons, which match the most abundant species of isoaccepting tRNAs, results in increased translational efficiency and accuracy. Previous work demonstrated that the experimental reduction of codon bias in the Drosophila alcohol dehydrogenase (Adh) gene led to a significant decrease in ADH protein expression. In this study we performed the converse experiment: we replaced seven suboptimal leucine codons that occur naturally in the Drosophila melanogaster Adh gene with the optimal codon. We then compared the in vivo ADH activities imparted by the wild-type and mutant alleles. The introduction of optimal leucine codons led to an increase in ADH activity in third-instar larvae. In adult flies, however, the introduction of optimal codons led to a decrease in ADH activity. There is no evidence that other selectively constrained features of the Adh gene, or its rate of transcription, were altered by the synonymous replacements. These results are consistent with translational selection for codon bias being stronger in the larval stage and suggest that there may be a selective conflict over optimal codon usage between different developmental stages.

  17. Cellulosic Ethanol Production by Recombinant Cellulolytic Bacteria Harbouring pdc and adh II Genes of Zymomonas mobilis

    PubMed Central

    Piriya, P. Sobana; Vasan, P. Thirumalai; Padma, V. S.; Vidhyadevi, U.; Archana, K.; Vennison, S. John

    2012-01-01

    The ethanol fermenting genes such as pyruvate decarboxylase (pdc) and alcohol dehydrogenase II (adh II) were cloned from Zymomonas mobilis and transformed into three different cellulolytic bacteria, namely Enterobacter cloacae JV, Proteus mirabilis JV and Erwinia chrysanthemi and their cellulosic ethanol production capability was studied. Recombinant E. cloacae JV was found to produce 4.5% and 3.5% (v/v) ethanol, respectively, when CMC and 4% NaOH pretreated bagasse were used as substrates, whereas recombinant P. mirabilis and E. chrysanthemi with the same substrates could only produce 4%, 3.5%, 1%, and 1.5 % of ethanol, respectively. The recombinant E. cloacae strain produced twofold higher percentage of ethanol than the wild type. The recombinant E. cloacae strain could be improved further by increasing its ethanol tolerance capability through media optimization and also by combining multigene cellulase expression for enhancing ethanol production from various types of lignocellulosic biomass so that it can be used for industrial level ethanol production. PMID:22919503

  18. Cellulosic Ethanol Production by Recombinant Cellulolytic Bacteria Harbouring pdc and adh II Genes of Zymomonas mobilis.

    PubMed

    Piriya, P Sobana; Vasan, P Thirumalai; Padma, V S; Vidhyadevi, U; Archana, K; Vennison, S John

    2012-01-01

    The ethanol fermenting genes such as pyruvate decarboxylase (pdc) and alcohol dehydrogenase II (adh II) were cloned from Zymomonas mobilis and transformed into three different cellulolytic bacteria, namely Enterobacter cloacae JV, Proteus mirabilis JV and Erwinia chrysanthemi and their cellulosic ethanol production capability was studied. Recombinant E. cloacae JV was found to produce 4.5% and 3.5% (v/v) ethanol, respectively, when CMC and 4% NaOH pretreated bagasse were used as substrates, whereas recombinant P. mirabilis and E. chrysanthemi with the same substrates could only produce 4%, 3.5%, 1%, and 1.5 % of ethanol, respectively. The recombinant E. cloacae strain produced twofold higher percentage of ethanol than the wild type. The recombinant E. cloacae strain could be improved further by increasing its ethanol tolerance capability through media optimization and also by combining multigene cellulase expression for enhancing ethanol production from various types of lignocellulosic biomass so that it can be used for industrial level ethanol production.

  19. Nucleotide sequence analysis of Adh genes estimates the time of geographic isolation of the Bogota population of Drosophila pseudoobscura.

    PubMed Central

    Schaeffer, S W; Miller, E L

    1991-01-01

    The population of Drosophila pseudoobscura at Bogota, Columbia, is geographically and partially reproductively isolated from populations in the main body of the species in North America. The degree of genetic differentiation and time of divergence between populations at Bogota and Apple Hill, CA, were estimated by comparison of 3388 nucleotides in the alcohol dehydrogenase region (Adh and Adh-Dup genes) of 18 strains. Of the 146 polymorphic nucleotide sites detected, 68 and 31 were unique to the Apple Hill and Bogota samples, respectively, and 53 were shared. On the basis of an observed net divergence per nucleotide site of 0.264% between the two samples, the Bogota and North American populations were estimated to have been separated for at least 155,000 years. This divergence time suggests that D. pseudoobscura extended its range from North America to South America in a period of Pleistocene glaciation, when habitat suitable for the species presumably existed in lowland Central America. PMID:2068088

  20. Maize Adh-1 promoter sequences control anaerobic regulation: addition of upstream promoter elements from constitutive genes is necessary for expression in tobacco

    PubMed Central

    Ellis, J.G.; Llewellyn, D.J.; Dennis, E.S.; Peacock, W.J.

    1987-01-01

    The promoter region of a maize alcohol dehydrogenase gene (Adh-1) was linked to a reporter gene encoding chloramphenicol acetyl transferase (CAT) and transformed stably into tobacco cells using T-DNA vectors. No CAT enzyme activity could be detected in transgenic tobacco plants unless upstream promoter elements from the octopine synthase gene or the cauliflower mosaic virus 35S promoter were supplied in addition to the maize promoter region. CAT enzyme activity and transcription of the chimaeric gene were then readily detected after anaerobic induction. The first 247 bp upstream of the translation initiation codon of the maize Adh-1 gene were sufficient to impose anaerobic regulation on the hybrid gene and S1 nuclease mapping confirmed mRNA initiation is from the normal maize Adh-1 transcription start point. ImagesFig. 1.Fig. 2.Fig. 3.Fig. 4.Fig. 5.Fig. 6. PMID:15981329

  1. The relationship between third-codon position nucleotide content, codon bias, mRNA secondary structure and gene expression in the drosophilid alcohol dehydrogenase genes Adh and Adhr.

    PubMed Central

    Carlini, D B; Chen, Y; Stephan, W

    2001-01-01

    To gain insights into the relationship between codon bias, mRNA secondary structure, third-codon position nucleotide distribution, and gene expression, we predicted secondary structures in two related drosophilid genes, Adh and Adhr, which differ in degree of codon bias and level of gene expression. Individual structural elements (helices) were inferred using the comparative method. For each gene, four types of randomization simulations were performed to maintain/remove codon bias and/or to maintain or alter third-codon position nucleotide composition (N3). In the weakly expressed, weakly biased gene Adhr, the potential for secondary structure formation was found to be much stronger than in the highly expressed, highly biased gene Adh. This is consistent with the observation of approximately equal G and C percentages in Adhr ( approximately 31% across species), whereas in Adh the N3 distribution is shifted toward C (42% across species). Perturbing the N3 distribution to approximately equal amounts of A, G, C, and T increases the potential for secondary structure formation in Adh, but decreases it in Adhr. On the other hand, simulations that reduce codon bias without changing N3 content indicate that codon bias per se has only a weak effect on the formation of secondary structures. These results suggest that, for these two drosophilid genes, secondary structure is a relatively independent, negative regulator of gene expression. Whereas the degree of codon bias is positively correlated with level of gene expression, strong individual secondary structural elements may be selected for to retard mRNA translation and to decrease gene expression. PMID:11606539

  2. Expression pattern, ethanol-metabolizing activities, and cellular localization of alcohol and aldehyde dehydrogenases in human large bowel: association of the functional polymorphisms of ADH and ALDH genes with hemorrhoids and colorectal cancer.

    PubMed

    Chiang, Chien-Ping; Jao, Shu-Wen; Lee, Shiao-Pieng; Chen, Pei-Chi; Chung, Chia-Chi; Lee, Shou-Lun; Nieh, Shin; Yin, Shih-Jiun

    2012-02-01

    Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) are principal enzymes responsible for metabolism of ethanol. Functional polymorphisms of ADH1B, ADH1C, and ALDH2 genes occur among racial populations. The goal of this study was to systematically determine the functional expressions and cellular localization of ADHs and ALDHs in human rectal mucosa, the lesions of adenocarcinoma and hemorrhoid, and the genetic association of allelic variations of ADH and ALDH with large bowel disorders. Twenty-one surgical specimens of rectal adenocarcinoma and the adjacent normal mucosa, including 16 paired tissues of rectal tumor, normal mucosae of rectum and sigmoid colon from the same individuals, and 18 surgical mixed hemorrhoid specimens and leukocyte DNA samples from 103 colorectal cancer patients, 67 hemorrhoid patients, and 545 control subjects recruited in previous study, were investigated. The isozyme/allozyme expression patterns of ADH and ALDH were identified by isoelectric focusing and the activities were assayed spectrophotometrically. The protein contents of ADH/ALDH isozymes were determined by immunoblotting using the corresponding purified class-specific antibodies; the cellular activity and protein localizations were detected by immunohistochemistry and histochemistry, respectively. Genotypes of ADH1B, ADH1C, and ALDH2 were determined by polymerase chain reaction-restriction fragment length polymorphisms. At 33mM ethanol, pH 7.5, the activity of ADH1C*1/1 phenotypes exhibited 87% higher than that of the ADH1C*1/*2 phenotypes in normal rectal mucosa. The activity of ALDH2-active phenotypes of rectal mucosa was 33% greater than ALDH2-inactive phenotypes at 200μM acetaldehyde. The protein contents in normal rectal mucosa were in the following order: ADH1>ALDH2>ADH3≈ALDH1A1, whereas those of ADH2, ADH4, and ALDH3A1 were fairly low. Both activity and content of ADH1 were significantly decreased in rectal tumors, whereas the ALDH activity remained

  3. Site-directed mutations reveal long-range compensatory interactions in the Adh gene of Drosophila melanogaster

    PubMed Central

    Parsch, John; Tanda, Soichi; Stephan, Wolfgang

    1997-01-01

    Long-range interactions between the 5′ and 3′ ends of mRNA molecules have been suggested to play a role in the initiation of translation and the regulation of gene expression. To identify such interactions and to study their molecular evolution, we used phylogenetic analysis to generate a model of mRNA higher-order structure in the Adh transcript of Drosophila melanogaster. This model predicts long-range, tertiary contacts between a region of the protein-encoding sequence just downstream of the start codon and a conserved sequence in the 3′ untranslated region (UTR). To further examine the proposed structure, site-directed mutations were generated in vitro in a cloned D. melanogaster Adh gene, and the mutant constructs were introduced into the Drosophila germ line through P-element mediated transformation. Transformants were spectrophotometrically assayed for alcohol dehydrogenase activity. Our results indicate that transformants containing a silent mutation near the start of the protein-encoding sequence show an ≈15% reduction in alcohol dehydrogenase activity relative to wild-type transformants. This activity can be restored to wild-type levels by a second, compensatory mutation in the 3′ UTR. These observations are consistent with a higher-order structure model that includes long-range interactions between the 5′ and 3′ ends of the Adh mRNA. However, our results do not fit the classical compensatory substitution model because the second mutation by itself (in the 3′ UTR) did not show a measurable reduction in gene expression. PMID:9023359

  4. Ethnic Related Selection for an ADH Class I Variant within East Asia

    PubMed Central

    Li, Hui; Gu, Sheng; Cai, Xiaoyun; Speed, William C.; Pakstis, Andrew J.; Golub, Efim I.; Kidd, Judith R.; Kidd, Kenneth K.

    2008-01-01

    Background The alcohol dehydrogenases (ADH) are widely studied enzymes and the evolution of the mammalian gene cluster encoding these enzymes is also well studied. Previous studies have shown that the ADH1B*47His allele at one of the seven genes in humans is associated with a decrease in the risk of alcoholism and the core molecular region with this allele has been selected for in some East Asian populations. As the frequency of ADH1B*47His is highest in East Asia, and very low in most of the rest of the world, we have undertaken more detailed investigation in this geographic region. Methodology/Principal Findings Here we report new data on 30 SNPs in the ADH7 and Class I ADH region in samples of 24 populations from China and Laos. These populations cover a wide geographic region and diverse ethnicities. Combined with our previously published East Asian data for these SNPs in 8 populations, we have typed populations from all of the 6 major linguistic phyla (Altaic including Korean-Japanese and inland Altaic, Sino-Tibetan, Hmong-Mien, Austro-Asiatic, Daic, and Austronesian). The ADH1B genotyping data are strongly related to ethnicity. Only some eastern ethnic phyla or subphyla (Korean-Japanese, Han Chinese, Hmong-Mien, Daic, and Austronesian) have a high frequency of ADH1B*47His. ADH1B haplotype data clustered the populations into linguistic subphyla, and divided the subphyla into eastern and western parts. In the Hmong-Mien and Altaic populations, the extended haplotype homozygosity (EHH) and relative EHH (REHH) tests for the ADH1B core were consistent with selection for the haplotype with derived SNP alleles. In the other ethnic phyla, the core showed only a weak signal of selection at best. Conclusions/Significance The selection distribution is more significantly correlated with the frequency of the derived ADH1B regulatory region polymorphism than the derived amino-acid altering allele ADH1B*47His. Thus, the real focus of selection may be the regulatory region

  5. Dosage compensation of the Drosophila pseudoobscura Hsp82 gene and the Drosophila melanogaster Adh gene at ectopic sites in D. melanogaster.

    PubMed Central

    Sass, H; Meselson, M

    1991-01-01

    Measurements were made of the amounts of larval RNA transcribed from the autosomal Adh gene of Drosophila melanogaster and the X chromosomal Hsp82 gene of Drosophila pseudoobscura carried on the same P-element transposon inserted at various sites in the D. melanogaster genome. Both genes were fully compensated at sites in euchromatic regions of the X chromosome but neither was compensated at a site in the centric beta-heterochromatin of the X chromosome. No compensation of the D. pseudoobscura Hsp82 gene was found at any of 10 autosomal insertion sites tested. The compensation behavior of the transposed genes was, therefore, not determined by closely linked sequences but instead was determined in each case by their new chromosomal environment. Images PMID:1907376

  6. Association and ancestry analysis of sequence variants in ADH and ALDH using alcohol-related phenotypes in a Native American community sample

    PubMed Central

    Peng, Qian; Gizer, Ian R.; Libiger, Ondrej; Bizon, Chris; Wilhelmsen, Kirk C.; Schork, Nicholas J.; Ehlers, Cindy L.

    2015-01-01

    Higher rates of alcohol use and other drug-dependence have been observed in some Native American populations relative to other ethnic groups in the U.S. Previous studies have shown that alcohol dehydrogenase (ADH) genes and aldehyde dehydrogenase (ALDH) genes may affect the risk of development of alcohol dependence, and that polymorphisms within these genes may differentially affect risk for the disorder depending on the ethnic group evaluated. We evaluated variations in the ADH and ALDH genes in a large study investigating risk factors for substance use in a Native American population. We assessed ancestry admixture and tested for associations between alcohol-related phenotypes in the genomic regions around the ADH1-7 and ALDH2 and ALDH1A1 genes. Seventy-two (72) ADH variants showed significant evidence of association with a severity level of alcohol drinking-related dependence symptoms phenotype. These significant variants spanned across the entire 7 ADH gene cluster regions. Two significant associations, one in ADH and one in ALDH2, were observed with alcohol dependence diagnosis. Seventeen (17) variants showed significant association with the largest number of alcohol drinks ingested during any 24-hour period. Variants in or near ADH7 were significantly negatively associated with alcohol-related phenotypes, suggesting a potential protective effect of this gene. In addition, our results suggested that a higher degree of Native American ancestry is associated with higher frequencies of potential risk variants and lower frequencies of potential protective variants for alcohol dependence phenotypes. PMID:25270064

  7. Association and ancestry analysis of sequence variants in ADH and ALDH using alcohol-related phenotypes in a Native American community sample.

    PubMed

    Peng, Qian; Gizer, Ian R; Libiger, Ondrej; Bizon, Chris; Wilhelmsen, Kirk C; Schork, Nicholas J; Ehlers, Cindy L

    2014-12-01

    Higher rates of alcohol use and other drug-dependence have been observed in some Native American (NA) populations relative to other ethnic groups in the US. Previous studies have shown that alcohol dehydrogenase (ADH) genes and aldehyde dehydrogenase (ALDH) genes may affect the risk of development of alcohol dependence, and that polymorphisms within these genes may differentially affect risk for the disorder depending on the ethnic group evaluated. We evaluated variations in the ADH and ALDH genes in a large study investigating risk factors for substance use in a NA population. We assessed ancestry admixture and tested for associations between alcohol-related phenotypes in the genomic regions around the ADH1-7 and ALDH2 and ALDH1A1 genes. Seventy-two ADH variants showed significant evidence of association with a severity level of alcohol drinking-related dependence symptoms phenotype. These significant variants spanned across the entire 7 ADH gene cluster regions. Two significant associations, one in ADH and one in ALDH2, were observed with alcohol dependence diagnosis. Seventeen variants showed significant association with the largest number of alcohol drinks ingested during any 24-hour period. Variants in or near ADH7 were significantly negatively associated with alcohol-related phenotypes, suggesting a potential protective effect of this gene. In addition, our results suggested that a higher degree of NA ancestry is associated with higher frequencies of potential risk variants and lower frequencies of potential protective variants for alcohol dependence phenotypes.

  8. [Verification and Validation on Single Nucleotide Polymorphism Analysis of Alcohol Metabolism-Related Genes ADH1B and ALDH2, Using Dried-Saliva Samples].

    PubMed

    Murata, Shigenori; Hayashida, Mariko; Ishiguro-Tanaka, Yuko; Imazeki, Hiromi; Hayashi, Emiko; Yokoyama, Akira; Kinoshita, Kenji

    2015-11-01

    We have developed a new method for unprocessed biological specimens as templates directly into the TaqMan assay. Saliva was needed to be put on a water-soluble paper and dried, because foreign substances, such as a filter paper, hinder fluorescence detection through the assay. Genotyping of alcohol metabolism-related genes ADH1B (rs1229984) and ALDH2 (rs671) polymorphisms was, subsequently, performed by TaqMan PCR assay using dried saliva in the present investigation. The optimized technique was tested on 114 samples of alcoholic patients. The PCR-RFLP methods with purified DNA from blood samples were employed for validation of the assay. Upon validation, complete concordance was observed between the two independent results. These results highlight the ability of TaqMan PCR assays using dried saliva on water-soluble paper in genotyping of ADH1B and ALDH2 genes. Our results showed a rapid, simple, reliable, and cost-effective method for SNP genotyping of mutations in ADH1B and ALDH2 genes. This will be very useful for large-scale association studies in various fields. [Original]. PMID:26995869

  9. Polymorphism of ADH and ALDH genes among four ethnic groups in China and effects upon the risk for alcoholism.

    PubMed

    Shen, Y C; Fan, J H; Edenberg, H J; Li, T K; Cui, Y H; Wang, Y F; Tian, C H; Zhou, C F; Zhou, R L; Wang, J; Zhao, Z L; Xia, G Y

    1997-10-01

    The alcohol dehydrogenases (ADHs) and aldehyde dehydrogenases (ALDHs) that metabolize ethanol are polymorphic. Different alleles encode subunits of the enzymes that differ in their rate of metabolizing ethanol. These polymorphisms are distributed differently among populations and have been shown to influence the risk for alcoholism in some Asian populations. We have examined the allele frequencies at the ADH2, ADH3, and ALDH2 loci in four populations from China (Han, Mongolian, Korean, and Elunchun) and in alcoholics within each population. The four populations differ in allele frequencies, with the Elunchun having a much lower frequency of ADH2*2 alleles, and the Mongolian and Elunchun having a much lower frequency of ALDH2*2 alleles. Within each population, alleles at one or more of these three loci are protective against alcoholism, although the populations differ in which loci play significant roles. The protective allele at each locus (ALDH2*2, ADH2*2, and ADH3*1) encodes a subunit that either metabolizes ethanol to acetaldehyde more rapidly or slows the conversion of acetaldehyde to acetate. Taken as a whole, data demonstrate that genetic differences in the enzymes that metabolize alcohol can substantially affect the risk for alcoholism.

  10. Molecular Variation of Adh and P6 Genes in an African Population of Drosophila Melanogaster and Its Relation to Chromosomal Inversions

    PubMed Central

    Benassi, V.; Aulard, S.; Mazeau, S.; Veuille, M.

    1993-01-01

    Four-cutter molecular polymorphism of Adh and P6, and chromosome inversion polymorphism of chromosome II were investigated in 95 isogenic lines of an Ivory Coast population of Drosophila melanogaster, a species assumed to have recently spread throughout the world from a West African origin. The P6 gene showed little linkage disequilibrium with the In(2L)t inversion, although it is located within this inversion. This suggests that the inversion and the P6 locus have extensively exchanged genetic information through either double crossover or gene conversion. Allozymic variation in ADH was in linkage disequilibrium with In(2L)t and In(2R)NS inversions. Evidence suggests either that inversion linkage with the Fast allele is selectively maintained, or that this allele only recently appeared. Molecular polymorphism at the Adh locus in the Ivory Coast is not higher than in North American populations. New haplotypes specific to the African population were found, some of them connect the ``Wa(s)-like'' haplotypes found at high frequencies in the United States to the other slow haplotypes. Their relation with In(2L)t supports the hypothesis that Wa(s) recently recombined away from an In(2L)t chromosome which may be the cause of its divergence from the other haplotypes. PMID:8349110

  11. Efficient production of lycopene in Saccharomyces cerevisiae by expression of synthetic crt genes from a plasmid harboring the ADH2 promoter.

    PubMed

    Bahieldin, Ahmed; Gadalla, Nour O; Al-Garni, Saleh M; Almehdar, Hussein; Noor, Samah; Hassan, Sabah M; Shokry, Ahmed M; Sabir, Jamal S M; Murata, Norio

    2014-03-01

    Lycopene is an effective antioxidant proposed as a possible treatment for some cancers and other degenerative human conditions. This study aims at generation of a yeast strain (Saccharomyces cerevisiae) of efficient productivity of lycopene by overexpressing synthetic genes derived from crtE, crtB and crtI genes of Erwinia uredovora. These synthetic genes were constructed in accordance with the preferred codon usage in S. cerevisiae but with no changes in amino acid sequences of the gene products. S. cerevisiae cells were transformed with these synthetic crt genes, whose expression was regulated by the ADH2 promoter, which is de-repressed upon glucose depletion. The RT-PCR and Western blotting analyses indicated that the synthetic crt genes were efficiently transcribed and translated in crt-transformed S. cerevisiae cells. The highest level of lycopene in one of the transformed lines was 3.3mglycopene/g dry cell weight, which is higher than the previously reported levels of lycopene in other microorganisms transformed with the three genes. These results suggest the excellence of using the synthetic crt genes and the ADH2 promoter in generation of recombinant S. cerevisiae that produces a high level of lycopene. The level of ergosterol was reversely correlated to that of lycopene in crt-transformed S. cerevisiae cells, suggesting that two pathways for lycopene and ergosterol syntheses compete for the use of farnesyl diphosphate.

  12. Neurite outgrowth resistance to rho kinase inhibitors in PC12 Adh cell.

    PubMed

    Yin, Hua; Hou, Xiaolin; Tao, Tingrui; Lv, Xiaoman; Zhang, Luyong; Duan, Weigang

    2015-05-01

    Rho kinase (ROCK) inhibitor is a promising agent for neural injury disorders, which mechanism is associated with neurite outgrowth. However, neurite outgrowth resistance occurred when PC12 Adh cell was treated with ROCK inhibitors for a longer time. PC12 Adh cells were treated with ROCK inhibitor Y27632 or NGF for different durations. Neurite outgrowth resistance occurred when PC12 Adh cell exposed to Y27632 (33 µM) for 3 or more days, but not happen when exposed to nerve growth factor (NGF, 100 ng/mL). The gene expression in the PC12 Adh cells treated with Y27632 (33 µM) or NGF (100 ng/mL) for 2 or 4 days was assayed by gene microarray, and the reliability of the results were confirmed by real-time RT-PCR. Cluster analysis proved that the gene expression profile of PC12 Adh cell treated with Y27632 for 4 days was different from that treated with Y27632 for 2 days and those treated with NGF for 2 and 4 days, respectively. Pathway analysis hinted that the neurite outgrowth resistance could be associated with up-regulation of inflammatory pathways, especially rno04610 (complement and coagulation cascades), and down-regulation of cell cycle pathways, especially rno04110.

  13. The Ccr4 Protein from Saccharomyces Cerevisiae Contains a Leucine-Rich Repeat Region Which Is Required for Its Control of Adh2 Gene Expression

    PubMed Central

    Malvar, T.; Biron, R. W.; Kaback, D. B.; Denis, C. L.

    1992-01-01

    The CCR4 gene from Saccharomyces cerevisiae is required for the transcription of the glucose-repressible alcohol dehydrogenase (ADH2). Mutations in CCR4 also suppress the transcription at the ADH2 and his4-912delta loci caused by defects in the SPT10 (CRE1) and SPT6 (CRE2) genes. The CCR4 gene was mapped to the left arm of chromosome I and cloned by complementation of function using previously isolated segments of chromosome I. DNA sequence analysis of the cloned gene defined CCR4 as a 2511 bp open reading frame that would encode a polypeptide of 837 amino acids. The CCR4 mRNA was found to be 2.8 kb in size and Western analysis identified CCR4 as a 95,000 D protein. Disruption of the CCR4 gene resulted in reduced levels of ADH2 expression under both glucose and ethanol growth conditions and in temperature sensitive growth on nonfermentative medium, phenotypes essentially indistinguishable from previously identified mutations in CCR4. The amino terminus of the CCR4 protein was found to be rich in glutamine residues similar to a number of genes which are required for transcription. More importantly, CCR4 showed similarity to a diverse set of proteins sharing a leucine-rich tandem repeat motif, the presence of which has been implicated in mediating protein-protein interactions. Deletions of several of the five leucine-rich repeats in CCR4 were shown to produce nonfunctional proteins indicating the importance of the repeats to CCR4 activity. This leucine-rich repeat region may mediate the contact CCR4 makes with another factor. PMID:1459446

  14. A comparison of two novel alcohol dehydrogenase enzymes (ADH1 and ADH2) from the extreme halophile Haloferax volcanii.

    PubMed

    Timpson, Leanne M; Liliensiek, Ann-Kathrin; Alsafadi, Diya; Cassidy, Jennifer; Sharkey, Michael A; Liddell, Susan; Allers, Thorsten; Paradisi, Francesca

    2013-01-01

    Haloarchaeal alcohol dehydrogenases are exciting biocatalysts with potential industrial applications. In this study, two alcohol dehydrogenase enzymes from the extremely halophilic archaeon Haloferax volcanii (HvADH1 and HvADH2) were homologously expressed and subsequently purified by immobilized metal-affinity chromatography. The proteins appeared to copurify with endogenous alcohol dehydrogenases, and a double Δadh2 Δadh1 gene deletion strain was constructed to prevent this occurrence. Purified HvADH1 and HvADH2 were compared in terms of stability and enzymatic activity over a range of pH values, salt concentrations, and temperatures. Both enzymes were haloalkaliphilic and thermoactive for the oxidative reaction and catalyzed the reductive reaction at a slightly acidic pH. While the NAD(+)-dependent HvADH1 showed a preference for short-chain alcohols and was inherently unstable, HvADH2 exhibited dual cofactor specificity, accepted a broad range of substrates, and, with respect to HvADH1, was remarkably stable. Furthermore, HvADH2 exhibited tolerance to organic solvents. HvADH2 therefore displays much greater potential as an industrially useful biocatalyst than HvADH1.

  15. Ethanol production by Escherichia coli strains co-expressing Zymomonas PDC and ADH genes

    DOEpatents

    Ingram, Lonnie O.; Conway, Tyrrell; Alterthum, Flavio

    1991-01-01

    A novel operon and plasmids comprising genes which code for the alcohol dehydrogenase and pyruvate decarboxylase activities of Zymomonas mobilis are described. Also disclosed are methods for increasing the growth of microorganisms or eukaryotic cells and methods for reducing the accumulation of undesirable metabolic products in the growth medium of microorganisms or cells.

  16. Inferring the Evolutionary Histories of the Adh and Adh-Dup Loci in Drosophila Melanogaster from Patterns of Polymorphism and Divergence

    PubMed Central

    Kreitman, M.; Hudson, R. R.

    1991-01-01

    The DNA sequences of 11 Drosophila melanogaster lines are compared across three contiguous regions, the Adh and Adh-dup loci and a noncoding 5' flanking region of Adh. Ninety-eight of approximately 4750 sites are segregating in the sample, 36 in the 5' flanking region, 38 in Adh and 24 in Adh-dup. Several methods are presented to test whether the patterns and levels of polymorphism are consistent with neutral molecular evolution. The analysis of within- and between-species polymorphism indicates that the region is evolving in a nonneutral and complex fashion. A graphical analysis of the data provides support for a hypothesized balanced polymorphism at or near position 1490, site of the amino acid replacement difference between Adh(f) and Adh(s). The Adh-dup locus is less polymorphic than Adh and all 24 of its polymorphisms occur at low frequency--suggestive of a recent selective substitution in the Adh-dup region. Adh(s) alleles form two distinct evolutionary lineages that differ one from another at a total of nineteen sites in the Adh and Adh-dup loci. The polymorphisms are in complete linkage disequilibrium. A recombination experiment failed to find evidence for recombination suppression between the two allelic classes. Two hypotheses are presented to account for the widespread distribution of the two divergent lineages in natural populations. Natural selection appears to have played an important role in governing the overall patterns of nucleotide variation across the two-gene region. PMID:1673107

  17. Persistence drives gene clustering in bacterial genomes

    PubMed Central

    Fang, Gang; Rocha, Eduardo PC; Danchin, Antoine

    2008-01-01

    Background Gene clustering plays an important role in the organization of the bacterial chromosome and several mechanisms have been proposed to explain its extent. However, the controversies raised about the validity of each of these mechanisms remind us that the cause of this gene organization remains an open question. Models proposed to explain clustering did not take into account the function of the gene products nor the likely presence or absence of a given gene in a genome. However, genomes harbor two very different categories of genes: those genes present in a majority of organisms – persistent genes – and those present in very few organisms – rare genes. Results We show that two classes of genes are significantly clustered in bacterial genomes: the highly persistent and the rare genes. The clustering of rare genes is readily explained by the selfish operon theory. Yet, genes persistently present in bacterial genomes are also clustered and we try to understand why. We propose a model accounting specifically for such clustering, and show that indispensability in a genome with frequent gene deletion and insertion leads to the transient clustering of these genes. The model describes how clusters are created via the gene flux that continuously introduces new genes while deleting others. We then test if known selective processes, such as co-transcription, physical interaction or functional neighborhood, account for the stabilization of these clusters. Conclusion We show that the strong selective pressure acting on the function of persistent genes, in a permanent state of flux of genes in bacterial genomes, maintaining their size fairly constant, that drives persistent genes clustering. A further selective stabilization process might contribute to maintaining the clustering. PMID:18179692

  18. Biological cluster evaluation for gene function prediction.

    PubMed

    Klie, Sebastian; Nikoloski, Zoran; Selbig, Joachim

    2014-06-01

    Recent advances in high-throughput omics techniques render it possible to decode the function of genes by using the "guilt-by-association" principle on biologically meaningful clusters of gene expression data. However, the existing frameworks for biological evaluation of gene clusters are hindered by two bottleneck issues: (1) the choice for the number of clusters, and (2) the external measures which do not take in consideration the structure of the analyzed data and the ontology of the existing biological knowledge. Here, we address the identified bottlenecks by developing a novel framework that allows not only for biological evaluation of gene expression clusters based on existing structured knowledge, but also for prediction of putative gene functions. The proposed framework facilitates propagation of statistical significance at each of the following steps: (1) estimating the number of clusters, (2) evaluating the clusters in terms of novel external structural measures, (3) selecting an optimal clustering algorithm, and (4) predicting gene functions. The framework also includes a method for evaluation of gene clusters based on the structure of the employed ontology. Moreover, our method for obtaining a probabilistic range for the number of clusters is demonstrated valid on synthetic data and available gene expression profiles from Saccharomyces cerevisiae. Finally, we propose a network-based approach for gene function prediction which relies on the clustering of optimal score and the employed ontology. Our approach effectively predicts gene function on the Saccharomyces cerevisiae data set and is also employed to obtain putative gene functions for an Arabidopsis thaliana data set.

  19. Taraxerone enhances alcohol oxidation via increases of alcohol dehyderogenase (ADH) and acetaldehyde dehydrogenase (ALDH) activities and gene expressions.

    PubMed

    Sung, Chang-Keun; Kim, Seung-Mi; Oh, Chang-Jin; Yang, Sun-A; Han, Byung-Hee; Mo, Eun-Kyoung

    2012-07-01

    The present study, taraxerone (d-friedoolean-14-en-3-one) was isolated from Sedum sarmentosum with purity 96.383%, and its enhancing effects on alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH) activities were determined: EC(50) values were 512.42 ± 3.12 and 500.16 ± 3.23 μM for ADH and ALDH, respectively. In order to obtain more information on taraxerone related with the alcohol metabolism, 40% ethanol (5 mL/kg body weight) with 0.5-1mM of taraxerone were administered to mice. The plasma alcohol and acetaldehyde concentrations of taraxerone-treated groups were significantly lowered than those of the control group (p<0.01): approximately 20-67% and 7-57% lowered for plasma alcohol and acetaldehyde, respectively. Compare to the control group, the ADH and ALDH expressions in the liver tissues were abruptly increased in the taraxerone-treated groups after ethanol exposure. In addition, taraxerone prevented catalase, superoxide dismutase, and reduced glutathione concentrations from the decrease induced by ethanol administration with the concentration dependent manner.

  20. ADH single nucleotide polymorphism associations with alcohol metabolism in vivo

    PubMed Central

    Birley, Andrew J.; James, Michael R.; Dickson, Peter A.; Montgomery, Grant W.; Heath, Andrew C.; Martin, Nicholas G.; Whitfield, John B.

    2009-01-01

    We have previously found that variation in alcohol metabolism in Europeans is linked to the chromosome 4q region containing the ADH gene family. We have now typed 103 single nucleotide polymorphisms (SNPs) across this region to test for allelic associations with variation in blood and breath alcohol concentrations after an alcohol challenge. In vivo alcohol metabolism was modelled with three parameters that identified the absorption and rise of alcohol concentration following ingestion, and the rate of elimination. Alleles of ADH7 SNPs were associated with the early stages of alcohol metabolism, with additional effects in the ADH1A, ADH1B and ADH4 regions. Rate of elimination was associated with SNPs in the intragenic region between ADH7 and ADH1C, and across ADH1C and ADH1B. SNPs affecting alcohol metabolism did not correspond to those reported to affect alcohol dependence or alcohol-related disease. The combined SNP associations with early- and late-stage metabolism only account for approximately 20% of the total genetic variance linked to the ADH region, and most of the variance for in vivo alcohol metabolism linked to this region is yet to be explained. PMID:19193628

  1. Molecular analysis of the Adh region of the genome of Drosophila melanogaster.

    PubMed

    Chia, W; Karp, R; McGill, S; Ashburner, M

    1985-12-20

    A small region of the genome of Drosophila melanogaster has been cloned in a series of overlapping phage. A length of 165 X 10(3) base-pairs of contiguous DNA that spans polytene chromosome region 35A4 to 35B1 and includes the structural gene for alcohol dehydrogenase (Adh) as well as at least two other genes, outspread (osp) and no-ocelli (noc), has been characterized by mapping chromosome aberrations to the DNA. The relationship between osp and Adh is surprising: of nine osp alleles associated with chromosome breakpoints, five map distal (i.e. 5') to Adh and four map proximal (i.e. 3') to this gene. None affects the expression of Adh. As defined by these and other breakpoints, the osp gene spans at least 52 X 10(3) base-pairs and overlaps the Adh gene. The noc gene, as defined by the mapping of nearly 30 breakpoints, is at least 50 X 10(3) base-pairs in size. Alleles of noc and noc- deletions show either of two kinds of interaction with the recessive lethality of l(2)br29ScoR+1, a lethal that maps immediately distal to noc. One class of noc allele is viable when heterozygous with ScoR+1, while the other class is lethal or semi-lethal. Both classes, however, are homozygous or hemizygous viable. The locations of these two classes of noc allele on the DNA fall into two clusters, with those that are viable with ScoR+1 located proximal to those that are not. The physical boundary between these classes lies at a site just distal to that of the breakpoint of the inversion associated with ScoR+1 itself.

  2. Cofactor Specificity of the Bifunctional Alcohol and Aldehyde Dehydrogenase (AdhE) in Wild-Type and Mutant Clostridium thermocellum and Thermoanaerobacterium saccharolyticum

    PubMed Central

    Zheng, Tianyong; Olson, Daniel G.; Tian, Liang; Bomble, Yannick J.; Himmel, Michael E.; Lo, Jonathan; Hon, Shuen; Shaw, A. Joe; van Dijken, Johannes P.

    2015-01-01

    ABSTRACT Clostridium thermocellum and Thermoanaerobacterium saccharolyticum are thermophilic bacteria that have been engineered to produce ethanol from the cellulose and hemicellulose fractions of biomass, respectively. Although engineered strains of T. saccharolyticum produce ethanol with a yield of 90% of the theoretical maximum, engineered strains of C. thermocellum produce ethanol at lower yields (∼50% of the theoretical maximum). In the course of engineering these strains, a number of mutations have been discovered in their adhE genes, which encode both alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) enzymes. To understand the effects of these mutations, the adhE genes from six strains of C. thermocellum and T. saccharolyticum were cloned and expressed in Escherichia coli, the enzymes produced were purified by affinity chromatography, and enzyme activity was measured. In wild-type strains of both organisms, NADH was the preferred cofactor for both ALDH and ADH activities. In high-ethanol-producing (ethanologen) strains of T. saccharolyticum, both ALDH and ADH activities showed increased NADPH-linked activity. Interestingly, the AdhE protein of the ethanologenic strain of C. thermocellum has acquired high NADPH-linked ADH activity while maintaining NADH-linked ALDH and ADH activities at wild-type levels. When single amino acid mutations in AdhE that caused increased NADPH-linked ADH activity were introduced into C. thermocellum and T. saccharolyticum, ethanol production increased in both organisms. Structural analysis of the wild-type and mutant AdhE proteins was performed to provide explanations for the cofactor specificity change on a molecular level. IMPORTANCE This work describes the characterization of the AdhE enzyme from different strains of C. thermocellum and T. saccharolyticum. C. thermocellum and T. saccharolyticum are thermophilic anaerobes that have been engineered to make high yields of ethanol and can solubilize components of

  3. Deletion of a conserved regulatory element in the Drosophila Adh gene leads to increased alcohol dehydrogenase activity but also delays development.

    PubMed Central

    Parsch, J; Russell, J A; Beerman, I; Hartl, D L; Stephan, W

    2000-01-01

    In vivo levels of enzymatic activity may be increased through either structural or regulatory changes. Here we use Drosophila melanogaster alcohol dehydrogenase (ADH) in an experimental test for selective differences between these two mechanisms. The well-known ADH-Slow (S)/Fast (F) amino acid replacement leads to a twofold increase in activity by increasing the catalytic efficiency of the enzyme. Disruption of a highly conserved, negative regulatory element in the Adh 3' UTR also leads to a twofold increase in activity, although this is achieved by increasing in vivo Adh mRNA and protein concentrations. These two changes appear to be under different types of selection, with positive selection favoring the amino acid replacement and purifying selection maintaining the 3' UTR sequence. Using transgenic experiments we show that deletion of the conserved 3' UTR element increases adult and larval Adh expression in both the ADH-F and ADH-S genetic backgrounds. However, the 3' UTR deletion also leads to a significant increase in developmental time in both backgrounds. ADH allozyme type has no detectable effect on development. These results demonstrate a negative fitness effect associated with Adh overexpression. This provides a mechanism whereby natural selection can discriminate between alternative pathways of increasing enzymatic activity. PMID:10978287

  4. Characterization of the Structure and Evolution of the Adh Region of Drosophila Hydei

    PubMed Central

    Menotti-Raymond, M.; Starmer, W. T.; Sullivan, D. T.

    1991-01-01

    Drosophila of the repleta group have a duplication of the gene which encodes alcohol dehydrogenase (ADH). We report the nucleotide sequence of an 8.4-kb region of genomic DNA of Drosophila hydei which includes the entire Adh region. Analysis of this sequence reveals similarity in organization to the Adh region of Drosophila mojavensis and Drosophila mulleri of the mulleri subgroup, with three genes ordered 5' to 3', Adh-ψ, Adh-2, Adh-1. Deletion of a nucleotide in the second codon of each pseudogene suggests that the first Adh duplication occurred before the divergence of the hydei and mulleri subgroups. However, Adh-1 and Adh-2 of D. hydei are significantly more alike than Adh-1 and Adh-2 of D. mojavensis. Models to account for the difference in similarity between the coding genes were tested by orthologous and paralogous comparisons of the extent of sequence divergence. A model which proposes that independent duplication events generated Adh-1 and Adh-2 in the two lineages is supported by these data. The D. hydei pseudogene is transcribed and the transcript is processed in a complex manner. An intron of greater than 6.2 kb exists between the first ``coding'' exon and an upstream exon which is approximately 250 nucleotides in length. PMID:2004708

  5. ADH (Antidiuretic Hormone) Test

    MedlinePlus

    ... Also known as: Vasopressin; AVP Formal name: Antidiuretic Hormone; Arginine Vasopressin Related tests: Osmolality , BUN , Creatinine , Sodium , ... should know? How is it used? The antidiuretic hormone (ADH) test is used to help detect, diagnose, ...

  6. Clustering of High Throughput Gene Expression Data

    PubMed Central

    Pirim, Harun; Ekşioğlu, Burak; Perkins, Andy; Yüceer, Çetin

    2012-01-01

    High throughput biological data need to be processed, analyzed, and interpreted to address problems in life sciences. Bioinformatics, computational biology, and systems biology deal with biological problems using computational methods. Clustering is one of the methods used to gain insight into biological processes, particularly at the genomics level. Clearly, clustering can be used in many areas of biological data analysis. However, this paper presents a review of the current clustering algorithms designed especially for analyzing gene expression data. It is also intended to introduce one of the main problems in bioinformatics - clustering gene expression data - to the operations research community. PMID:23144527

  7. Clustering of gene ontology terms in genomes.

    PubMed

    Tiirikka, Timo; Siermala, Markku; Vihinen, Mauno

    2014-10-25

    Although protein coding genes occupy only a small fraction of genomes in higher species, they are not randomly distributed within or between chromosomes. Clustering of genes with related function(s) and/or characteristics has been evident at several different levels. To study how common the clustering of functionally related genes is and what kind of functions the end products of these genes are involved, we collected gene ontology (GO) terms for complete genomes and developed a method to detect previously undefined gene clustering. Exhaustive analysis was performed for seven widely studied species ranging from human to Escherichia coli. To overcome problems related to varying gene lengths and densities, a novel method was developed and a fixed number of genes were analyzed irrespective of the genome span covered. Statistically very significant GO term clustering was apparent in all the investigated genomes. The analysis window, which ranged from 5 to 50 consecutive genes, revealed extensive GO term clusters for genes with widely varying functions. Here, the most interesting and significant results are discussed and the complete dataset for each analyzed species is available at the GOme database at http://bioinf.uta.fi/GOme. The results indicated that clusters of genes with related functions are very common, not only in bacteria, in which operons are frequent, but also in all the studied species irrespective of how complex they are. There are some differences between species but in all of them GO term clusters are common and of widely differing sizes. The presented method can be applied to analyze any genome or part of a genome for which descriptive features are available, and thus is not restricted to ontology terms. This method can also be applied to investigate gene and protein expression patterns. The results pave a way for further studies of mechanisms that shape genome structure and evolutionary forces related to them. PMID:24995610

  8. Genetic polymorphisms of ADH1B, ADH1C and ALDH2 in Turkish alcoholics: lack of association with alcoholism and alcoholic cirrhosis.

    PubMed

    Vatansever, Sezgin; Tekin, Fatih; Salman, Esin; Altintoprak, Ender; Coskunol, Hakan; Akarca, Ulus Salih

    2015-05-17

    No data exists regarding the alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) gene polymorphisms in Turkish alcoholic cirrhotics. We studied the polymorphisms of ADH1B, ADH1C and ALDH2 genes in alcoholic cirrhotics and compared the results with non-cirrhotic alcoholics and healthy volunteers. Overall, 237 subjects were included for the study: 156 alcoholic patients (78 cirrhotics, 78 non-cirrhotic alcoholics) and 81 healthy volunteers. Three different single-nucleotide-polymorphism genotyping methods were used. ADH1C genotyping was performed using a polymerase chain reaction-restriction fragment length polymorphism method. The identified ADH1C genotypes were named according to the presence or absence of the enzyme restriction sites. ADH1B (Arg47Hys) genotyping was performed using the allele specific primer extension method, and ALDH2 (Glu487Lys) genotyping was performed by a multiplex polymerase chain reaction using two allele-specific primer pairs. For ADH1B, the frequency of allele *1 in the cirrhotics, non-cirrhotic alcoholics and healthy volunteers was 97.4%, 94.9% and 99.4%, respectively. For ADH1C, the frequency of allele *1 in the cirrhotics, non-cirrhotic alcoholics and healthy volunteers was 47%, 36.3% and 45%, respectively. There was no statistical difference between the groups for ADH1B and ADH1C (p>0.05). All alcoholic and non-alcoholic subjects (100%) had the allele *1 for ALDH2. The obtained results for ADH1B, ADH1C, and ALDH gene polymorphisms in the present study are similar to the results of Caucasian studies. ADH1B and ADH1C genetic variations are not related to the development of alcoholism or susceptibility to alcoholic cirrhosis. ALDH2 gene has no genetic variation in the Turkish population.

  9. Chicken rRNA Gene Cluster Structure

    PubMed Central

    Dyomin, Alexander G.; Koshel, Elena I.; Kiselev, Artem M.; Saifitdinova, Alsu F.; Galkina, Svetlana A.; Fukagawa, Tatsuo; Kostareva, Anna A.

    2016-01-01

    Ribosomal RNA (rRNA) genes, whose activity results in nucleolus formation, constitute an extremely important part of genome. Despite the extensive exploration into avian genomes, no complete description of avian rRNA gene primary structure has been offered so far. We publish a complete chicken rRNA gene cluster sequence here, including 5’ETS (1836 bp), 18S rRNA gene (1823 bp), ITS1 (2530 bp), 5.8S rRNA gene (157 bp), ITS2 (733 bp), 28S rRNA gene (4441 bp) and 3’ETS (343 bp). The rRNA gene cluster sequence of 11863 bp was assembled from raw reads and deposited to GenBank under KT445934 accession number. The assembly was validated through in situ fluorescent hybridization analysis on chicken metaphase chromosomes using computed and synthesized specific probes, as well as through the reference assembly against de novo assembled rRNA gene cluster sequence using sequenced fragments of BAC-clone containing chicken NOR (nucleolus organizer region). The results have confirmed the chicken rRNA gene cluster validity. PMID:27299357

  10. Conserved enhancer and silencer elements responsible for differential Adh transcription in Drosophila cell lines.

    PubMed Central

    Ayer, S; Benyajati, C

    1990-01-01

    The distal promoter of Adh is differentially expressed in Drosophila tissue culture cell lines. After transfection with an exogenous Adh gene, there was a specific increase in distal alcohol dehydrogenase (ADH) transcripts in ADH-expressing (ADH+) cells above the levels observed in transfected ADH-nonexpressing (ADH-) cells. We used deletion mutations and a comparative transient-expression assay to identify the cis-acting elements responsible for enhanced Adh distal transcription in ADH+ cells. DNA sequences controlling high levels of distal transcription were localized to a 15-base-pair (bp) region nearly 500 bp upstream of the distal RNA start site. In addition, a 61-bp negative cis-acting element was found upstream from and adjacent to the enhancer. When this silencer element was deleted, distal transcription increased only in the ADH+ cell line. These distant upstream elements must interact with the promoter elements, the Adf-1-binding site and the TATA box, as they only influenced transcription when at least one of these two positive distal promoter elements was present. Internal deletions targeted to the Adf-1-binding site or the TATA box reduced transcription in both cell types but did not affect the transcription initiation site. Distal transcription in transfected ADH- cells appears to be controlled primarily through these promoter elements and does not involve the upstream regulatory elements. Evolutionary conservation in distantly related Drosophila species suggests the importance of these upstream elements in correct developmental and tissue-specific expression of ADH. Images PMID:1694013

  11. Protein variation in Adh and Adh-related in Drosophila pseudoobscura. Linkage disequilibrium between single nucleotide polymorphisms and protein alleles.

    PubMed Central

    Schaeffer, S W; Walthour, C S; Toleno, D M; Olek, A T; Miller, E L

    2001-01-01

    A 3.5-kb segment of the alcohol dehydrogenase (Adh) region that includes the Adh and Adh-related genes was sequenced in 139 Drosophila pseudoobscura strains collected from 13 populations. The Adh gene encodes four protein alleles and rejects a neutral model of protein evolution with the McDonald-Kreitman test, although the number of segregating synonymous sites is too high to conclude that adaptive selection has operated. The Adh-related gene encodes 18 protein haplotypes and fails to reject an equilibrium neutral model. The populations fail to show significant geographic differentiation of the Adh-related haplotypes. Eight of 404 single nucleotide polymorphisms (SNPs) in the Adh region were in significant linkage disequilibrium with three ADHR protein alleles. Coalescent simulations with and without recombination were used to derive the expected levels of significant linkage disequilibrium between SNPs and 18 protein haplotypes. Maximum levels of linkage disequilibrium are expected for protein alleles at moderate frequencies. In coalescent models without recombination, linkage disequilibrium decays between SNPs and high frequency haplotypes because common alleles mutate to haplotypes that are rare or that reach moderate frequency. The implication of this study is that linkage disequilibrium mapping has the highest probability of success with disease-causing alleles at frequencies of 10%. PMID:11606543

  12. Transcriptomic Identification of ADH1B as a Novel Candidate Gene for Obesity and Insulin Resistance in Human Adipose Tissue in Mexican Americans from the Veterans Administration Genetic Epidemiology Study (VAGES)

    PubMed Central

    Winnier, Deidre A.; Fourcaudot, Marcel; Norton, Luke; Abdul-Ghani, Muhammad A.; Hu, Shirley L.; Farook, Vidya S.; Coletta, Dawn K.; Kumar, Satish; Puppala, Sobha; Chittoor, Geetha; Dyer, Thomas D.; Arya, Rector; Carless, Melanie; Lehman, Donna M.; Curran, Joanne E.; Cromack, Douglas T.; Tripathy, Devjit; Blangero, John; Duggirala, Ravindranath; Göring, Harald H. H.; DeFronzo, Ralph A.; Jenkinson, Christopher P.

    2015-01-01

    Type 2 diabetes (T2D) is a complex metabolic disease that is more prevalent in ethnic groups such as Mexican Americans, and is strongly associated with the risk factors obesity and insulin resistance. The goal of this study was to perform whole genome gene expression profiling in adipose tissue to detect common patterns of gene regulation associated with obesity and insulin resistance. We used phenotypic and genotypic data from 308 Mexican American participants from the Veterans Administration Genetic Epidemiology Study (VAGES). Basal fasting RNA was extracted from adipose tissue biopsies from a subset of 75 unrelated individuals, and gene expression data generated on the Illumina BeadArray platform. The number of gene probes with significant expression above baseline was approximately 31,000. We performed multiple regression analysis of all probes with 15 metabolic traits. Adipose tissue had 3,012 genes significantly associated with the traits of interest (false discovery rate, FDR ≤ 0.05). The significance of gene expression changes was used to select 52 genes with significant (FDR ≤ 10-4) gene expression changes across multiple traits. Gene sets/Pathways analysis identified one gene, alcohol dehydrogenase 1B (ADH1B) that was significantly enriched (P < 10-60) as a prime candidate for involvement in multiple relevant metabolic pathways. Illumina BeadChip derived ADH1B expression data was consistent with quantitative real time PCR data. We observed significant inverse correlations with waist circumference (2.8 x 10-9), BMI (5.4 x 10-6), and fasting plasma insulin (P < 0.001). These findings are consistent with a central role for ADH1B in obesity and insulin resistance and provide evidence for a novel genetic regulatory mechanism for human metabolic diseases related to these traits. PMID:25830378

  13. ADH IB expression, but not ADH III, is decreased in human lung cancer.

    PubMed

    Mutka, Sarah C; Green, Lucia H; Verderber, Evie L; Richards, Jane P; Looker, Doug L; Chlipala, Elizabeth A; Rosenthal, Gary J

    2012-01-01

    Endogenous S-nitrosothiols, including S-nitrosoglutathione (GSNO), mediate nitric oxide (NO)-based signaling, inflammatory responses, and smooth muscle function. Reduced GSNO levels have been implicated in several respiratory diseases, and inhibition of GSNO reductase, (GSNOR) the primary enzyme that metabolizes GSNO, represents a novel approach to treating inflammatory lung diseases. Recently, an association between decreased GSNOR expression and human lung cancer risk was proposed in part based on immunohistochemical staining using a polyclonal GSNOR antibody. GSNOR is an isozyme of the alcohol dehydrogenase (ADH) family, and we demonstrate that the antibody used in those studies cross reacts substantially with other ADH proteins and may not be an appropriate reagent. We evaluated human lung cancer tissue arrays using monoclonal antibodies highly specific for human GSNOR with minimal cross reactivity to other ADH proteins. We verified the presence of GSNOR in ≥85% of specimens examined, and extensive analysis of these samples demonstrated no difference in GSNOR protein expression between cancerous and normal lung tissues. Additionally, GSNOR and other ADH mRNA levels were evaluated quantitatively in lung cancer cDNA arrays by qPCR. Consistent with our immunohistochemical findings, GSNOR mRNA levels were not changed in lung cancer tissues, however the expression levels of other ADH genes were decreased. ADH IB mRNA levels were reduced (>10-fold) in 65% of the lung cancer cDNA specimens. We conclude that the previously reported results showed an incorrect association of GSNOR and human lung cancer risk, and a decrease in ADH IB, rather than GSNOR, correlates with human lung cancer.

  14. ADH IB Expression, but Not ADH III, Is Decreased in Human Lung Cancer

    PubMed Central

    Mutka, Sarah C.; Green, Lucia H.; Verderber, Evie L.; Richards, Jane P.; Looker, Doug L.; Chlipala, Elizabeth A.; Rosenthal, Gary J.

    2012-01-01

    Endogenous S-nitrosothiols, including S-nitrosoglutathione (GSNO), mediate nitric oxide (NO)-based signaling, inflammatory responses, and smooth muscle function. Reduced GSNO levels have been implicated in several respiratory diseases, and inhibition of GSNO reductase, (GSNOR) the primary enzyme that metabolizes GSNO, represents a novel approach to treating inflammatory lung diseases. Recently, an association between decreased GSNOR expression and human lung cancer risk was proposed in part based on immunohistochemical staining using a polyclonal GSNOR antibody. GSNOR is an isozyme of the alcohol dehydrogenase (ADH) family, and we demonstrate that the antibody used in those studies cross reacts substantially with other ADH proteins and may not be an appropriate reagent. We evaluated human lung cancer tissue arrays using monoclonal antibodies highly specific for human GSNOR with minimal cross reactivity to other ADH proteins. We verified the presence of GSNOR in ≥85% of specimens examined, and extensive analysis of these samples demonstrated no difference in GSNOR protein expression between cancerous and normal lung tissues. Additionally, GSNOR and other ADH mRNA levels were evaluated quantitatively in lung cancer cDNA arrays by qPCR. Consistent with our immunohistochemical findings, GSNOR mRNA levels were not changed in lung cancer tissues, however the expression levels of other ADH genes were decreased. ADH IB mRNA levels were reduced (>10-fold) in 65% of the lung cancer cDNA specimens. We conclude that the previously reported results showed an incorrect association of GSNOR and human lung cancer risk, and a decrease in ADH IB, rather than GSNOR, correlates with human lung cancer. PMID:23285246

  15. ADH IB expression, but not ADH III, is decreased in human lung cancer.

    PubMed

    Mutka, Sarah C; Green, Lucia H; Verderber, Evie L; Richards, Jane P; Looker, Doug L; Chlipala, Elizabeth A; Rosenthal, Gary J

    2012-01-01

    Endogenous S-nitrosothiols, including S-nitrosoglutathione (GSNO), mediate nitric oxide (NO)-based signaling, inflammatory responses, and smooth muscle function. Reduced GSNO levels have been implicated in several respiratory diseases, and inhibition of GSNO reductase, (GSNOR) the primary enzyme that metabolizes GSNO, represents a novel approach to treating inflammatory lung diseases. Recently, an association between decreased GSNOR expression and human lung cancer risk was proposed in part based on immunohistochemical staining using a polyclonal GSNOR antibody. GSNOR is an isozyme of the alcohol dehydrogenase (ADH) family, and we demonstrate that the antibody used in those studies cross reacts substantially with other ADH proteins and may not be an appropriate reagent. We evaluated human lung cancer tissue arrays using monoclonal antibodies highly specific for human GSNOR with minimal cross reactivity to other ADH proteins. We verified the presence of GSNOR in ≥85% of specimens examined, and extensive analysis of these samples demonstrated no difference in GSNOR protein expression between cancerous and normal lung tissues. Additionally, GSNOR and other ADH mRNA levels were evaluated quantitatively in lung cancer cDNA arrays by qPCR. Consistent with our immunohistochemical findings, GSNOR mRNA levels were not changed in lung cancer tissues, however the expression levels of other ADH genes were decreased. ADH IB mRNA levels were reduced (>10-fold) in 65% of the lung cancer cDNA specimens. We conclude that the previously reported results showed an incorrect association of GSNOR and human lung cancer risk, and a decrease in ADH IB, rather than GSNOR, correlates with human lung cancer. PMID:23285246

  16. Clustering Genes of Common Evolutionary History

    PubMed Central

    Gori, Kevin; Suchan, Tomasz; Alvarez, Nadir; Goldman, Nick; Dessimoz, Christophe

    2016-01-01

    Phylogenetic inference can potentially result in a more accurate tree using data from multiple loci. However, if the loci are incongruent—due to events such as incomplete lineage sorting or horizontal gene transfer—it can be misleading to infer a single tree. To address this, many previous contributions have taken a mechanistic approach, by modeling specific processes. Alternatively, one can cluster loci without assuming how these incongruencies might arise. Such “process-agnostic” approaches typically infer a tree for each locus and cluster these. There are, however, many possible combinations of tree distance and clustering methods; their comparative performance in the context of tree incongruence is largely unknown. Furthermore, because standard model selection criteria such as AIC cannot be applied to problems with a variable number of topologies, the issue of inferring the optimal number of clusters is poorly understood. Here, we perform a large-scale simulation study of phylogenetic distances and clustering methods to infer loci of common evolutionary history. We observe that the best-performing combinations are distances accounting for branch lengths followed by spectral clustering or Ward’s method. We also introduce two statistical tests to infer the optimal number of clusters and show that they strongly outperform the silhouette criterion, a general-purpose heuristic. We illustrate the usefulness of the approach by 1) identifying errors in a previous phylogenetic analysis of yeast species and 2) identifying topological incongruence among newly sequenced loci of the globeflower fly genus Chiastocheta. We release treeCl, a new program to cluster genes of common evolutionary history (http://git.io/treeCl). PMID:26893301

  17. Clustering Genes of Common Evolutionary History.

    PubMed

    Gori, Kevin; Suchan, Tomasz; Alvarez, Nadir; Goldman, Nick; Dessimoz, Christophe

    2016-06-01

    Phylogenetic inference can potentially result in a more accurate tree using data from multiple loci. However, if the loci are incongruent-due to events such as incomplete lineage sorting or horizontal gene transfer-it can be misleading to infer a single tree. To address this, many previous contributions have taken a mechanistic approach, by modeling specific processes. Alternatively, one can cluster loci without assuming how these incongruencies might arise. Such "process-agnostic" approaches typically infer a tree for each locus and cluster these. There are, however, many possible combinations of tree distance and clustering methods; their comparative performance in the context of tree incongruence is largely unknown. Furthermore, because standard model selection criteria such as AIC cannot be applied to problems with a variable number of topologies, the issue of inferring the optimal number of clusters is poorly understood. Here, we perform a large-scale simulation study of phylogenetic distances and clustering methods to infer loci of common evolutionary history. We observe that the best-performing combinations are distances accounting for branch lengths followed by spectral clustering or Ward's method. We also introduce two statistical tests to infer the optimal number of clusters and show that they strongly outperform the silhouette criterion, a general-purpose heuristic. We illustrate the usefulness of the approach by 1) identifying errors in a previous phylogenetic analysis of yeast species and 2) identifying topological incongruence among newly sequenced loci of the globeflower fly genus Chiastocheta We release treeCl, a new program to cluster genes of common evolutionary history (http://git.io/treeCl). PMID:26893301

  18. Bifunctional aldehyde/alcohol dehydrogenase (ADHE) in chlorophyte algal mitochondria.

    PubMed

    Atteia, Ariane; van Lis, Robert; Mendoza-Hernández, Guillermo; Henze, Katrin; Martin, William; Riveros-Rosas, Hector; González-Halphen, Diego

    2003-09-01

    Protein profiles of mitochondria isolated from the heterotrophic chlorophyte Polytomella sp. grown on ethanol at pH 6.0 and pH 3.7 were analyzed by Blue Native and denaturing polyacrylamide gel electrophoresis. Steady-state levels of oxidative phosphorylation complexes were influenced by external pH. Levels of an abundant, soluble, mitochondrial protein of 85 kDa and its corresponding mRNA increased at pH 6.0 relative to pH 3.7. N-terminal and internal sequencing of the 85 kDa mitochondrial protein together with the corresponding cDNA identified it as a bifunctional aldehyde/alcohol dehydrogenase (ADHE) with strong similarity to homologues from eubacteria and amitochondriate protists. A mitochondrial targeting sequence of 27 amino acids precedes the N-terminus of the mature mitochondrial protein. A gene encoding an ADHE homologue was also identified in the genome of Chlamydomonas reinhardtii, a photosynthetic relative of Polytomella. ADHE reveals a complex picture of sequence similarity among homologues. The lack of ADHE from archaebacteria indicates a eubacterial origin for the eukaryotic enzyme. Among eukaryotes, ADHE has hitherto been characteristic of anaerobes since it is essential to cytosolic energy metabolism of amitochondriate protists such as Giardia intestinalis and Entamoeba histolytica. Its abundance and expression pattern suggest an important role for ADHE in mitochondrial metabolism of Polytomella under the conditions studied. The current data are compatible with the view that Polytomella ADHE could be involved either in ethanol production or assimilation, or both, depending upon environmental conditions. Presence of ADHE in an oxygen-respiring algal mitochondrion and co-expression at ambient oxygen levels with respiratory chain components is unexpected with respect to the view that eukaryotes acquired ADHE genes specifically as an adaptation to an anaerobic lifestyle.

  19. Bifunctional aldehyde/alcohol dehydrogenase (ADHE) in chlorophyte algal mitochondria.

    PubMed

    Atteia, Ariane; van Lis, Robert; Mendoza-Hernández, Guillermo; Henze, Katrin; Martin, William; Riveros-Rosas, Hector; González-Halphen, Diego

    2003-09-01

    Protein profiles of mitochondria isolated from the heterotrophic chlorophyte Polytomella sp. grown on ethanol at pH 6.0 and pH 3.7 were analyzed by Blue Native and denaturing polyacrylamide gel electrophoresis. Steady-state levels of oxidative phosphorylation complexes were influenced by external pH. Levels of an abundant, soluble, mitochondrial protein of 85 kDa and its corresponding mRNA increased at pH 6.0 relative to pH 3.7. N-terminal and internal sequencing of the 85 kDa mitochondrial protein together with the corresponding cDNA identified it as a bifunctional aldehyde/alcohol dehydrogenase (ADHE) with strong similarity to homologues from eubacteria and amitochondriate protists. A mitochondrial targeting sequence of 27 amino acids precedes the N-terminus of the mature mitochondrial protein. A gene encoding an ADHE homologue was also identified in the genome of Chlamydomonas reinhardtii, a photosynthetic relative of Polytomella. ADHE reveals a complex picture of sequence similarity among homologues. The lack of ADHE from archaebacteria indicates a eubacterial origin for the eukaryotic enzyme. Among eukaryotes, ADHE has hitherto been characteristic of anaerobes since it is essential to cytosolic energy metabolism of amitochondriate protists such as Giardia intestinalis and Entamoeba histolytica. Its abundance and expression pattern suggest an important role for ADHE in mitochondrial metabolism of Polytomella under the conditions studied. The current data are compatible with the view that Polytomella ADHE could be involved either in ethanol production or assimilation, or both, depending upon environmental conditions. Presence of ADHE in an oxygen-respiring algal mitochondrion and co-expression at ambient oxygen levels with respiratory chain components is unexpected with respect to the view that eukaryotes acquired ADHE genes specifically as an adaptation to an anaerobic lifestyle. PMID:14756315

  20. Combined clustering models for the analysis of gene expression

    SciTech Connect

    Angelova, M. Ellman, J.

    2010-02-15

    Clustering has become one of the fundamental tools for analyzing gene expression and producing gene classifications. Clustering models enable finding patterns of similarity in order to understand gene function, gene regulation, cellular processes and sub-types of cells. The clustering results however have to be combined with sequence data or knowledge about gene functionality in order to make biologically meaningful conclusions. In this work, we explore a new model that integrates gene expression with sequence or text information.

  1. Delineation of Cis-Acting Sequences Required for Expression of Drosophila Mojavensis Adh-1

    PubMed Central

    Bayer, C. A.; Curtiss, S. W.; Weaver, J. A.; Sullivan, D. T.

    1992-01-01

    The control of expression of the Adh-1 gene of Drosophila mojavensis has been analyzed by transforming ADH null Drosophila melanogaster hosts with P element constructs which contain D. mojavensis Adh-1 having deletions of different extent in the 5' and 3' ends. Adh-1 expression in the D. melanogaster hosts is qualitatively similar to expression in D. mojavensis, although expression is quantitatively lower in transformants. Deletions of the 5' end indicate that information required for normal temporal and tissue expression in larvae is contained within 70 bp of the transcription start site. However, deletion constructs to -70 are deficient in ovarian nurse cell expression, whereas the additional upstream sequences present in constructs containing deletions to -257 do support expression in the ovary. Comparison of the nucleotide sequence in the -257 to -70 region of Adh-1 of four species: D. mojavensis and Drosophila arizona, which express Adh-1 in the ovary, and Drosophila mulleri and Drosophila navojoa, which do not, has led to the identification of regions of sequence similarity that correlate with ovary expression. One of these bears a striking similarity to a conserved sequence located upstream of the three heat shock genes that have constitutive ovarian expression and may be an ovarian control element. We have identified an aberrant aspect of Adh-1 expression. In transformants which carry an Adh-1 gene without a functional upstream Adh-2 gene Adh-1 expression continues into the adult stage instead of ceasing at the onset of metamorphosis. In transformants with a functional Adh-2 gene, Adh-1 expression ceases in the third larval instar stage and aberrant expression in the adult stage does not occur. PMID:1317314

  2. Genetic Variation in the Expression of ADH in DROSOPHILA MELANOGASTER

    PubMed Central

    Maroni, G.; Laurie-Ahlberg, C. C.; Adams, D. A.; Wilton, A. N.

    1982-01-01

    Several chromosomes derived from natural populations have been identified that affect the expression of alcohol dehydrogenase (ADH). Second chromosomes, which also carry the structural gene Adh, show a great deal of polymorphism of genetic elements that determine how much enzyme protein accumulates. The level of enzyme was measured in third instar larvae, 6-to-8-day-old males and in larval fat bodies and alimentary canals. In general, activities in the different organs and stages are highly correlated with one another. One line was found, however, in which the ADH level in the fat body is more than twice the level one would expect on the basis of the activity in alimentary canal. We have also found evidence of third-chromosome elements that affect the level of ADH. PMID:6816669

  3. Functional Gene Group Summarization by Clustering MEDLINE Abstract Sentences

    PubMed Central

    Yang, Jianji; Cohen, Aaron M.; Hersh, William R.

    2006-01-01

    Tools to automatically summarize functional gene group information from the biomedical literature will help genomics researchers both better interpret gene expression data and understand biological pathways. In this study, we built a system that takes in a set of genes and MEDLINE records and outputs clusters of genes along with summaries of each cluster by sentence extraction from MEDLINE abstracts. Our preliminary use-case evaluation shows that this approach can identify gene clusters similar to manually generated groupings. PMID:17238770

  4. Sequence variation of alcohol dehydrogenase (Adh) paralogs in cactophilic Drosophila.

    PubMed Central

    Matzkin, Luciano M; Eanes, Walter F

    2003-01-01

    This study focuses on the population genetics of alcohol dehydrogenase (Adh) in cactophilic Drosophila. Drosophila mojavensis and D. arizonae utilize cactus hosts, and each host contains a characteristic mixture of alcohol compounds. In these Drosophila species there are two functional Adh loci, an adult form (Adh-2) and a larval and ovarian form (Adh-1). Overall, the greater level of variation segregating in D. arizonae than in D. mojavensis suggests a larger population size for D. arizonae. There are markedly different patterns of variation between the paralogs across both species. A 16-bp intron haplotype segregates in both species at Adh-2, apparently the product of an ancient gene conversion event between the paralogs, which suggests that there is selection for the maintenance of the intron structure possibly for the maintenance of pre-mRNA structure. We observe a pattern of variation consistent with adaptive protein evolution in the D. mojavensis lineage at Adh-1, suggesting that the cactus host shift that occurred in the divergence of D. mojavensis from D. arizonae had an effect on the evolution of the larval expressed paralog. Contrary to previous work we estimate a recent time for both the divergence of D. mojavensis and D. arizonae (2.4 +/- 0.7 MY) and the age of the gene duplication (3.95 +/- 0.45 MY). PMID:12586706

  5. Cloning and sequencing of the gene cluster encoding two subunits of membrane-bound alcohol dehydrogenase from Acetobacter polyoxogenes.

    PubMed

    Tamaki, T; Fukaya, M; Takemura, H; Tayama, K; Okumura, H; Kawamura, Y; Nishiyama, M; Horinouchi, S; Beppu, T

    1991-02-16

    The membrane-bound alcohol dehydrogenase (ADH) from Acetobacter polyoxogenes NBI1028 is composed of a 72 kDa subunit and a 44 kDa cytochrome c subunit. The amino acid sequences of the two regions of the 72 kDa subunit were determined to prepare oligonucleotides for the purpose of amplification of a DNA fragment corresponding to the intermediate region by the polymerase chain reaction. A 0.5 kb DNA fragment thus amplified was used as the probe to clone a 7.0 kb PstI fragment coding for the whole 72 kDa subunit. Nucleotide sequencing and immunoblot analysis revealed that the cloned fragment contained the full structural genes for the 72 kDa and the 44 kDa subunits and they were clustered with the same transcription polarity. The predicted amino acid sequence of the gene for the 72 kDa subunit showed homology with that of the 72 kDa subunit from ADH of A. aceti and those of methanol dehydrogenase from methylotrophic bacteria. The 72 and 44 kDa subunits contained one and three typical haem binding sequences, respectively.

  6. Detection of putative virulence genes of Lactococcus garvieae.

    PubMed

    Ture, Mustafa; Altinok, Ilhan

    2016-04-12

    Lactococcus garvieae is the causative agent of lactococcosis and has been isolated from a wide variety of animals. In the present study, 34 strains of L. garvieae isolated from fish from different sources and locations were tested for the presence or absence of the following putative virulence genes: a capsule gene cluster (CGC), hemolysins 1, 2, and 3 (hly1, -2, -3), NADH oxidase, superoxide dismutase (sod), phosphoglucomutase (pgm), adhesin Pav (adhPav), adhesin PsaA (adhPsaA), enolase (eno), LPxTG-containing surface proteins 1, 2, 3, and 4 (LPxTG-1, LPxTG-2, LPxTG-3, LPxTG-4; where LPxTG means Leu-Pro-any-Thr-Gly), adhesin clusters 1 and 2 (adhCI, adhCII), and adhesin (adh). To determine the presence of the CGC, we developed a multiplex PCR. All strains of L. garvieae had the hly1, -2, -3, NADH oxidase, pgm, adhPav, LPxTG-2, LPxTG-3, sod, eno, adhPsaA, adhCII, and adhCII genes, while only the Lg2 strain contained the CGC. The virulent Lg2 strain contained all 17 virulent genes. All Turkish, Spanish, Italian, and French strains did not contain the CGC. The multiplex PCR assay was useful for the detection of the CGC genes. In conclusion, the CGC is not the only virulent factor in L. garvieae because strains that lack the CGC are virulent to rainbow trout. Single genes also might not be responsible for the virulence of L. garvieae.

  7. Detection of putative virulence genes of Lactococcus garvieae.

    PubMed

    Ture, Mustafa; Altinok, Ilhan

    2016-04-12

    Lactococcus garvieae is the causative agent of lactococcosis and has been isolated from a wide variety of animals. In the present study, 34 strains of L. garvieae isolated from fish from different sources and locations were tested for the presence or absence of the following putative virulence genes: a capsule gene cluster (CGC), hemolysins 1, 2, and 3 (hly1, -2, -3), NADH oxidase, superoxide dismutase (sod), phosphoglucomutase (pgm), adhesin Pav (adhPav), adhesin PsaA (adhPsaA), enolase (eno), LPxTG-containing surface proteins 1, 2, 3, and 4 (LPxTG-1, LPxTG-2, LPxTG-3, LPxTG-4; where LPxTG means Leu-Pro-any-Thr-Gly), adhesin clusters 1 and 2 (adhCI, adhCII), and adhesin (adh). To determine the presence of the CGC, we developed a multiplex PCR. All strains of L. garvieae had the hly1, -2, -3, NADH oxidase, pgm, adhPav, LPxTG-2, LPxTG-3, sod, eno, adhPsaA, adhCII, and adhCII genes, while only the Lg2 strain contained the CGC. The virulent Lg2 strain contained all 17 virulent genes. All Turkish, Spanish, Italian, and French strains did not contain the CGC. The multiplex PCR assay was useful for the detection of the CGC genes. In conclusion, the CGC is not the only virulent factor in L. garvieae because strains that lack the CGC are virulent to rainbow trout. Single genes also might not be responsible for the virulence of L. garvieae. PMID:27068503

  8. Microarray gene cluster identification and annotation through cluster ensemble and EM-based informative textual summarization.

    PubMed

    Hu, Xiaohua; Park, E K; Zhang, Xiaodan

    2009-09-01

    Generating high-quality gene clusters and identifying the underlying biological mechanism of the gene clusters are the important goals of clustering gene expression analysis. To get high-quality cluster results, most of the current approaches rely on choosing the best cluster algorithm, in which the design biases and assumptions meet the underlying distribution of the dataset. There are two issues for this approach: 1) usually, the underlying data distribution of the gene expression datasets is unknown and 2) there are so many clustering algorithms available and it is very challenging to choose the proper one. To provide a textual summary of the gene clusters, the most explored approach is the extractive approach that essentially builds upon techniques borrowed from the information retrieval, in which the objective is to provide terms to be used for query expansion, and not to act as a stand-alone summary for the entire document sets. Another drawback is that the clustering quality and cluster interpretation are treated as two isolated research problems and are studied separately. In this paper, we design and develop a unified system Gene Expression Miner to address these challenging issues in a principled and general manner by integrating cluster ensemble, text clustering, and multidocument summarization and provide an environment for comprehensive gene expression data analysis. We present a novel cluster ensemble approach to generate high-quality gene cluster. In our text summarization module, given a gene cluster, our expectation-maximization based algorithm can automatically identify subtopics and extract most probable terms for each topic. Then, the extracted top k topical terms from each subtopic are combined to form the biological explanation of each gene cluster. Experimental results demonstrate that our system can obtain high-quality clusters and provide informative key terms for the gene clusters.

  9. Evolution of Hox gene clusters in deuterostomes.

    PubMed

    Pascual-Anaya, Juan; D'Aniello, Salvatore; Kuratani, Shigeru; Garcia-Fernàndez, Jordi

    2013-01-01

    Hox genes, with their similar roles in animals as evolutionarily distant as humans and flies, have fascinated biologists since their discovery nearly 30 years ago. During the last two decades, reports on Hox genes from a still growing number of eumetazoan species have increased our knowledge on the Hox gene contents of a wide range of animal groups. In this review, we summarize the current Hox inventory among deuterostomes, not only in the well-known teleosts and tetrapods, but also in the earlier vertebrate and invertebrate groups. We draw an updated picture of the ancestral repertoires of the different lineages, a sort of "genome Hox bar-code" for most clades. This scenario allows us to infer differential gene or cluster losses and gains that occurred during deuterostome evolution, which might be causally linked to the morphological changes that led to these widely diverse animal taxa. Finally, we focus on the challenging family of posterior Hox genes, which probably originated through independent tandem duplication events at the origin of each of the ambulacrarian, cephalochordate and vertebrate/urochordate lineages.

  10. Evolution of Hox gene clusters in deuterostomes

    PubMed Central

    2013-01-01

    Hox genes, with their similar roles in animals as evolutionarily distant as humans and flies, have fascinated biologists since their discovery nearly 30 years ago. During the last two decades, reports on Hox genes from a still growing number of eumetazoan species have increased our knowledge on the Hox gene contents of a wide range of animal groups. In this review, we summarize the current Hox inventory among deuterostomes, not only in the well-known teleosts and tetrapods, but also in the earlier vertebrate and invertebrate groups. We draw an updated picture of the ancestral repertoires of the different lineages, a sort of “genome Hox bar-code” for most clades. This scenario allows us to infer differential gene or cluster losses and gains that occurred during deuterostome evolution, which might be causally linked to the morphological changes that led to these widely diverse animal taxa. Finally, we focus on the challenging family of posterior Hox genes, which probably originated through independent tandem duplication events at the origin of each of the ambulacrarian, cephalochordate and vertebrate/urochordate lineages. PMID:23819519

  11. The rise of operon-like gene clusters in plants.

    PubMed

    Boycheva, Svetlana; Daviet, Laurent; Wolfender, Jean-Luc; Fitzpatrick, Teresa B

    2014-07-01

    Gene clusters are common features of prokaryotic genomes also present in eukaryotes. Most clustered genes known are involved in the biosynthesis of secondary metabolites. Although horizontal gene transfer is a primary source of prokaryotic gene cluster (operon) formation and has been reported to occur in eukaryotes, the predominant source of cluster formation in eukaryotes appears to arise de novo or through gene duplication followed by neo- and sub-functionalization or translocation. Here we aim to provide an overview of the current knowledge and open questions related to plant gene cluster functioning, assembly, and regulation. We also present potential research approaches and point out the benefits of a better understanding of gene clusters in plants for both fundamental and applied plant science.

  12. Streptomyces coelicolor as an expression host for heterologous gene clusters.

    PubMed

    Gomez-Escribano, Juan Pablo; Bibb, Mervyn J

    2012-01-01

    The expression of a gene or a set of genes from one organism in a different species is known as "heterologous expression." In actinomycetes, prolific producers of natural products, heterologous gene expression has been used to confirm the clustering of secondary metabolite biosynthetic genes, to analyze natural product biosynthesis, to produce variants of natural products by genetic engineering, and to discover new compounds by screening genomic libraries. Recent advances in DNA sequencing have enabled the rapid and affordable sequencing of actinomycete genomes and revealed a large number of secondary metabolite gene clusters with no known products. Heterologous expression of these cryptic gene clusters combined with comparative metabolic profiling provides an important means to identify potentially novel compounds. In this chapter, the methods and strategies used to heterologously express actinomycete gene clusters, including the techniques used for cloning secondary metabolite gene clusters, the Streptomyces hosts used for their expression, and the techniques employed to analyze their products by metabolic profiling, are described.

  13. Genetic characteristics of vancomycin resistance gene cluster in Enterococcus spp.

    PubMed

    Chunhui, Chen; Xiaogang, Xu

    2015-05-01

    Vancomycin resistant enterococci has become an important nosocomial pathogen since it is discovered in late 1980s. The products, encoded by vancomycin resistant gene cluster in enterococci, catalyze the synthesis of peptidoglycan precursors with low affinity with glycopeptide antibiotics including vancomycin and teicoplanin and lead to resistance. These vancomycin resistant gene clusters are classified into nine types according to their gene sequences and organization, or D-Ala:D-Lac (VanA, VanB, VanD and VanM) and D-Ala:D-Ser (VanC, VanE, VanG, VanL and VanN) ligase gene clusters based on the differences of their encoded ligases. Moreover, these gene clusters are characterized by their different resistance levels and infection models. In this review, we summarize the classification, gene organization and infection model of vancomycin resistant gene cluster in Enterococcus spp.

  14. The Alcohol Dehydrogenase Gene Family in Melon (Cucumis melo L.): Bioinformatic Analysis and Expression Patterns

    PubMed Central

    Jin, Yazhong; Zhang, Chong; Liu, Wei; Tang, Yufan; Qi, Hongyan; Chen, Hao; Cao, Songxiao

    2016-01-01

    Alcohol dehydrogenases (ADH), encoded by multigene family in plants, play a critical role in plant growth, development, adaptation, fruit ripening and aroma production. Thirteen ADH genes were identified in melon genome, including 12 ADHs and one formaldehyde dehydrogenease (FDH), designated CmADH1-12 and CmFDH1, in which CmADH1 and CmADH2 have been isolated in Cantaloupe. ADH genes shared a lower identity with each other at the protein level and had different intron-exon structure at nucleotide level. No typical signal peptides were found in all CmADHs, and CmADH proteins might locate in the cytoplasm. The phylogenetic tree revealed that 13 ADH genes were divided into three groups respectively, namely long-, medium-, and short-chain ADH subfamily, and CmADH1,3-11, which belongs to the medium-chain ADH subfamily, fell into six medium-chain ADH subgroups. CmADH12 may belong to the long-chain ADH subfamily, while CmFDH1 may be a Class III ADH and serve as an ancestral ADH in melon. Expression profiling revealed that CmADH1, CmADH2, CmADH10 and CmFDH1 were moderately or strongly expressed in different vegetative tissues and fruit at medium and late developmental stages, while CmADH8 and CmADH12 were highly expressed in fruit after 20 days. CmADH3 showed preferential expression in young tissues. CmADH4 only had slight expression in root. Promoter analysis revealed several motifs of CmADH genes involved in the gene expression modulated by various hormones, and the response pattern of CmADH genes to ABA, IAA and ethylene were different. These CmADHs were divided into ethylene-sensitive and –insensitive groups, and the functions of CmADHs were discussed. PMID:27242871

  15. The Alcohol Dehydrogenase Gene Family in Melon (Cucumis melo L.): Bioinformatic Analysis and Expression Patterns.

    PubMed

    Jin, Yazhong; Zhang, Chong; Liu, Wei; Tang, Yufan; Qi, Hongyan; Chen, Hao; Cao, Songxiao

    2016-01-01

    Alcohol dehydrogenases (ADH), encoded by multigene family in plants, play a critical role in plant growth, development, adaptation, fruit ripening and aroma production. Thirteen ADH genes were identified in melon genome, including 12 ADHs and one formaldehyde dehydrogenease (FDH), designated CmADH1-12 and CmFDH1, in which CmADH1 and CmADH2 have been isolated in Cantaloupe. ADH genes shared a lower identity with each other at the protein level and had different intron-exon structure at nucleotide level. No typical signal peptides were found in all CmADHs, and CmADH proteins might locate in the cytoplasm. The phylogenetic tree revealed that 13 ADH genes were divided into three groups respectively, namely long-, medium-, and short-chain ADH subfamily, and CmADH1,3-11, which belongs to the medium-chain ADH subfamily, fell into six medium-chain ADH subgroups. CmADH12 may belong to the long-chain ADH subfamily, while CmFDH1 may be a Class III ADH and serve as an ancestral ADH in melon. Expression profiling revealed that CmADH1, CmADH2, CmADH10 and CmFDH1 were moderately or strongly expressed in different vegetative tissues and fruit at medium and late developmental stages, while CmADH8 and CmADH12 were highly expressed in fruit after 20 days. CmADH3 showed preferential expression in young tissues. CmADH4 only had slight expression in root. Promoter analysis revealed several motifs of CmADH genes involved in the gene expression modulated by various hormones, and the response pattern of CmADH genes to ABA, IAA and ethylene were different. These CmADHs were divided into ethylene-sensitive and -insensitive groups, and the functions of CmADHs were discussed. PMID:27242871

  16. Ethanol-Induced Alcohol Dehydrogenase E (AdhE) Potentiates Pneumolysin in Streptococcus pneumoniae

    PubMed Central

    Luong, Truc Thanh; Kim, Eun-Hye; Bak, Jong Phil; Nguyen, Cuong Thach; Choi, Sangdun; Briles, David E.; Pyo, Suhkneung

    2014-01-01

    Alcohol impairs the host immune system, rendering the host more vulnerable to infection. Therefore, alcoholics are at increased risk of acquiring serious bacterial infections caused by Streptococcus pneumoniae, including pneumonia. Nevertheless, how alcohol affects pneumococcal virulence remains unclear. Here, we showed that the S. pneumoniae type 2 D39 strain is ethanol tolerant and that alcohol upregulates alcohol dehydrogenase E (AdhE) and potentiates pneumolysin (Ply). Hemolytic activity, colonization, and virulence of S. pneumoniae, as well as host cell myeloperoxidase activity, proinflammatory cytokine secretion, and inflammation, were significantly attenuated in adhE mutant bacteria (ΔadhE strain) compared to D39 wild-type bacteria. Therefore, AdhE might act as a pneumococcal virulence factor. Moreover, in the presence of ethanol, S. pneumoniae AdhE produced acetaldehyde and NADH, which subsequently led Rex (redox-sensing transcriptional repressor) to dissociate from the adhE promoter. An increase in AdhE level under the ethanol condition conferred an increase in Ply and H2O2 levels. Consistently, S. pneumoniae D39 caused higher cytotoxicity to RAW 264.7 cells than the ΔadhE strain under the ethanol stress condition, and ethanol-fed mice (alcoholic mice) were more susceptible to infection with the D39 wild-type bacteria than with the ΔadhE strain. Taken together, these data indicate that AdhE increases Ply under the ethanol stress condition, thus potentiating pneumococcal virulence. PMID:25312953

  17. Ethanol-induced alcohol dehydrogenase E (AdhE) potentiates pneumolysin in Streptococcus pneumoniae.

    PubMed

    Luong, Truc Thanh; Kim, Eun-Hye; Bak, Jong Phil; Nguyen, Cuong Thach; Choi, Sangdun; Briles, David E; Pyo, Suhkneung; Rhee, Dong-Kwon

    2015-01-01

    Alcohol impairs the host immune system, rendering the host more vulnerable to infection. Therefore, alcoholics are at increased risk of acquiring serious bacterial infections caused by Streptococcus pneumoniae, including pneumonia. Nevertheless, how alcohol affects pneumococcal virulence remains unclear. Here, we showed that the S. pneumoniae type 2 D39 strain is ethanol tolerant and that alcohol upregulates alcohol dehydrogenase E (AdhE) and potentiates pneumolysin (Ply). Hemolytic activity, colonization, and virulence of S. pneumoniae, as well as host cell myeloperoxidase activity, proinflammatory cytokine secretion, and inflammation, were significantly attenuated in adhE mutant bacteria (ΔadhE strain) compared to D39 wild-type bacteria. Therefore, AdhE might act as a pneumococcal virulence factor. Moreover, in the presence of ethanol, S. pneumoniae AdhE produced acetaldehyde and NADH, which subsequently led Rex (redox-sensing transcriptional repressor) to dissociate from the adhE promoter. An increase in AdhE level under the ethanol condition conferred an increase in Ply and H2O2 levels. Consistently, S. pneumoniae D39 caused higher cytotoxicity to RAW 264.7 cells than the ΔadhE strain under the ethanol stress condition, and ethanol-fed mice (alcoholic mice) were more susceptible to infection with the D39 wild-type bacteria than with the ΔadhE strain. Taken together, these data indicate that AdhE increases Ply under the ethanol stress condition, thus potentiating pneumococcal virulence.

  18. A Nomadic Subtelomeric Disease Resistance Gene Cluster in Common Bean

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The B4 resistance (R)-gene cluster, located in subtelomeric region of chromosome 4, is one of the largest clusters known in common bean (Phaseolus vulgaris, Pv). We sequenced 650 kb spanning this locus and annotated 97 genes, 26 of which correspond to Coiled-coil-Nucleotide-Binding-Site-Leucine-Rich...

  19. Ethanol formation in adh0 mutants reveals the existence of a novel acetaldehyde-reducing activity in Saccharomyces cerevisiae.

    PubMed Central

    Drewke, C; Thielen, J; Ciriacy, M

    1990-01-01

    A strain of Saccharomyces cerevisiae has been constructed which is deficient in the four alcohol dehydrogenase (ADH) isozymes known at present. This strain (adh0), being irreversibly mutated in the genes ADH1, ADH3, and ADH4 and carrying a point mutation in the gene ADH2 coding for the glucose-repressible isozyme ADHII, still produces up to one third of the theoretical maximum yield of ethanol in a homofermentative conversion of glucose to ethanol. Analysis of the glucose metabolism of adh0 cells shows that the lack of all known ADH isozymes results in the formation of glycerol as a major fermentation product, accompanied by a significant production of acetaldehyde and acetate. Treatment of glucose-growing adh0 cells with the respiratory-chain inhibitor antimycin A leads to an immediate cessation of ethanol production, demonstrating that ethanol production in adh0 cells is dependent on mitochondrial electron transport. Reduction of acetaldehyde to ethanol in isolated mitochondria could also be demonstrated. This reduction is apparently linked to the oxidation of acetaldehyde to acetate. Preliminary data suggest that this novel type of ethanol formation in S. cerevisiae is associated with the inner mitochondrial membrane. Images PMID:2193925

  20. Allelic variants of ADH, ALDH and the five factor model of personality in alcohol dependence syndrome

    PubMed Central

    Salujha, S. K.; Chaudhury, S.; Menon, P. K.; Srivastava, K.; Gupta, A.

    2014-01-01

    Background: The etiology of alcohol dependence is a complex interplay of biopsychosocial factors. The genes for alcohol-metabolizing enzymes: Alcohol dehydrogenase (ADH2 and ADH3) and aldehyde dehydrogenase (ALDH2) exhibit functional polymorphisms. Vulnerability of alcohol dependence may also be in part due to heritable personality traits. Aim: To determine whether any association exists between polymorphisms of ADH2, ADH3 and ALDH2 and alcohol dependence syndrome in a group of Asian Indians. In addition, the personality of these patients was assessed to identify traits predisposing to alcoholism. Materials and Methods: In this study, 100 consecutive males with alcohol dependence syndrome attending the psychiatric outpatient department of a tertiary care service hospital and an equal number of matched healthy controls were included with their consent. Blood samples of all the study cases and controls were collected and genotyped for the ADH2, ADH3 and ALDH2 loci. Personality was evaluated using the neuroticism, extraversion, openness (NEO) personality inventory and sensation seeking scale. Results: Allele frequencies of ADH2*2 (0.50), ADH3*1 (0.67) and ALSH2*2 (0.09) were significantly low in the alcohol dependent subjects. Personality traits of NEO personality inventory and sensation seeking were significantly higher when compared to controls. Conclusions: The functional polymorphisms of genes coding for alcohol metabolizing enzymes and personality traits of NEO and sensation seeking may affect the propensity to develop dependence. PMID:25535445

  1. Selection variability for Arg48His in alcohol dehydrogenase ADH1B among Asian populations.

    PubMed

    Evsyukov, Alexey; Ivanov, Denis

    2013-08-01

    The variant His at codon 48 of the alcohol dehydrogenase gene (ADH1B) results in more efficient ethanol metabolism than with the "typical" codon 48Arg. In this study we introduced selection properties of Arg48His genotypes of ADH1B and estimated fitness in four ethnic-geographical clusters in Asia. Population genetics models were employed that derive observed gene frequencies from fitness relationships among genotypes, to infer the selection pattern of polymorphisms in an indirect manner. The data were analyzed using the model of "complete stationary distribution" by Wright that takes into account random genetic drift, pressure of migrations, mutations, and selection as influential factors of gene frequency. We found that the different population groups showed some variation in the types of selection for Arg48His. Han Chinese from eastern and southeastern China and the Japanese and Korean populations showed stabilizing selection, while the groups from Central Asian and Indochina showed divergent selection. However, all the groups demonstrated a strong positive selection for Arg48His. PMID:25019189

  2. Estimating the number of clusters via system evolution for cluster analysis of gene expression data.

    PubMed

    Wang, Kaijun; Zheng, Jie; Zhang, Junying; Dong, Jiyang

    2009-09-01

    The estimation of the number of clusters (NC) is one of crucial problems in the cluster analysis of gene expression data. Most approaches available give their answers without the intuitive information about separable degrees between clusters. However, this information is useful for understanding cluster structures. To provide this information, we propose system evolution (SE) method to estimate NC based on partitioning around medoids (PAM) clustering algorithm. SE analyzes cluster structures of a dataset from the viewpoint of a pseudothermodynamics system. The system will go to its stable equilibrium state, at which the optimal NC is found, via its partitioning process and merging process. The experimental results on simulated and real gene expression data demonstrate that the SE works well on the data with well-separated clusters and the one with slightly overlapping clusters. PMID:19527960

  3. The ADH7 Promoter of Saccharomyces cerevisiae is Vanillin-Inducible and Enables mRNA Translation Under Severe Vanillin Stress.

    PubMed

    Nguyen, Trinh T M; Iwaki, Aya; Izawa, Shingo

    2015-01-01

    Vanillin is one of the major phenolic aldehyde compounds derived from lignocellulosic biomass and acts as a potent fermentation inhibitor to repress the growth and fermentative ability of yeast. Vanillin can be reduced to its less toxic form, vanillyl alcohol, by the yeast NADPH-dependent medium chain alcohol dehydrogenases, Adh6 and Adh7. However, there is little information available regarding the regulation of their gene expression upon severe vanillin stress, which has been shown to repress the bulk translation activity in yeast cells. Therefore, in this study, we investigated expression patterns of the ADH6 and ADH7 genes in the presence of high concentrations of vanillin. We found that although both genes were transcriptionally upregulated by vanillin stress, they showed different protein expression patterns in response to vanillin. Expression of Adh6 was constitutive and gradually decreased under vanillin stress, whereas expression of Adh7 was inducible, and, importantly, occurred under severe vanillin stress. The null mutants of ADH6 or ADH7 genes were hypersensitive to vanillin and reduced vanillin less efficiently than the wild type, confirming the importance of Adh6 and Adh7 in vanillin detoxification. Additionally, we demonstrate that the ADH7 promoter is vanillin-inducible and enables effective protein synthesis even under severe vanillin stress, and it may be useful for the improvement of vanillin-tolerance and biofuel production efficiency via modification of yeast gene expression in the presence of high concentrations of vanillin. PMID:26696995

  4. The ADH7 Promoter of Saccharomyces cerevisiae is Vanillin-Inducible and Enables mRNA Translation Under Severe Vanillin Stress

    PubMed Central

    Nguyen, Trinh T. M.; Iwaki, Aya; Izawa, Shingo

    2015-01-01

    Vanillin is one of the major phenolic aldehyde compounds derived from lignocellulosic biomass and acts as a potent fermentation inhibitor to repress the growth and fermentative ability of yeast. Vanillin can be reduced to its less toxic form, vanillyl alcohol, by the yeast NADPH-dependent medium chain alcohol dehydrogenases, Adh6 and Adh7. However, there is little information available regarding the regulation of their gene expression upon severe vanillin stress, which has been shown to repress the bulk translation activity in yeast cells. Therefore, in this study, we investigated expression patterns of the ADH6 and ADH7 genes in the presence of high concentrations of vanillin. We found that although both genes were transcriptionally upregulated by vanillin stress, they showed different protein expression patterns in response to vanillin. Expression of Adh6 was constitutive and gradually decreased under vanillin stress, whereas expression of Adh7 was inducible, and, importantly, occurred under severe vanillin stress. The null mutants of ADH6 or ADH7 genes were hypersensitive to vanillin and reduced vanillin less efficiently than the wild type, confirming the importance of Adh6 and Adh7 in vanillin detoxification. Additionally, we demonstrate that the ADH7 promoter is vanillin-inducible and enables effective protein synthesis even under severe vanillin stress, and it may be useful for the improvement of vanillin-tolerance and biofuel production efficiency via modification of yeast gene expression in the presence of high concentrations of vanillin. PMID:26696995

  5. The ADH7 Promoter of Saccharomyces cerevisiae is Vanillin-Inducible and Enables mRNA Translation Under Severe Vanillin Stress.

    PubMed

    Nguyen, Trinh T M; Iwaki, Aya; Izawa, Shingo

    2015-01-01

    Vanillin is one of the major phenolic aldehyde compounds derived from lignocellulosic biomass and acts as a potent fermentation inhibitor to repress the growth and fermentative ability of yeast. Vanillin can be reduced to its less toxic form, vanillyl alcohol, by the yeast NADPH-dependent medium chain alcohol dehydrogenases, Adh6 and Adh7. However, there is little information available regarding the regulation of their gene expression upon severe vanillin stress, which has been shown to repress the bulk translation activity in yeast cells. Therefore, in this study, we investigated expression patterns of the ADH6 and ADH7 genes in the presence of high concentrations of vanillin. We found that although both genes were transcriptionally upregulated by vanillin stress, they showed different protein expression patterns in response to vanillin. Expression of Adh6 was constitutive and gradually decreased under vanillin stress, whereas expression of Adh7 was inducible, and, importantly, occurred under severe vanillin stress. The null mutants of ADH6 or ADH7 genes were hypersensitive to vanillin and reduced vanillin less efficiently than the wild type, confirming the importance of Adh6 and Adh7 in vanillin detoxification. Additionally, we demonstrate that the ADH7 promoter is vanillin-inducible and enables effective protein synthesis even under severe vanillin stress, and it may be useful for the improvement of vanillin-tolerance and biofuel production efficiency via modification of yeast gene expression in the presence of high concentrations of vanillin.

  6. Super-paramagnetic clustering of yeast gene expression profiles

    NASA Astrophysics Data System (ADS)

    Getz, G.; Levine, E.; Domany, E.; Zhang, M. Q.

    2000-04-01

    High-density DNA arrays, used to monitor gene expression at a genomic scale, have produced vast amounts of information which require the development of efficient computational methods to analyze them. The important first step is to extract the fundamental patterns of gene expression inherent in the data. This paper describes the application of a novel clustering algorithm, super-paramagnetic clustering (SPC) to analysis of gene expression profiles that were generated recently during a study of the yeast cell cycle. SPC was used to organize genes into biologically relevant clusters that are suggestive for their co-regulation. Some of the advantages of SPC are its robustness against noise and initialization, a clear signature of cluster formation and splitting, and an unsupervised self-organized determination of the number of clusters at each resolution. Our analysis revealed interesting correlated behavior of several groups of genes which has not been previously identified.

  7. GE-Miner: integration of cluster ensemble and text mining for comprehensive gene expression analysis.

    PubMed

    Hu, Xiaohua

    2006-01-01

    Generating high quality gene clusters and identifying the underlying biological mechanism of the gene clusters are the important goals of clustering gene expression analysis. Based on this consideration, we design and develop a unified system Gene Expression Miner (GE-Miner) by integrating cluster ensemble, text clustering and multidocument summarisation and provide an environment for comprehensive gene expression data analysis. Experimental results demonstrate that our systems can obtain high quality clusters and provide concise and informative textual summary for the gene clusters.

  8. Gene expression data clustering using a multiobjective symmetry based clustering technique.

    PubMed

    Saha, Sriparna; Ekbal, Asif; Gupta, Kshitija; Bandyopadhyay, Sanghamitra

    2013-11-01

    The invention of microarrays has rapidly changed the state of biological and biomedical research. Clustering algorithms play an important role in clustering microarray data sets where identifying groups of co-expressed genes are a very difficult task. Here we have posed the problem of clustering the microarray data as a multiobjective clustering problem. A new symmetry based fuzzy clustering technique is developed to solve this problem. The effectiveness of the proposed technique is demonstrated on five publicly available benchmark data sets. Results are compared with some widely used microarray clustering techniques. Statistical and biological significance tests have also been carried out. PMID:24209942

  9. Remodelling of a homeobox gene cluster by multiple independent gene reunions in Drosophila.

    PubMed

    Chan, Carolus; Jayasekera, Suvini; Kao, Bryant; Páramo, Moisés; von Grotthuss, Marcin; Ranz, José M

    2015-01-01

    Genome clustering of homeobox genes is often thought to reflect arrangements of tandem gene duplicates maintained by advantageous coordinated gene regulation. Here we analyse the chromosomal organization of the NK homeobox genes, presumed to be part of a single cluster in the Bilaterian ancestor, across 20 arthropods. We find that the ProtoNK cluster was extensively fragmented in some lineages, showing that NK clustering in Drosophila species does not reflect selectively maintained gene arrangements. More importantly, the arrangement of NK and neighbouring genes across the phylogeny supports that, in two instances within the Drosophila genus, some cluster remnants became reunited via large-scale chromosomal rearrangements. Simulated scenarios of chromosome evolution indicate that these reunion events are unlikely unless the genome neighbourhoods harbouring the participating genes tend to colocalize in the nucleus. Our results underscore how mechanisms other than tandem gene duplication can result in paralogous gene clustering during genome evolution. PMID:25739651

  10. Prokaryotic Gene Clusters: A Rich Toolbox for Synthetic Biology

    PubMed Central

    Fischbach, Michael; Voigt, Christopher A.

    2014-01-01

    Bacteria construct elaborate nanostructures, obtain nutrients and energy from diverse sources, synthesize complex molecules, and implement signal processing to react to their environment. These complex phenotypes require the coordinated action of multiple genes, which are often encoded in a contiguous region of the genome, referred to as a gene cluster. Gene clusters sometimes contain all of the genes necessary and sufficient for a particular function. As an evolutionary mechanism, gene clusters facilitate the horizontal transfer of the complete function between species. Here, we review recent work on a number of clusters whose functions are relevant to biotechnology. Engineering these clusters has been hindered by their regulatory complexity, the need to balance the expression of many genes, and a lack of tools to design and manipulate DNA at this scale. Advances in synthetic biology will enable the large-scale bottom-up engineering of the clusters to optimize their functions, wake up cryptic clusters, or to transfer them between organisms. Understanding and manipulating gene clusters will move towards an era of genome engineering, where multiple functions can be “mixed-and-matched” to create a designer organism. PMID:21154668

  11. Recommending pathway genes using a compendium of clustering solutions.

    PubMed

    Ng, David M; Woehrmann, Marcos H; Stuart, Joshua M

    2007-01-01

    A common approach for identifying pathways from gene expression data is to cluster the genes without using prior information about a pathway, which often identifies only the dominant coexpression groups. Recommender systems are well-suited for using the known genes of a pathway to identify the appropriate experiments for predicting new members. However, existing systems, such as the GeneRecommender, ignore how genes naturally group together within specific experiments. We present a collaborative filtering approach which uses the pattern of how genes cluster together in different experiments to recommend new genes in a pathway. Clusters are first identified within a single experiment series. Informative clusters, in which the user-supplied query genes appear together, are identified. New genes that cluster with the known genes, in a significant fraction of the informative clusters, are recommended. We implemented a prototype of our system and measured its performance on hundreds of pathways. We find that our method performs as well as an established approach while significantly increasing the speed and scalability of searching large datasets. [Supplemental material is available online at sysbio.soe.ucsc.edu/cluegene/psb07.

  12. Bioinformatics Prediction of Polyketide Synthase Gene Clusters from Mycosphaerella fijiensis.

    PubMed

    Noar, Roslyn D; Daub, Margaret E

    2016-01-01

    Mycosphaerella fijiensis, causal agent of black Sigatoka disease of banana, is a Dothideomycete fungus closely related to fungi that produce polyketides important for plant pathogenicity. We utilized the M. fijiensis genome sequence to predict PKS genes and their gene clusters and make bioinformatics predictions about the types of compounds produced by these clusters. Eight PKS gene clusters were identified in the M. fijiensis genome, placing M. fijiensis into the 23rd percentile for the number of PKS genes compared to other Dothideomycetes. Analysis of the PKS domains identified three of the PKS enzymes as non-reducing and two as highly reducing. Gene clusters contained types of genes frequently found in PKS clusters including genes encoding transporters, oxidoreductases, methyltransferases, and non-ribosomal peptide synthases. Phylogenetic analysis identified a putative PKS cluster encoding melanin biosynthesis. None of the other clusters were closely aligned with genes encoding known polyketides, however three of the PKS genes fell into clades with clusters encoding alternapyrone, fumonisin, and solanapyrone produced by Alternaria and Fusarium species. A search for homologs among available genomic sequences from 103 Dothideomycetes identified close homologs (>80% similarity) for six of the PKS sequences. One of the PKS sequences was not similar (< 60% similarity) to sequences in any of the 103 genomes, suggesting that it encodes a unique compound. Comparison of the M. fijiensis PKS sequences with those of two other banana pathogens, M. musicola and M. eumusae, showed that these two species have close homologs to five of the M. fijiensis PKS sequences, but three others were not found in either species. RT-PCR and RNA-Seq analysis showed that the melanin PKS cluster was down-regulated in infected banana as compared to growth in culture. Three other clusters, however were strongly upregulated during disease development in banana, suggesting that they may encode

  13. Bioinformatics Prediction of Polyketide Synthase Gene Clusters from Mycosphaerella fijiensis

    PubMed Central

    Noar, Roslyn D.; Daub, Margaret E.

    2016-01-01

    Mycosphaerella fijiensis, causal agent of black Sigatoka disease of banana, is a Dothideomycete fungus closely related to fungi that produce polyketides important for plant pathogenicity. We utilized the M. fijiensis genome sequence to predict PKS genes and their gene clusters and make bioinformatics predictions about the types of compounds produced by these clusters. Eight PKS gene clusters were identified in the M. fijiensis genome, placing M. fijiensis into the 23rd percentile for the number of PKS genes compared to other Dothideomycetes. Analysis of the PKS domains identified three of the PKS enzymes as non-reducing and two as highly reducing. Gene clusters contained types of genes frequently found in PKS clusters including genes encoding transporters, oxidoreductases, methyltransferases, and non-ribosomal peptide synthases. Phylogenetic analysis identified a putative PKS cluster encoding melanin biosynthesis. None of the other clusters were closely aligned with genes encoding known polyketides, however three of the PKS genes fell into clades with clusters encoding alternapyrone, fumonisin, and solanapyrone produced by Alternaria and Fusarium species. A search for homologs among available genomic sequences from 103 Dothideomycetes identified close homologs (>80% similarity) for six of the PKS sequences. One of the PKS sequences was not similar (< 60% similarity) to sequences in any of the 103 genomes, suggesting that it encodes a unique compound. Comparison of the M. fijiensis PKS sequences with those of two other banana pathogens, M. musicola and M. eumusae, showed that these two species have close homologs to five of the M. fijiensis PKS sequences, but three others were not found in either species. RT-PCR and RNA-Seq analysis showed that the melanin PKS cluster was down-regulated in infected banana as compared to growth in culture. Three other clusters, however were strongly upregulated during disease development in banana, suggesting that they may encode

  14. Sesterterpene ophiobolin biosynthesis involving multiple gene clusters in Aspergillus ustus

    PubMed Central

    Chai, Hangzhen; Yin, Ru; Liu, Yongfeng; Meng, Huiying; Zhou, Xianqiang; Zhou, Guolin; Bi, Xupeng; Yang, Xue; Zhu, Tonghan; Zhu, Weiming; Deng, Zixin; Hong, Kui

    2016-01-01

    Terpenoids are the most diverse and abundant natural products among which sesterterpenes account for less than 2%, with very few reports on their biosynthesis. Ophiobolins are tricyclic 5–8–5 ring sesterterpenes with potential pharmaceutical application. Aspergillus ustus 094102 from mangrove rizhosphere produces ophiobolin and other terpenes. We obtained five gene cluster knockout mutants, with altered ophiobolin yield using genome sequencing and in silico analysis, combined with in vivo genetic manipulation. Involvement of the five gene clusters in ophiobolin synthesis was confirmed by investigation of the five key terpene synthesis relevant enzymes in each gene cluster, either by gene deletion and complementation or in vitro verification of protein function. The results demonstrate that ophiobolin skeleton biosynthesis involves five gene clusters, which are responsible for C15, C20, C25, and C30 terpenoid biosynthesis. PMID:27273151

  15. Prodigiosin biosynthesis gene cluster in the roseophilin producer Streptomyces griseoviridis.

    PubMed

    Kawasaki, Takashi; Sakurai, Fumi; Nagatsuka, Shun-ya; Hayakawa, Yoichi

    2009-05-01

    Streptomyces griseoviridis 2464-S5 produces prodigiosin R1, a tripyrrole antibiotic, and roseophilin, a structurally related compound containing two pyrrole and one furan rings. A gene cluster for the biosynthesis of a prodigiosin was identified in S. griseoviridis. The cluster consisted of 24 open reading frames, including 21 genes (rphD-rphZ) homologous to prodigiosin biosynthesis genes in the red cluster in Streptomyces coelicolor A3(2). The expression of rphN in S. coelicolor lacking redN restored the production of prodigiosin.

  16. Genomic analyses of bacterial porin-cytochrome gene clusters

    DOE PAGES

    Shi, Liang; Fredrickson, James K.; Zachara, John M.

    2014-11-26

    In this study, the porin-cytochrome (Pcc) protein complex is responsible for trans-outer membrane electron transfer during extracellular reduction of Fe(III) by the dissimilatory metal-reducing bacterium Geobacter sulfurreducens PCA. The identified and characterized Pcc complex of G. sulfurreducens PCA consists of a porin-like outer-membrane protein, a periplasmic 8-heme c type cytochrome (c-Cyt) and an outer-membrane 12-heme c-Cyt, and the genes encoding the Pcc proteins are clustered in the same regions of genome (i.e., the pcc gene clusters) of G. sulfurreducens PCA. A survey of additionally microbial genomes has identified the pcc gene clusters in all sequenced Geobacter spp. and other bacteriamore » from six different phyla, including Anaeromyxobacter dehalogenans 2CP-1, A. dehalogenans 2CP-C, Anaeromyxobacter sp. K, Candidatus Kuenenia stuttgartiensis, Denitrovibrio acetiphilus DSM 12809, Desulfurispirillum indicum S5, Desulfurivibrio alkaliphilus AHT2, Desulfurobacterium thermolithotrophum DSM 11699, Desulfuromonas acetoxidans DSM 684, Ignavibacterium album JCM 16511, and Thermovibrio ammonificans HB-1. The numbers of genes in the pcc gene clusters vary, ranging from two to nine. Similar to the metal-reducing (Mtr) gene clusters of other Fe(III)-reducing bacteria, such as Shewanella spp., additional genes that encode putative c-Cyts with predicted cellular localizations at the cytoplasmic membrane, periplasm and outer membrane often associate with the pcc gene clusters. This suggests that the Pcc-associated c-Cyts may be part of the pathways for extracellular electron transfer reactions. The presence of pcc gene clusters in the microorganisms that do not reduce solid-phase Fe(III) and Mn(IV) oxides, such as D. alkaliphilus AHT2 and I. album JCM 16511, also suggests that some of the pcc gene clusters may be involved in extracellular electron transfer reactions with the substrates other than Fe(III) and Mn(IV) oxides.« less

  17. Relevant and significant supervised gene clusters for microarray cancer classification.

    PubMed

    Maji, Pradipta; Das, Chandra

    2012-06-01

    An important application of microarray data in functional genomics is to classify samples according to their gene expression profiles such as to classify cancer versus normal samples or to classify different types or subtypes of cancer. One of the major tasks with gene expression data is to find co-regulated gene groups whose collective expression is strongly associated with sample categories. In this regard, a gene clustering algorithm is proposed to group genes from microarray data. It directly incorporates the information of sample categories in the grouping process for finding groups of co-regulated genes with strong association to the sample categories, yielding a supervised gene clustering algorithm. The average expression of the genes from each cluster acts as its representative. Some significant representatives are taken to form the reduced feature set to build the classifiers for cancer classification. The mutual information is used to compute both gene-gene redundancy and gene-class relevance. The performance of the proposed method, along with a comparison with existing methods, is studied on six cancer microarray data sets using the predictive accuracy of naive Bayes classifier, K-nearest neighbor rule, and support vector machine. An important finding is that the proposed algorithm is shown to be effective for identifying biologically significant gene clusters with excellent predictive capability. PMID:22552589

  18. Determination of the effects of alcohol dehydrogenase (ADH) 1B and ADH1C polymorphisms on alcohol dependence in Turkey.

    PubMed

    Aktas, Ekin Ozgur; Kocak, Aytaç; Senol, Ender; Celik, Handan Ak; Coskunol, Hakan; Berdeli, Afig; Aydin, Hikmet Hakan

    2012-03-01

    Alcoholism is a complex genetically influenced disorder which refers to alcohol abuse and alcohol dependence. There are controversial results on the role of gene polymorphisms in alcohol dependence in the literature. Differences in population groups and selective inclusion criteria for alcohol dependence may affect results. In this study, we investigated the role of ADH1B Arg48His (rs1229984) and, ADH1C Ile350Val (rs698) gene polymorphisms in Turkish population. 100 healthy volunteers and 75 patients who were admitted to Ege University Alcohol Dependence Unit enrolled in the study. We found significant increase both in ADH1B (Arg48His) polymorphism Arg allele and Arg/Arg genotype frequency in patients. No profound connection between alcohol dependence and ADH1C Ile350Val gene polymorphism was detected. Alcohol dependence is an important health problem that depends on many genetic and environmental factors but we think that it is possible to interpret genetic risk for developing early diagnostic methods and treatment strategies by comprehensive linkage and association studies.

  19. Hox gene clusters in the Indonesian coelacanth, Latimeria menadoensis.

    PubMed

    Koh, Esther G L; Lam, Kevin; Christoffels, Alan; Erdmann, Mark V; Brenner, Sydney; Venkatesh, Byrappa

    2003-02-01

    The Hox genes encode transcription factors that play a key role in specifying body plans of metazoans. They are organized into clusters that contain up to 13 paralogue group members. The complex morphology of vertebrates has been attributed to the duplication of Hox clusters during vertebrate evolution. In contrast to the single Hox cluster in the amphioxus (Branchiostoma floridae), an invertebrate-chordate, mammals have four clusters containing 39 Hox genes. Ray-finned fishes (Actinopterygii) such as zebrafish and fugu possess more than four Hox clusters. The coelacanth occupies a basal phylogenetic position among lobe-finned fishes (Sarcopterygii), which gave rise to the tetrapod lineage. The lobe fins of sarcopterygians are considered to be the evolutionary precursors of tetrapod limbs. Thus, the characterization of Hox genes in the coelacanth should provide insights into the origin of tetrapod limbs. We have cloned the complete second exon of 33 Hox genes from the Indonesian coelacanth, Latimeria menadoensis, by extensive PCR survey and genome walking. Phylogenetic analysis shows that 32 of these genes have orthologs in the four mammalian HOX clusters, including three genes (HoxA6, D1, and D8) that are absent in ray-finned fishes. The remaining coelacanth gene is an ortholog of hoxc1 found in zebrafish but absent in mammals. Our results suggest that coelacanths have four Hox clusters bearing a gene complement more similar to mammals than to ray-finned fishes, but with an additional gene, HoxC1, which has been lost during the evolution of mammals from lobe-finned fishes.

  20. SMART: Unique Splitting-While-Merging Framework for Gene Clustering

    PubMed Central

    Fa, Rui; Roberts, David J.; Nandi, Asoke K.

    2014-01-01

    Successful clustering algorithms are highly dependent on parameter settings. The clustering performance degrades significantly unless parameters are properly set, and yet, it is difficult to set these parameters a priori. To address this issue, in this paper, we propose a unique splitting-while-merging clustering framework, named “splitting merging awareness tactics” (SMART), which does not require any a priori knowledge of either the number of clusters or even the possible range of this number. Unlike existing self-splitting algorithms, which over-cluster the dataset to a large number of clusters and then merge some similar clusters, our framework has the ability to split and merge clusters automatically during the process and produces the the most reliable clustering results, by intrinsically integrating many clustering techniques and tasks. The SMART framework is implemented with two distinct clustering paradigms in two algorithms: competitive learning and finite mixture model. Nevertheless, within the proposed SMART framework, many other algorithms can be derived for different clustering paradigms. The minimum message length algorithm is integrated into the framework as the clustering selection criterion. The usefulness of the SMART framework and its algorithms is tested in demonstration datasets and simulated gene expression datasets. Moreover, two real microarray gene expression datasets are studied using this approach. Based on the performance of many metrics, all numerical results show that SMART is superior to compared existing self-splitting algorithms and traditional algorithms. Three main properties of the proposed SMART framework are summarized as: (1) needing no parameters dependent on the respective dataset or a priori knowledge about the datasets, (2) extendible to many different applications, (3) offering superior performance compared with counterpart algorithms. PMID:24714159

  1. Examining emergence of functional gene clustering in a simulated evolution.

    PubMed

    Yerushalmi, Uri; Teicher, Mina

    2007-10-01

    Recent research suggests that rather than being random, gene order may be coupled with gene functionality. These findings may be explained by mechanisms that require physical proximity such as co-expression and co-regulation. Alternatively, they may be due to evolutionary-dynamics forces, as expressed in genetic drift or linkage disequilibrium. This paper proposes a biologically plausible model for evolutionary development. Using the model, which includes natural selection and the development of gene networks and cellular organisms, the co-evolution of recombination rate and gene functionality is examined. The results presented here are compatible with previous biological findings showing that functionally related genes are clustered. These results imply that evolutionary pressure in a complex environment is sufficient for the emergence of gene order that is coupled with functionality. They shed further light on the mechanisms that may cause such gene clusters.

  2. Overexpression of the genes PDC1 and ADH1 activates glycerol conversion to ethanol in the thermotolerant yeast Ogataea (Hansenula) polymorpha.

    PubMed

    Kata, Iwona; Semkiv, Marta V; Ruchala, Justyna; Dmytruk, Kostyantyn V; Sibirny, Andriy A

    2016-08-01

    Conversion of byproduct from biodiesel production glycerol to high-value compounds is of great importance. Ethanol is considered a promising product of glycerol bioconversion. The methylotrophic thermotolerant yeast Ogataea (Hansenula) polymorpha is of great interest for this purpose as the glycerol byproduct contains methanol and heavy metals as contaminants, and this yeast utilizes methanol and is relatively resistant to heavy metals. Besides, O. polymorpha shows robust growth on glycerol and produces ethanol from various carbon sources. The thermotolerance of this yeast is an additional advantage, allowing increased fermentation temperature to 45-48 °C, leading to increased rate of the fermentation process and a fall in the cost of distillation. The wild-type strain of O. polymorpha produces insignificant amounts of ethanol from glycerol (0.8 g/l). Overexpression of PDC1 coding for pyruvate decarboxylase enhanced ethanol production up to 3.1 g/l, whereas simultaneous overexpression of PDC1 and ADH1 (coding for alcohol dehydrogenase) led to further increase in ethanol production from glycerol. Moreover, the increased temperature of fermentation up to 45 °C stimulated the production of ethanol from glycerol used as the only carbon source up to 5.0 g/l, which exceeds the data obtained by methylotrophic yeast strains reported so far. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27256876

  3. Functional optimization of gene clusters by combinatorial design and assembly.

    PubMed

    Smanski, Michael J; Bhatia, Swapnil; Zhao, Dehua; Park, YongJin; B A Woodruff, Lauren; Giannoukos, Georgia; Ciulla, Dawn; Busby, Michele; Calderon, Johnathan; Nicol, Robert; Gordon, D Benjamin; Densmore, Douglas; Voigt, Christopher A

    2014-12-01

    Large microbial gene clusters encode useful functions, including energy utilization and natural product biosynthesis, but genetic manipulation of such systems is slow, difficult and complicated by complex regulation. We exploit the modularity of a refactored Klebsiella oxytoca nitrogen fixation (nif) gene cluster (16 genes, 103 parts) to build genetic permutations that could not be achieved by starting from the wild-type cluster. Constraint-based combinatorial design and DNA assembly are used to build libraries of radically different cluster architectures by varying part choice, gene order, gene orientation and operon occupancy. We construct 84 variants of the nifUSVWZM operon, 145 variants of the nifHDKY operon, 155 variants of the nifHDKYENJ operon and 122 variants of the complete 16-gene pathway. The performance and behavior of these variants are characterized by nitrogenase assay and strand-specific RNA sequencing (RNA-seq), and the results are incorporated into subsequent design cycles. We have produced a fully synthetic cluster that recovers 57% of wild-type activity. Our approach allows the performance of genetic parts to be quantified simultaneously in hundreds of genetic contexts. This parallelized design-build-test-learn cycle, which can access previously unattainable regions of genetic space, should provide a useful, fast tool for genetic optimization and hypothesis testing.

  4. Characterization of the largest effector gene cluster of Ustilago maydis.

    PubMed

    Brefort, Thomas; Tanaka, Shigeyuki; Neidig, Nina; Doehlemann, Gunther; Vincon, Volker; Kahmann, Regine

    2014-07-01

    In the genome of the biotrophic plant pathogen Ustilago maydis, many of the genes coding for secreted protein effectors modulating virulence are arranged in gene clusters. The vast majority of these genes encode novel proteins whose expression is coupled to plant colonization. The largest of these gene clusters, cluster 19A, encodes 24 secreted effectors. Deletion of the entire cluster results in severe attenuation of virulence. Here we present the functional analysis of this genomic region. We show that a 19A deletion mutant behaves like an endophyte, i.e. is still able to colonize plants and complete the infection cycle. However, tumors, the most conspicuous symptoms of maize smut disease, are only rarely formed and fungal biomass in infected tissue is significantly reduced. The generation and analysis of strains carrying sub-deletions identified several genes significantly contributing to tumor formation after seedling infection. Another of the effectors could be linked specifically to anthocyanin induction in the infected tissue. As the individual contributions of these genes to tumor formation were small, we studied the response of maize plants to the whole cluster mutant as well as to several individual mutants by array analysis. This revealed distinct plant responses, demonstrating that the respective effectors have discrete plant targets. We propose that the analysis of plant responses to effector mutant strains that lack a strong virulence phenotype may be a general way to visualize differences in effector function.

  5. Fusarium oxysporum Adh1 has dual fermentative and oxidative functions and is involved in fungal virulence in tomato plants.

    PubMed

    Corrales Escobosa, Alma Rosa; Rangel Porras, Rosa Angelica; Meza Carmen, Victor; Gonzalez Hernandez, Gloria Angélica; Torres Guzman, Juan Carlos; Wrobel, Kazimierz; Wrobel, Katarzyna; Roncero, M Isabel G; Gutierrez Corona, J Felix

    2011-09-01

    An alcohol dehydrogenase gene, adh1, has been identified in the vascular wilt fungus Fusarium oxysporum f. sp. lycopersici. Reverse transcription polymerase chain reaction (RT-PCR) analysis revealed that adh1 is highly expressed in mycelia grown in potato dextrose liquid medium (PDB) under hypoxic conditions, as compared to mycelia grown under aerobic conditions. One spontaneous allyl alcohol-resistant (Ally(R)) mutant exhibited insertion of an incomplete F.oxysporum transposable element, while another mutant contained a short (13 nucleotide) deletion, in both cases interrupting the coding region of the adh1 gene. These mutations caused deficiency in Adh activity due to loss of the main constitutive isoform of Adh1, as well as alteration of different physiological parameters related to carbon and energy metabolism, including the ability to use ethanol as a carbon source under aerobic conditions; impaired growth under hypoxic conditions with glucose as the carbon source; and diminished production of ethanol in glucose-containing medium. Interestingly, the adh1 mutations resulted in a significant delay in fungal disease development in tomato plants. Complementation with the wild-type adh1 allele repaired all defects caused by mutation, indicating that the product of the adh1 gene has dual enzymatic functions (fermentative and oxidative), depending on culture conditions, and is also required for full fungal virulence.

  6. Identification of nitrogen-fixing genes and gene clusters from metagenomic library of acid mine drainage.

    PubMed

    Dai, Zhimin; Guo, Xue; Yin, Huaqun; Liang, Yili; Cong, Jing; Liu, Xueduan

    2014-01-01

    Biological nitrogen fixation is an essential function of acid mine drainage (AMD) microbial communities. However, most acidophiles in AMD environments are uncultured microorganisms and little is known about the diversity of nitrogen-fixing genes and structure of nif gene cluster in AMD microbial communities. In this study, we used metagenomic sequencing to isolate nif genes in the AMD microbial community from Dexing Copper Mine, China. Meanwhile, a metagenome microarray containing 7,776 large-insertion fosmids was constructed to screen novel nif gene clusters. Metagenomic analyses revealed that 742 sequences were identified as nif genes including structural subunit genes nifH, nifD, nifK and various additional genes. The AMD community is massively dominated by the genus Acidithiobacillus. However, the phylogenetic diversity of nitrogen-fixing microorganisms is much higher than previously thought in the AMD community. Furthermore, a 32.5-kb genomic sequence harboring nif, fix and associated genes was screened by metagenome microarray. Comparative genome analysis indicated that most nif genes in this cluster are most similar to those of Herbaspirillum seropedicae, but the organization of the nif gene cluster had significant differences from H. seropedicae. Sequence analysis and reverse transcription PCR also suggested that distinct transcription units of nif genes exist in this gene cluster. nifQ gene falls into the same transcription unit with fixABCX genes, which have not been reported in other diazotrophs before. All of these results indicated that more novel diazotrophs survive in the AMD community.

  7. Identification of Nitrogen-Fixing Genes and Gene Clusters from Metagenomic Library of Acid Mine Drainage

    PubMed Central

    Yin, Huaqun; Liang, Yili; Cong, Jing; Liu, Xueduan

    2014-01-01

    Biological nitrogen fixation is an essential function of acid mine drainage (AMD) microbial communities. However, most acidophiles in AMD environments are uncultured microorganisms and little is known about the diversity of nitrogen-fixing genes and structure of nif gene cluster in AMD microbial communities. In this study, we used metagenomic sequencing to isolate nif genes in the AMD microbial community from Dexing Copper Mine, China. Meanwhile, a metagenome microarray containing 7,776 large-insertion fosmids was constructed to screen novel nif gene clusters. Metagenomic analyses revealed that 742 sequences were identified as nif genes including structural subunit genes nifH, nifD, nifK and various additional genes. The AMD community is massively dominated by the genus Acidithiobacillus. However, the phylogenetic diversity of nitrogen-fixing microorganisms is much higher than previously thought in the AMD community. Furthermore, a 32.5-kb genomic sequence harboring nif, fix and associated genes was screened by metagenome microarray. Comparative genome analysis indicated that most nif genes in this cluster are most similar to those of Herbaspirillum seropedicae, but the organization of the nif gene cluster had significant differences from H. seropedicae. Sequence analysis and reverse transcription PCR also suggested that distinct transcription units of nif genes exist in this gene cluster. nifQ gene falls into the same transcription unit with fixABCX genes, which have not been reported in other diazotrophs before. All of these results indicated that more novel diazotrophs survive in the AMD community. PMID:24498417

  8. Inferring the evolutionary history of gene clusters from phylogenetic and gene order data.

    PubMed

    Lajoie, Mathieu; Bertrand, Denis; El-Mabrouk, Nadia

    2010-04-01

    Gene duplication is frequent within gene clusters and plays a fundamental role in evolution by providing a source of new genetic material upon which natural selection can act. Although classical phylogenetic inference methods provide some insight into the evolutionary history of a gene cluster, they are not sufficient alone to differentiate single- from multiple gene duplication events and to answer other questions regarding the nature and size of evolutionary events. In this paper, we present an algorithm allowing to infer a set of optimal evolutionary histories for a gene cluster in a single species, according to a general cost model involving variable length duplications (in tandem or inverted), deletions, and inversions. We applied our algorithm to the human olfactory receptor and protocadherin gene clusters, showing that the duplication size distribution differs significantly between the two gene families. The algorithm is available through a web interface at http://www-lbit.iro.umontreal.ca/DILTAG/.

  9. A genetic analysis of Adh1 regulation

    SciTech Connect

    Freeling, M.

    1992-01-01

    The overall goal of our research proposal is to understand the meaning of the various cis-acting sites responsible for AdH1 expression in the entire maize plant. Progress is reported in the following areas: Studies on the TATA box and analysis of revertants of the Adh1-3F1124 allele; screening for more different mutants that affect Adh1 expression differentially; studies on cis-acting sequences required for root-specific Adh1 expression; refinement of the use of the particle gun; and functional analysis of a non- glycolytic anaerobic protein.

  10. Evolution of the Leucine Gene Cluster in Buchnera aphidicola: Insights from Chromosomal Versions of the Cluster

    PubMed Central

    Sabater-Muñoz, Beatriz; van Ham, Roeland C. H. J.; Moya, Andrés; Silva, Francisco J.; Latorre, Amparo

    2004-01-01

    In Buchnera aphidicola strains associated with the aphid subfamilies Thelaxinae, Lachninae, Pterocommatinae, and Aphidinae, the four leucine genes (leuA, -B, -C, and -D) are located on a plasmid. However, these genes are located on the main chromosome in B. aphidicola strains associated with the subfamilies Pemphiginae and Chaitophorinae. The sequence of the chromosomal fragment containing the leucine cluster and flanking genes has different positions in the chromosome in B. aphidicola strains associated with three tribes of the subfamily Pemphiginae and one tribe of the subfamily Chaitophorinae. Due to the extreme gene order conservation of the B. aphidicola genomes, the variability in the position of the leucine cluster in the chromosome may be interpreted as resulting from independent insertions from an ancestral plasmid-borne leucine gene. These findings do not support a chromosomal origin for the leucine genes in the ancestral B. aphidicola and do support a back transfer evolutionary scenario from a plasmid to the main chromosome. PMID:15090505

  11. Recommended nomenclature for the vertebrate alcohol dehydrogenase gene family.

    PubMed

    Duester, G; Farrés, J; Felder, M R; Holmes, R S; Höög, J O; Parés, X; Plapp, B V; Yin, S J; Jörnvall, H

    1999-08-01

    The alcohol dehydrogenase (ADH) gene family encodes enzymes that metabolize a wide variety of substrates, including ethanol, retinol, other aliphatic alcohols, hydroxysteroids, and lipid peroxidation products. Studies on 19 vertebrate animals have identified ADH orthologs across several species, and this has now led to questions of how best to name ADH proteins and genes. Seven distinct classes of vertebrate ADH encoded by non-orthologous genes have been defined based upon sequence homology as well as unique catalytic properties or gene expression patterns. Each class of vertebrate ADH shares <70% sequence identity with other classes of ADH in the same species. Classes may be further divided into multiple closely related isoenzymes sharing >80% sequence identity such as the case for class I ADH where humans have three class I ADH genes, horses have two, and mice have only one. Presented here is a nomenclature that uses the widely accepted vertebrate ADH class system as its basis. It follows the guidelines of human and mouse gene nomenclature committees, which recommend coordinating names across species boundaries and eliminating Roman numerals and Greek symbols. We recommend that enzyme subunits be referred to by the symbol "ADH" (alcohol dehydrogenase) followed by an Arabic number denoting the class; i.e. ADH1 for class I ADH. For genes we recommend the italicized root symbol "ADH" for human and "Adh" for mouse, followed by the appropriate Arabic number for the class; i.e. ADH1 or Adh1 for class I ADH genes. For organisms where multiple species-specific isoenzymes exist within a class, we recommend adding a capital letter after the Arabic number; i.e. ADH1A, ADH1B, and ADH1C for human alpha, beta, and gamma class I ADHs, respectively. This nomenclature will accommodate newly discovered members of the vertebrate ADH family, and will facilitate functional and evolutionary studies. PMID:10424757

  12. Genomic analyses of bacterial porin-cytochrome gene clusters

    SciTech Connect

    Shi, Liang; Fredrickson, James K.; Zachara, John M.

    2014-11-26

    In this study, the porin-cytochrome (Pcc) protein complex is responsible for trans-outer membrane electron transfer during extracellular reduction of Fe(III) by the dissimilatory metal-reducing bacterium Geobacter sulfurreducens PCA. The identified and characterized Pcc complex of G. sulfurreducens PCA consists of a porin-like outer-membrane protein, a periplasmic 8-heme c type cytochrome (c-Cyt) and an outer-membrane 12-heme c-Cyt, and the genes encoding the Pcc proteins are clustered in the same regions of genome (i.e., the pcc gene clusters) of G. sulfurreducens PCA. A survey of additionally microbial genomes has identified the pcc gene clusters in all sequenced Geobacter spp. and other bacteria from six different phyla, including Anaeromyxobacter dehalogenans 2CP-1, A. dehalogenans 2CP-C, Anaeromyxobacter sp. K, Candidatus Kuenenia stuttgartiensis, Denitrovibrio acetiphilus DSM 12809, Desulfurispirillum indicum S5, Desulfurivibrio alkaliphilus AHT2, Desulfurobacterium thermolithotrophum DSM 11699, Desulfuromonas acetoxidans DSM 684, Ignavibacterium album JCM 16511, and Thermovibrio ammonificans HB-1. The numbers of genes in the pcc gene clusters vary, ranging from two to nine. Similar to the metal-reducing (Mtr) gene clusters of other Fe(III)-reducing bacteria, such as Shewanella spp., additional genes that encode putative c-Cyts with predicted cellular localizations at the cytoplasmic membrane, periplasm and outer membrane often associate with the pcc gene clusters. This suggests that the Pcc-associated c-Cyts may be part of the pathways for extracellular electron transfer reactions. The presence of pcc gene clusters in the microorganisms that do not reduce solid-phase Fe(III) and Mn(IV) oxides, such as D. alkaliphilus AHT2 and I. album JCM 16511, also suggests that some of the pcc gene clusters may be involved in extracellular

  13. Identification of the Scopularide Biosynthetic Gene Cluster in Scopulariopsis brevicaulis

    PubMed Central

    Lukassen, Mie Bech; Saei, Wagma; Sondergaard, Teis Esben; Tamminen, Anu; Kumar, Abhishek; Kempken, Frank; Wiebe, Marilyn G.; Sørensen, Jens Laurids

    2015-01-01

    Scopularide A is a promising potent anticancer lipopeptide isolated from a marine derived Scopulariopsis brevicaulis strain. The compound consists of a reduced carbon chain (3-hydroxy-methyldecanoyl) attached to five amino acids (glycine, l-valine, d-leucine, l-alanine, and l-phenylalanine). Using the newly sequenced S. brevicaulis genome we were able to identify the putative biosynthetic gene cluster using genetic information from the structurally related emericellamide A from Aspergillus nidulans and W493-B from Fusarium pseudograminearum. The scopularide A gene cluster includes a nonribosomal peptide synthetase (NRPS1), a polyketide synthase (PKS2), a CoA ligase, an acyltransferase, and a transcription factor. Homologous recombination was low in S. brevicaulis so the local transcription factor was integrated randomly under a constitutive promoter, which led to a three to four-fold increase in scopularide A production. This indirectly verifies the identity of the proposed biosynthetic gene cluster. PMID:26184239

  14. Phage cluster relationships identified through single gene analysis

    PubMed Central

    2013-01-01

    Background Phylogenetic comparison of bacteriophages requires whole genome approaches such as dotplot analysis, genome pairwise maps, and gene content analysis. Currently mycobacteriophages, a highly studied phage group, are categorized into related clusters based on the comparative analysis of whole genome sequences. With the recent explosion of phage isolation, a simple method for phage cluster prediction would facilitate analysis of crude or complex samples without whole genome isolation and sequencing. The hypothesis of this study was that mycobacteriophage-cluster prediction is possible using comparison of a single, ubiquitous, semi-conserved gene. Tape Measure Protein (TMP) was selected to test the hypothesis because it is typically the longest gene in mycobacteriophage genomes and because regions within the TMP gene are conserved. Results A single gene, TMP, identified the known Mycobacteriophage clusters and subclusters using a Gepard dotplot comparison or a phylogenetic tree constructed from global alignment and maximum likelihood comparisons. Gepard analysis of 247 mycobacteriophage TMP sequences appropriately recovered 98.8% of the subcluster assignments that were made by whole-genome comparison. Subcluster-specific primers within TMP allow for PCR determination of the mycobacteriophage subcluster from DNA samples. Using the single-gene comparison approach for siphovirus coliphages, phage groupings by TMP comparison reflected relationships observed in a whole genome dotplot comparison and confirm the potential utility of this approach to another widely studied group of phages. Conclusions TMP sequence comparison and PCR results support the hypothesis that a single gene can be used for distinguishing phage cluster and subcluster assignments. TMP single-gene analysis can quickly and accurately aid in mycobacteriophage classification. PMID:23777341

  15. Identification of caerulomycin A gene cluster implicates a tailoring amidohydrolase.

    PubMed

    Zhu, Yiguang; Fu, Peng; Lin, Qinheng; Zhang, Guangtao; Zhang, Haibo; Li, Sumei; Ju, Jianhua; Zhu, Weiming; Zhang, Changsheng

    2012-06-01

    The biosynthetic gene cluster for caerulomycin A (1) was cloned and characterized from the marine actinomycete Actinoalloteichus cyanogriseus WH1-2216-6, which revealed an unusual hybrid polyketide synthase (PKS)/nonribosomal peptide synthetase (NRPS) system. The crmL disruption mutant accumulated caerulomycin L (2) with an extended L-leucine at C-7, implicating an amidohydrolase activity for CrmL. The leucine-removing activity was confirmed for crude CrmL enzymes. Heterologous expression of the 1 gene cluster led to 1 production in Streptomyces coelicolor.

  16. Expression, crystallization and preliminary X-ray crystallographic analysis of alcohol dehydrogenase (ADH) from Kangiella koreensis.

    PubMed

    Ngo, Ho-Phuong-Thuy; Hong, Seung-Hye; Hong, Myoung-Ki; Pham, Tan-Viet; Oh, Deok-Kun; Kang, Lin-Woo

    2013-09-01

    Alcohol dehydrogenases (ADHs) are a group of dehydrogenase enzymes that facilitate the interconversion between alcohols and aldehydes or ketones with the reduction of NAD(+) to NADH. In bacteria, some alcohol dehydrogenases catalyze the opposite reaction as part of fermentation to ensure a constant supply of NAD(+). The adh gene from Kangiella koreensis was cloned and the protein (KkADH) was expressed, purified and crystallized. A KkADH crystal diffracted to 2.5 Å resolution and belonged to the monoclinic space group P2(1), with unit-cell parameters a = 94.1, b = 80.9, c = 115.6 Å, β = 111.9°. Four monomers were present in the asymmetric unit, with a corresponding VM of 2.55 Å(3) Da(-1) and a solvent content of 51.8%.

  17. Expression, crystallization and preliminary X-ray crystallographic analysis of alcohol dehydrogenase (ADH) from Kangiella koreensis

    PubMed Central

    Ngo, Ho-Phuong-Thuy; Hong, Seung-Hye; Hong, Myoung-Ki; Pham, Tan-Viet; Oh, Deok-Kun; Kang, Lin-Woo

    2013-01-01

    Alcohol dehydrogenases (ADHs) are a group of dehydrogenase enzymes that facilitate the interconversion between alcohols and aldehydes or ketones with the reduction of NAD+ to NADH. In bacteria, some alcohol dehydrogenases catalyze the opposite reaction as part of fermentation to ensure a constant supply of NAD+. The adh gene from Kangiella koreensis was cloned and the protein (KkADH) was expressed, purified and crystallized. A KkADH crystal diffracted to 2.5 Å resolution and belonged to the monoclinic space group P21, with unit-cell parameters a = 94.1, b = 80.9, c = 115.6 Å, β = 111.9°. Four monomers were present in the asymmetric unit, with a corresponding V M of 2.55 Å3 Da−1 and a solvent content of 51.8%. PMID:23989158

  18. IGSA: Individual Gene Sets Analysis, including Enrichment and Clustering

    PubMed Central

    Liu, Lei; Ma, Hongzhe; Yang, Jingbo; Xie, Hongbo; Liu, Bo; Jin, Qing

    2016-01-01

    Analysis of gene sets has been widely applied in various high-throughput biological studies. One weakness in the traditional methods is that they neglect the heterogeneity of genes expressions in samples which may lead to the omission of some specific and important gene sets. It is also difficult for them to reflect the severities of disease and provide expression profiles of gene sets for individuals. We developed an application software called IGSA that leverages a powerful analytical capacity in gene sets enrichment and samples clustering. IGSA calculates gene sets expression scores for each sample and takes an accumulating clustering strategy to let the samples gather into the set according to the progress of disease from mild to severe. We focus on gastric, pancreatic and ovarian cancer data sets for the performance of IGSA. We also compared the results of IGSA in KEGG pathways enrichment with David, GSEA, SPIA, ssGSEA and analyzed the results of IGSA clustering and different similarity measurement methods. Notably, IGSA is proved to be more sensitive and specific in finding significant pathways, and can indicate related changes in pathways with the severity of disease. In addition, IGSA provides with significant gene sets profile for each sample. PMID:27764138

  19. Characterization of the temperate bacteriophage phi adh and plasmid transduction in Lactobacillus acidophilus ADH.

    PubMed

    Raya, R R; Kleeman, E G; Luchansky, J B; Klaenhammer, T R

    1989-09-01

    Lactobacillus acidophilus ADH is lysogenic and harbors an inducible prophage, phi adh. Bacteriophage were detected in cell lysates induced by treatment with mitomycin C or UV light. Electron microscopy of lysates revealed phage particles with a hexagonal head (62 nm) and a long, noncontractile, flexible tail (398 nm) ending in at last five short fibers. Phage phi adh was classified within Bradley's B1 phage group and the Siphoviridae family. The phi adh genome is a linear double-stranded DNA molecule of 41.7 kilobase pairs with cohesive ends: a physical map of the phi adh genome was constructed. A prophage-cured derivative of strain ADH, designated NCK102, was isolated from cells that survived UV exposure. NCK102 did not exhibit mitomycin C-induced lysis, but broth cultures lysed upon addition of phage. Phage phi adh produced clear plaques on NCK102 in media containing 10 mM CaCl2 at pH values between 5.2 and 5.5. A relysogenized derivative (NCK103) of NCK102 was isolated that exhibited mitomycin C-induced lysis and superinfection immunity to phage phi adh. Hybridization experiments showed that the phi adh genome was present in the ADH and NCK103 chromosomes, but absent in NCK102. These results demonstrated classic lytic and lysogenic cycles of replication for the temperate phage phi adh induced from L. acidophilus ADH. Phage phi adh also mediates transduction of plasmid DNA. Transductants of strain ADH containing pC194, pGK12, pGB354, and pVA797 were detected at frequencies in the range of 3.6 x 10(-8) to 8.3 x 10(-10) per PFU. Rearrangements or deletions were not detected in these plasmids as a consequence of transduction. This is the first description of plasmid transduction in the genus Lactobacillus.

  20. The use of gene clusters to infer functional coupling.

    SciTech Connect

    Overbeek, R.; Fonstein, M.; D'Souza, M.; Pusch, G. D.; Mathematics and Computer Science; Integrated Genomics; Univ. of Chicago

    1999-03-01

    Previously, we presented evidence that it is possible to predict functional coupling between genes based on conservation of gene clusters between genomes. With the rapid increase in the availability of prokaryotic sequence data, it has become possible to verify and apply the technique. In this paper, we extend our characterization of the parameters that determine the utility of the approach, and we generalize the approach in a way that supports detection of common classes of functionally coupled genes (e.g., transport and signal transduction clusters). Now that the analysis includes over 30 complete or nearly complete genomes, it has become clear that this approach will play a significant role in supporting efforts to assign functionality to the remaining uncharacterized genes in sequenced genomes.

  1. The Use of Gene Clusters to Infer Functional Coupling

    NASA Astrophysics Data System (ADS)

    Overbeek, Ross; Fonstein, Michael; D'Souza, Mark; Pusch, Gordon D.; Maltsev, Natalia

    1999-03-01

    Previously, we presented evidence that it is possible to predict functional coupling between genes based on conservation of gene clusters between genomes. With the rapid increase in the availability of prokaryotic sequence data, it has become possible to verify and apply the technique. In this paper, we extend our characterization of the parameters that determine the utility of the approach, and we generalize the approach in a way that supports detection of common classes of functionally coupled genes (e.g., transport and signal transduction clusters). Now that the analysis includes over 30 complete or nearly complete genomes, it has become clear that this approach will play a significant role in supporting efforts to assign functionality to the remaining uncharacterized genes in sequenced genomes.

  2. ADH and ALDH polymorphisms and alcohol dependence in Mexican and Native Americans

    PubMed Central

    Ehlers, Cindy L.; Liang, Tiebing; Gizer, Ian R.

    2012-01-01

    Background Ethanol is primarily metabolized in the liver by 2 rate-limiting reactions: conversion of ethanol to acetaldehyde by alcohol dehydrogenase (ADH) and subsequent conversion of acetaldehyde to acetate by aldehyde dehydrogenase (ALDH). ADH and ALDH exist in multiple isozymes that differ in their kinetic properties. Notably, polymorphisms within the genes that encode for these isozymes vary in their allele frequencies between ethnic groups, and thus, they have been considered as candidate genes that may differentially influence risk for the development of alcohol dependence across ethnic groups. Objectives and Methods Associations between alcohol dependence and polymorphisms in ADH1B, ADH1C, and ALDH2, were compared in a community sample of Native Americans living on reservations (n=791) and Mexican Americans (n=391) living within the same county. Results Two Mexican Americans and no Native Americans possessed one ALDH2*2 allele. Presence of at least one ADH1B*2 allele was found in 7% of the Native Americans and 13% of the Mexican Americans, but was only associated with protection against alcohol dependence in the Mexican Americans. Presence of at least one ADH1B*3 allele was found in 4% if the Native Americans and 2% of the Mexican Americans, but was associated with protection against alcohol dependence only in the Native Americans. No associations between alcohol dependence and polymorphisms in ADH1C were found. Conclusions and Scientific Significance Polymorphisms in ADH1B are protective against alcoholism in these two populations; however, these findings do not explain the high prevalence of alcoholism in these populations. PMID:22931071

  3. ADH and ALDH polymorphisms and alcohol dependence in Mexican and Native Americans

    PubMed Central

    Ehlers, Cindy L.; Liang, Tiebing; Gizer, Ian R.

    2012-01-01

    Background Ethanol is primarily metabolized in the liver by 2 rate-limiting reactions: conversion of ethanol to acetaldehyde by alcohol dehydrogenase (ADH) and subsequent conversion of acetaldehyde to acetate by aldehyde dehydrogenase (ALDH). ADH and ALDH exist in multiple isozymes that differ in their kinetic properties. Notably, polymorphisms within the genes that encode for these isozymes vary in their allele frequencies between ethnic groups, and thus, they have been considered as candidate genes that may differentially influence risk for the development of alcohol dependence across ethnic groups. Objectives and Methods Associations between alcohol dependence and polymorphisms in ADH1B, ADH1C, and ALDH2, were compared in a community sample of Native Americans living on reservations (n=791) and Mexican Americans (n=391) living within the same county. Results Two Mexican Americans and no Native Americans possessed one ALDH2*2 allele. Presence of at least one ADH1B*2 allele was found in 7% of the Native Americans and 13% of the Mexican Americans, but was only associated with protection against alcohol dependence in the Mexican Americans. Presence of at least one ADH1B*3 allele was found in 4% if the Native Americans and 2% of the Mexican Americans, but was associated with protection against alcohol dependence only in the Native Americans. No associations between alcohol dependence and polymorphisms in ADH1C were found. Conclusions and Scientific Significance Polymorphisms in ADH1B are protective against alcoholism in these two populations; however, these findings do not explain the high prevalence of alcoholism in these populations. PMID:22931071

  4. The multifunctional isopropyl alcohol dehydrogenase of Phytomonas sp. could be the result of a horizontal gene transfer from a bacterium to the trypanosomatid lineage.

    PubMed

    Molinas, Sara M; Altabe, Silvia G; Opperdoes, Fred R; Rider, Mark H; Michels, Paul A M; Uttaro, Antonio D

    2003-09-19

    Isopropyl alcohol dehydrogenase (iPDH) is a dimeric mitochondrial alcohol dehydrogenase (ADH), so far detected within the Trypanosomatidae only in the genus Phytomonas. The cloning, sequencing, and heterologous expression of the two gene alleles of the enzyme revealed that it is a zinc-dependent medium-chain ADH. Both polypeptides have 361 amino acids. A mitochondrial targeting sequence was identified. The mature proteins each have 348 amino acids and a calculated molecular mass of 37 kDa. They differ only in one amino acid, which can explain the three isoenzymes and their respective isoelectric points previously found. A phylogenetic analysis locates iPDH within a cluster with fermentative ADHs from bacteria, sharing 74% similarity and 60% identity with Ralstonia eutropha ADH. The characterization of the two bacterially expressed Phytomonas enzymes and the comparison of their kinetic properties with those of the wild-type iPDH and of the R. eutropha ADH strongly support the idea of a horizontal gene transfer event from a bacterium to a trypanosomatid to explain the origin of the iPDH in Phytomonas. Phytomonas iPDH and R. eutropha ADH are able to use a wide range of substrates with similar Km values such as primary and secondary alcohols, diols, and aldehydes, as well as ketones such as acetone, diacetyl, and acetoin. We speculate that, as for R. eutropha ADH, Phytomonas iPDH acts as a safety valve for the release of excess reducing power. PMID:12853449

  5. Quantitative Methylation Analysis of the PCDHB Gene Cluster.

    PubMed

    Banelli, Barbara; Romani, Massimo

    2015-01-01

    Long Range Epigenetic Silencing (LRES) is a repressed chromatin state of large chromosomal regions caused by DNA hypermethylation and histone modifications and is commonly observed in cancer. At 5q31 a LRES region of 800 kb includes three multi-gene clusters (PCDHA@, PCDHB@, and PCDHG@, respectively). Multiple experimental evidences have led to consider the PCDHB cluster as a DNA methylation marker of aggressiveness in neuroblastoma, second most common solid tumor in childhood. Because of its potential involvement not only in neuroblastoma but also in other malignancies, an easy and fast assay to screen the DNA methylation content of the PCDHB cluster might be useful for the precise stratification of the patients into risk groups and hence for choosing the most appropriate therapeutic protocol. Accordingly, we have developed a simple and cost-effective Pyrosequencing(®) assay to evaluate the methylation level of 17 genes in the protocadherin B cluster (PCDHB@). The rationale behind this Pyrosequencing assay can in principle be applied to analyze the DNA methylation level of any gene cluster with high homologies for screening purposes. PMID:26103900

  6. The Fusarium graminearum Genome Reveals More Secondary Metabolite Gene Clusters and Hints of Horizontal Gene Transfer

    PubMed Central

    Wong, Philip; Münsterkötter, Martin; Mewes, Hans-Werner; Schmeitzl, Clemens; Varga, Elisabeth; Berthiller, Franz; Adam, Gerhard; Güldener, Ulrich

    2014-01-01

    Fungal secondary metabolite biosynthesis genes are of major interest due to the pharmacological properties of their products (like mycotoxins and antibiotics). The genome of the plant pathogenic fungus Fusarium graminearum codes for a large number of candidate enzymes involved in secondary metabolite biosynthesis. However, the chemical nature of most enzymatic products of proteins encoded by putative secondary metabolism biosynthetic genes is largely unknown. Based on our analysis we present 67 gene clusters with significant enrichment of predicted secondary metabolism related enzymatic functions. 20 gene clusters with unknown metabolites exhibit strong gene expression correlation in planta and presumably play a role in virulence. Furthermore, the identification of conserved and over-represented putative transcription factor binding sites serves as additional evidence for cluster co-regulation. Orthologous cluster search provided insight into the evolution of secondary metabolism clusters. Some clusters are characteristic for the Fusarium phylum while others show evidence of horizontal gene transfer as orthologs can be found in representatives of the Botrytis or Cochliobolus lineage. The presented candidate clusters provide valuable targets for experimental examination. PMID:25333987

  7. Cloning and Heterologous Expression of the Grecocycline Biosynthetic Gene Cluster

    PubMed Central

    Bilyk, Oksana; Sekurova, Olga N.; Zotchev, Sergey B.; Luzhetskyy, Andriy

    2016-01-01

    Transformation-associated recombination (TAR) in yeast is a rapid and inexpensive method for cloning and assembly of large DNA fragments, which relies on natural homologous recombination. Two vectors, based on p15a and F-factor replicons that can be maintained in yeast, E. coli and streptomycetes have been constructed. These vectors have been successfully employed for assembly of the grecocycline biosynthetic gene cluster from Streptomyces sp. Acta 1362. Fragments of the cluster were obtained by PCR and transformed together with the “capture” vector into the yeast cells, yielding a construct carrying the entire gene cluster. The obtained construct was heterologously expressed in S. albus J1074, yielding several grecocycline congeners. Grecocyclines have unique structural moieties such as a dissacharide side chain, an additional amino sugar at the C-5 position and a thiol group. Enzymes from this pathway may be used for the derivatization of known active angucyclines in order to improve their desired biological properties. PMID:27410036

  8. Cloning and Heterologous Expression of the Grecocycline Biosynthetic Gene Cluster.

    PubMed

    Bilyk, Oksana; Sekurova, Olga N; Zotchev, Sergey B; Luzhetskyy, Andriy

    2016-01-01

    Transformation-associated recombination (TAR) in yeast is a rapid and inexpensive method for cloning and assembly of large DNA fragments, which relies on natural homologous recombination. Two vectors, based on p15a and F-factor replicons that can be maintained in yeast, E. coli and streptomycetes have been constructed. These vectors have been successfully employed for assembly of the grecocycline biosynthetic gene cluster from Streptomyces sp. Acta 1362. Fragments of the cluster were obtained by PCR and transformed together with the "capture" vector into the yeast cells, yielding a construct carrying the entire gene cluster. The obtained construct was heterologously expressed in S. albus J1074, yielding several grecocycline congeners. Grecocyclines have unique structural moieties such as a dissacharide side chain, an additional amino sugar at the C-5 position and a thiol group. Enzymes from this pathway may be used for the derivatization of known active angucyclines in order to improve their desired biological properties. PMID:27410036

  9. Duplications of hox gene clusters and the emergence of vertebrates.

    PubMed

    Soshnikova, Natalia; Dewaele, Romain; Janvier, Philippe; Krumlauf, Robb; Duboule, Denis

    2013-06-15

    The vertebrate body plan is characterized by an increased complexity relative to that of all other chordates and large-scale gene amplifications have been associated with key morphological innovations leading to their remarkable evolutionary success. Here, we use compound full Hox clusters deletions to investigate how Hox genes duplications may have contributed to the emergence of vertebrate-specific innovations. We show that the combined deletion of HoxA and HoxB leads to an atavistic heart phenotype, suggesting that the ancestral HoxA/B cluster was co-opted to help in diversifying the complex organ in vertebrates. Other phenotypic effects observed seem to illustrate the resurgence of ancestral (plesiomorphic) features. This indicates that the duplications of Hox clusters were associated with the recruitment or formation of novel cis-regulatory controls, which were key to the evolution of many vertebrate features and hence to the evolutionary radiation of this group.

  10. From Green to Red: Horizontal Gene Transfer of the Phycoerythrin Gene Cluster between Planktothrix Strains

    PubMed Central

    Sogge, Hanne; Rounge, Trine Ballestad; Nederbragt, Alexander J.; Lagesen, Karin; Glöckner, Gernot; Hayes, Paul K.; Rohrlack, Thomas

    2013-01-01

    Horizontal gene transfer is common in cyanobacteria, and transfer of large gene clusters may lead to acquisition of new functions and conceivably niche adaption. In the present study, we demonstrate that horizontal gene transfer between closely related Planktothrix strains can explain the production of the same oligopeptide isoforms by strains of different colors. Comparison of the genomes of eight Planktothrix strains revealed that strains producing the same oligopeptide isoforms are closely related, regardless of color. We have investigated genes involved in the synthesis of the photosynthetic pigments phycocyanin and phycoerythrin, which are responsible for green and red appearance, respectively. Sequence comparisons suggest the transfer of a functional phycoerythrin gene cluster generating a red phenotype in a strain that is otherwise more closely related to green strains. Our data show that the insertion of a DNA fragment containing the 19.7-kb phycoerythrin gene cluster has been facilitated by homologous recombination, also replacing a region of the phycocyanin operon. These findings demonstrate that large DNA fragments spanning entire functional gene clusters can be effectively transferred between closely related cyanobacterial strains and result in a changed phenotype. Further, the results shed new light on the discussion of the role of horizontal gene transfer in the sporadic distribution of large gene clusters in cyanobacteria, as well as the appearance of red and green strains. PMID:23995927

  11. Comparative Genomics of Natural Killer Cell Receptor Gene Clusters

    PubMed Central

    Kelley, James; Walter, Lutz; Trowsdale, John

    2005-01-01

    Many receptors on natural killer (NK) cells recognize major histocompatibility complex class I molecules in order to monitor unhealthy tissues, such as cells infected with viruses, and some tumors. Genes encoding families of NK receptors and related sequences are organized into two main clusters in humans: the natural killer complex on Chromosome 12p13.1, which encodes C-type lectin molecules, and the leukocyte receptor complex on Chromosome 19q13.4, which encodes immunoglobulin superfamily molecules. The composition of these gene clusters differs markedly between closely related species, providing evidence for rapid, lineage-specific expansions or contractions of sets of loci. The choice of NK receptor genes is polarized in the two species most studied, mouse and human. In mouse, the C-type lectin-related Ly49 gene family predominates. Conversely, the single Ly49 sequence is a pseudogene in humans, and the immunoglobulin superfamily KIR gene family is extensive. These different gene sets encode proteins that are comparable in function and genetic diversity, even though they have undergone species-specific expansions. Understanding the biological significance of this curious situation may be aided by studying which NK receptor genes are used in other vertebrates, especially in relation to species-specific differences in genes for major histocompatibility complex class I molecules. PMID:16132082

  12. The evolution of small gene clusters: evidence for an independent origin of the maltase gene cluster in Drosophila virilis and Drosophila melanogaster.

    PubMed

    Vieira, C P; Vieira, J; Hartl, D L

    1997-10-01

    We analyzed a 5,770-bp genomic region of Drosophila virilis that contains a cluster of two maltase genes showing sequence similarity with genes in a cluster of three maltase genes previously identified in Drosophila melanogaster. The D. virilis maltase genes are designated Mav1 and Mav2. In addition to being different in gene number, the cluster of genes in D. virilis differs dramatically in intron-exon structure from the maltase genes in D. melanogaster, the transcriptional orientation of the genes in the cluster also differs between the species. Our findings support a model in which the maltase gene cluster in D. virilis and D. melanogaster evolved independently. Furthermore, while in D. melanogaster the maltase gene cluster lies only 10 kb distant from the larval cuticle gene cluster, the maltase and larval cuticle gene clusters in D. virilis are located very far apart and on a different chromosome than that expected from the known chromosome arm homologies between D. virilis and D. melanogaster. A region of the genome containing the maltase and larval cuticle gene clusters appears to have been relocated between nonhomologous chromosomes.

  13. An alanine tRNA gene cluster from Nephila clavipes.

    PubMed

    Luciano, E; Candelas, G C

    1996-06-01

    We report the sequence of a 2.3-kb genomic DNA fragment from the orb-web spider, Nephila clavipes (Nc). The fragment contains four regions of high homology to tRNA(Ala). The members of this irregularly spaced cluster of genes are oriented in the same direction and have the same anticodon (GCA), but their sequence differs at several positions. Initiation and termination signals, as well as consensus intragenic promoter sequences characteristic of tRNA genes, have been identified in all genes. tRNA(Ala) are involved in the regulation of the fibroin synthesis in the large ampullate Nc glands.

  14. Evolutionary conservation of regulatory elements in vertebrate HOX gene clusters

    SciTech Connect

    Santini, Simona; Boore, Jeffrey L.; Meyer, Axel

    2003-12-31

    Due to their high degree of conservation, comparisons of DNA sequences among evolutionarily distantly-related genomes permit to identify functional regions in noncoding DNA. Hox genes are optimal candidate sequences for comparative genome analyses, because they are extremely conserved in vertebrates and occur in clusters. We aligned (Pipmaker) the nucleotide sequences of HoxA clusters of tilapia, pufferfish, striped bass, zebrafish, horn shark, human and mouse (over 500 million years of evolutionary distance). We identified several highly conserved intergenic sequences, likely to be important in gene regulation. Only a few of these putative regulatory elements have been previously described as being involved in the regulation of Hox genes, while several others are new elements that might have regulatory functions. The majority of these newly identified putative regulatory elements contain short fragments that are almost completely conserved and are identical to known binding sites for regulatory proteins (Transfac). The conserved intergenic regions located between the most rostrally expressed genes in the developing embryo are longer and better retained through evolution. We document that presumed regulatory sequences are retained differentially in either A or A clusters resulting from a genome duplication in the fish lineage. This observation supports both the hypothesis that the conserved elements are involved in gene regulation and the Duplication-Deletion-Complementation model.

  15. Coupled two-way clustering analysis of gene microarray data

    NASA Astrophysics Data System (ADS)

    Getz, Gad; Levine, Erel; Domany, Eytan

    2000-10-01

    We present a coupled two-way clustering approach to gene microarray data analysis. The main idea is to identify subsets of the genes and samples, such that when one of these is used to cluster the other, stable and significant partitions emerge. The search for such subsets is a computationally complex task. We present an algorithm, based on iterative clustering, that performs such a search. This analysis is especially suitable for gene microarray data, where the contributions of a variety of biological mechanisms to the gene expression levels are entangled in a large body of experimental data. The method was applied to two gene microarray data sets, on colon cancer and leukemia. By identifying relevant subsets of the data and focusing on them we were able to discover partitions and correlations that were masked and hidden when the full dataset was used in the analysis. Some of these partitions have clear biological interpretation; others can serve to identify possible directions for future research.

  16. Characterization of alcohol dehydrogenase (ADH12) from Haloarcula marismortui, an extreme halophile from the Dead Sea.

    PubMed

    Timpson, Leanne M; Alsafadi, Diya; Mac Donnchadha, Cillín; Liddell, Susan; Sharkey, Michael A; Paradisi, Francesca

    2012-01-01

    Haloarchaeal alcohol dehydrogenases are of increasing interest as biocatalysts in the field of white biotechnology. In this study, the gene adh12 from the extreme halophile Haloarcula marismortui (HmADH12), encoding a 384 residue protein, was cloned into two vectors: pRV1 and pTA963. The resulting constructs were used to transform host strains Haloferax volcanii (DS70) and (H1209), respectively. Overexpressed His-tagged recombinant HmADH12 was purified by immobilized metal-affinity chromatography (IMAC). The His-tagged protein was visualized by SDS-PAGE, with a subunit molecular mass of 41.6 kDa, and its identity was confirmed by mass spectrometry. Purified HmADH12 catalyzed the interconversion between alcohols and aldehydes and ketones, being optimally active in the presence of 2 M KCl. It was thermoactive, with maximum activity registered at 60°C. The NADP(H) dependent enzyme was haloalkaliphilic for the oxidative reaction with optimum activity at pH 10.0. It favored a slightly acidic pH of 6.0 for catalysis of the reductive reaction. HmADH12 was significantly more tolerant than mesophilic ADHs to selected organic solvents, making it a much more suitable biocatalyst for industrial application.

  17. The Adh adhesin domain is required for trimeric autotransporter Apa1-mediated Actinobacillus pleuropneumoniae adhesion, autoaggregation, biofilm formation and pathogenicity.

    PubMed

    Wang, Lei; Qin, Wanhai; Yang, Shuxin; Zhai, Ruidong; Zhou, Liang; Sun, Changjiang; Pan, Fengguang; Ji, Qun; Wang, Yu; Gu, Jingmin; Feng, Xin; Du, Chongtao; Han, Wenyu; Langford, P R; Lei, Liancheng

    2015-05-15

    Actinobacillus pleuropneumoniae is a causative agent of porcine pleuropneumonia, which is a highly contagious endemic disease of pigs. Adhesion is a critical first step in the infection process. Trimeric autotransporter adhesions (TAAs) have been identified as novel virulence factors; however, little is known on their roles in A. pleuropneumoniae pathogenicity. Here, our data show that YadA-like head region (Adh) of Apa1 was the optimal adhesion functional domain via segment expression and adhesion assays in vitro. Additionally, Adh induced partial protection against A. pleuropneumoniae 5b L20 and serotypes 1, 3, and 5a in mice. The deletion of Adh gene significantly decreased autoaggregation, biofilm formation and adherence to host cells in vitro. Furthermore, with delaying of clinical symptoms, reducing production of pro-inflammatory cytokines and lessening the lung injury after infection, Adh deletion strain (5bϕAdh) significantly reduced the pathogenicity to piglets. To elucidate the mechanism of lung injury, the differentially expressed genes in the lung tissues of piglets infected with the 5b L20 or 5bϕAdh strains were investigated using microarray analysis and validated by qRT-PCR. Compared with the 5b L20 infected piglets, 495 genes were differentially expressed in 5bϕAdh infected lung tissue (221 upregulated and 274 downregulated). Especially, the antigen processing and presentation gene IFI30 was increased following infection with the 5bϕAdh strain. Thus, Adh may enhance pathogenicity by depressing host immune recognition. We conclude that the head domain of the A. pleuropneumoniae trimeric autotransporter Apa1 regulates autoagglutination, biofilm formation, adhesion to host cells and pathogenicity.

  18. High diversity of polyketide synthase genes and the melanin biosynthesis gene cluster in Penicillium marneffei.

    PubMed

    Woo, Patrick C Y; Tam, Emily W T; Chong, Ken T K; Cai, James J; Tung, Edward T K; Ngan, Antonio H Y; Lau, Susanna K P; Yuen, Kwok-Yung

    2010-09-01

    Despite the unique phenotypic properties and clinical importance of Penicillium marneffei, the polyketide synthase genes in its genome have never been characterized. Twenty-three putative polyketide synthase genes and two putative polyketide synthase nonribosomal peptide-synthase hybrid genes were identified in the P. marneffei genome, a diversity much higher than found in other pathogenic thermal dimorphic fungi, such as Histoplasma capsulatum (one polyketide synthase gene) and Coccidioides immitis (10 polyketide synthase genes). These genes were evenly distributed on the phylogenetic tree with polyketide synthase genes of Aspergillus and other fungi, indicating that the high diversity was not a result of lineage-specific gene expansion through recent gene duplication. The melanin-biosynthesis gene cluster had gene order and orientations identical to those in the Talaromyces stipitatus (a teleomorph of Penicillium emmonsii) genome. Phylogenetically, all six genes of the melanin-biosynthesis gene cluster in P. marneffei were also most closely related to those in T. stipitatus, with high bootstrap supports. The polyketide synthase gene of the melanin-biosynthesis gene cluster (alb1) in P. marneffei was knocked down, which was accompanied by loss of melanin pigment production and reduced ornamentation in conidia. The survival of mice challenged with the alb1 knockdown mutant was significantly better than those challenged with wild-type P. marneffei (P < 0.005). The sterilizing doses of hydrogen peroxide, leading to a 50% reduction in survival of conidia, were 11 min for wild-type P. marneffei and 6 min for the alb1 knockdown mutant of P. marneffei, implying that the melanin-biosynthesis gene cluster contributed to virulence through decreased susceptibility to killing by hydrogen peroxide. PMID:20718860

  19. Evolution of chemical diversity by coordinated gene swaps in type II polyketide gene clusters

    PubMed Central

    Hillenmeyer, Maureen E.; Vandova, Gergana A.; Berlew, Erin E.; Charkoudian, Louise K.

    2015-01-01

    Natural product biosynthetic pathways generate molecules of enormous structural complexity and exquisitely tuned biological activities. Studies of natural products have led to the discovery of many pharmaceutical agents, particularly antibiotics. Attempts to harness the catalytic prowess of biosynthetic enzyme systems, for both compound discovery and engineering, have been limited by a poor understanding of the evolution of the underlying gene clusters. We developed an approach to study the evolution of biosynthetic genes on a cluster-wide scale, integrating pairwise gene coevolution information with large-scale phylogenetic analysis. We used this method to infer the evolution of type II polyketide gene clusters, tracing the path of evolution from the single ancestor to those gene clusters surviving today. We identified 10 key gene types in these clusters, most of which were swapped in from existing cellular processes and subsequently specialized. The ancestral type II polyketide gene cluster likely comprised a core set of five genes, a roster that expanded and contracted throughout evolution. A key C24 ancestor diversified into major classes of longer and shorter chain length systems, from which a C20 ancestor gave rise to the majority of characterized type II polyketide antibiotics. Our findings reveal that (i) type II polyketide structure is predictable from its gene roster, (ii) only certain gene combinations are compatible, and (iii) gene swaps were likely a key to evolution of chemical diversity. The lessons learned about how natural selection drives polyketide chemical innovation can be applied to the rational design and guided discovery of chemicals with desired structures and properties. PMID:26499248

  20. Fetal ADH2*3, maternal alcohol consumption, and fetal growth.

    PubMed

    Arfsten, Darryl P; Silbergeld, Ellen K; Loffredo, Christopher A

    2004-01-01

    There is some evidence suggesting the allele for alcohol dehydrogenase 2*3 (ADH2*3) is associated with a protective effect against alcohol-related intrauterine growth retardation (IUGR). This study was conducted to explore the affect of the ADH2*3 allele on fetal growth. Bloodspots (n = 1016) belonging to individual infants of a subgroup of the Baltimore-Washington Infant Study (BWIS) were assayed for the presence of the ADH2*3 allele by a polymerase chain reaction (PCR)-based method. Infants genotyped for ADH2*3 were those for whom bloodspots were identified and obtained from the Maryland Newborn Screening Program. The effect of ADH2*3 and maternal alcohol consumption on intrauterine growth was explored by multivariable linear regression analysis. Twenty-six percent of the 306 blood spots belonging to African-American infants were positive for ADH2*3 (4% were homozygous and 22% were heterozygous). Only a small percentage of bloodspots for Caucasian (1.3%) were positive for the ADH2*3 allele. Consequently, further analysis concentrated on gene-exposure interactions for African-American infants. It was found that the incidence of being small-for-gestation-age (SGA) was lower for ADH2*3-positive infants (2.5% versus 8.8%; p = .08). SGA infants had elevated odds for being ADH2*3 negative (OR: 3.15, 95% C.I.: 0.70-14.26) and for being born to mothers that consumed alcohol during pregnancy (OR: 2.31, 95% C.I.: 0.77-6.91). A negative trend between maternal alcohol consumption and mean offspring birthweight was found; however, ADH2*3 did not have a significant impact on mean birthweight for infants born to mothers that drank during pregnancy. These results could be interpreted as possible support for the hypothesis that ADH2 genotype in the infant may impact risk for alcohol-related IUGR. However, this study has limitations in that it is a "nested study of convenience" and involves a relatively small number of infants born to mothers reporting moderate to heavy alcohol

  1. Cluster of genes controlling proline degradation in Salmonella typhimurium.

    PubMed Central

    Ratzkin, B; Roth, J

    1978-01-01

    A cluster of genes essential for degradation of proline to glutamate (put) is located between the pyrC and pyrD loci at min 22 of the Salmonella chromosome. A series of 25 deletion mutants of this region have been isolated and used to construct a fine-structure map of the put genes. The map includes mutations affecting the proline degradative activities, proline oxidase and pyrroline-5-carboxylic dehydrogenase. Also included are mutations affecting the major proline permease and a regulatory mutation that affects both enzyme and permease production. The two enzymatic activities appear to be encoded by a single gene (putA). The regulatory mutation maps between the putA gene and the proline permease gene (putP). PMID:342507

  2. Identification of genes and gene clusters involved in mycotoxin synthesis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Research methods to identify and characterize genes involved in mycotoxin biosynthetic pathways have evolved considerably over the years. Before whole genome sequences were available (e.g. pre-genomics), work focused primarily on chemistry, biosynthetic mutant strains and molecular analysis of sing...

  3. Transcription mediated insulation and interference direct gene cluster expression switches.

    PubMed

    Nguyen, Tania; Fischl, Harry; Howe, Françoise S; Woloszczuk, Ronja; Serra Barros, Ana; Xu, Zhenyu; Brown, David; Murray, Struan C; Haenni, Simon; Halstead, James M; O'Connor, Leigh; Shipkovenska, Gergana; Steinmetz, Lars M; Mellor, Jane

    2014-11-19

    In yeast, many tandemly arranged genes show peak expression in different phases of the metabolic cycle (YMC) or in different carbon sources, indicative of regulation by a bi-modal switch, but it is not clear how these switches are controlled. Using native elongating transcript analysis (NET-seq), we show that transcription itself is a component of bi-modal switches, facilitating reciprocal expression in gene clusters. HMS2, encoding a growth-regulated transcription factor, switches between sense- or antisense-dominant states that also coordinate up- and down-regulation of transcription at neighbouring genes. Engineering HMS2 reveals alternative mono-, di- or tri-cistronic and antisense transcription units (TUs), using different promoter and terminator combinations, that underlie state-switching. Promoters or terminators are excluded from functional TUs by read-through transcriptional interference, while antisense TUs insulate downstream genes from interference. We propose that the balance of transcriptional insulation and interference at gene clusters facilitates gene expression switches during intracellular and extracellular environmental change.

  4. Transcription mediated insulation and interference direct gene cluster expression switches

    PubMed Central

    Nguyen, Tania; Brown, David; Murray, Struan C; Haenni, Simon; Halstead, James M; O'Connor, Leigh; Shipkovenska, Gergana; Steinmetz, Lars M; Mellor, Jane

    2014-01-01

    In yeast, many tandemly arranged genes show peak expression in different phases of the metabolic cycle (YMC) or in different carbon sources, indicative of regulation by a bi-modal switch, but it is not clear how these switches are controlled. Using native elongating transcript analysis (NET-seq), we show that transcription itself is a component of bi-modal switches, facilitating reciprocal expression in gene clusters. HMS2, encoding a growth-regulated transcription factor, switches between sense- or antisense-dominant states that also coordinate up- and down-regulation of transcription at neighbouring genes. Engineering HMS2 reveals alternative mono-, di- or tri-cistronic and antisense transcription units (TUs), using different promoter and terminator combinations, that underlie state-switching. Promoters or terminators are excluded from functional TUs by read-through transcriptional interference, while antisense TUs insulate downstream genes from interference. We propose that the balance of transcriptional insulation and interference at gene clusters facilitates gene expression switches during intracellular and extracellular environmental change. DOI: http://dx.doi.org/10.7554/eLife.03635.001 PMID:25407679

  5. The Relationship between CmADHs and the Diversity of Volatile Organic Compounds of Three Aroma Types of Melon (Cucumis melo)

    PubMed Central

    Chen, Hao; Cao, Songxiao; Jin, Yazhong; Tang, Yufan; Qi, Hongyan

    2016-01-01

    Alcohol dehydrogenase (ADH) plays an important role in aroma volatile compounds synthesis of plants. In this paper, we tried to explore the relationship between CmADHs and the volatile organic compounds (VOCs) in oriental melon. Three different aroma types of melon were used as materials. The principle component analysis of three types of melon fruit was conducted. We also measured the CmADHs expression level and enzymatic activities of ADH and alcohol acyl-transferase (AAT) on different stages of fruit ripening. An incubation experiment was carried out to investigate the effect of substrates and inhibitor (4-MP, 4-methylpyrazole) on CmADHs expression, ADH activity, and the main compounds of oriental melon. The results illustrated that ethyl acetate, hexyl acetate (E,Z)-3,6-nonadien-1-ol and 2-ethyl-2hexen-1-ol were the four principal volatile compounds of these three types of melon. AAT activity was increasing with fruit ripening, and the AAT activity in CH were the highest, whereas ADH activity peaked on 32 DAP, 2 days before maturation, and the ADH activity in CB and CG were higher than that in CH. The expression pattern of 11 CmADH genes from 24 to 36 day after pollination (DAP) was found to vary in three melon varieties. CmADH4 was only expressed in CG and the expression levels of CmADH3 and CmADH12 in CH and CB were much higher than that in CG, and they both peaked 2 days before fruit ripening. Ethanol and 4-MP decreased the reductase activity of ADH, the expression of most CmADHs and ethyl acetate or hexyl acetate contents of CB, except for 0.1 mM 4-MP, while aldehyde improved the two acetate ester contents. In addition, we found a positive correlation between the expression of CmADH3 and CmADH12 and the key volatile compound of CB. The relationship between CmADHs and VOCs synthesis of oriental melon was discussed. PMID:27445845

  6. The Relationship between CmADHs and the Diversity of Volatile Organic Compounds of Three Aroma Types of Melon (Cucumis melo).

    PubMed

    Chen, Hao; Cao, Songxiao; Jin, Yazhong; Tang, Yufan; Qi, Hongyan

    2016-01-01

    Alcohol dehydrogenase (ADH) plays an important role in aroma volatile compounds synthesis of plants. In this paper, we tried to explore the relationship between CmADHs and the volatile organic compounds (VOCs) in oriental melon. Three different aroma types of melon were used as materials. The principle component analysis of three types of melon fruit was conducted. We also measured the CmADHs expression level and enzymatic activities of ADH and alcohol acyl-transferase (AAT) on different stages of fruit ripening. An incubation experiment was carried out to investigate the effect of substrates and inhibitor (4-MP, 4-methylpyrazole) on CmADHs expression, ADH activity, and the main compounds of oriental melon. The results illustrated that ethyl acetate, hexyl acetate (E,Z)-3,6-nonadien-1-ol and 2-ethyl-2hexen-1-ol were the four principal volatile compounds of these three types of melon. AAT activity was increasing with fruit ripening, and the AAT activity in CH were the highest, whereas ADH activity peaked on 32 DAP, 2 days before maturation, and the ADH activity in CB and CG were higher than that in CH. The expression pattern of 11 CmADH genes from 24 to 36 day after pollination (DAP) was found to vary in three melon varieties. CmADH4 was only expressed in CG and the expression levels of CmADH3 and CmADH12 in CH and CB were much higher than that in CG, and they both peaked 2 days before fruit ripening. Ethanol and 4-MP decreased the reductase activity of ADH, the expression of most CmADHs and ethyl acetate or hexyl acetate contents of CB, except for 0.1 mM 4-MP, while aldehyde improved the two acetate ester contents. In addition, we found a positive correlation between the expression of CmADH3 and CmADH12 and the key volatile compound of CB. The relationship between CmADHs and VOCs synthesis of oriental melon was discussed. PMID:27445845

  7. The Relationship between CmADHs and the Diversity of Volatile Organic Compounds of Three Aroma Types of Melon (Cucumis melo).

    PubMed

    Chen, Hao; Cao, Songxiao; Jin, Yazhong; Tang, Yufan; Qi, Hongyan

    2016-01-01

    Alcohol dehydrogenase (ADH) plays an important role in aroma volatile compounds synthesis of plants. In this paper, we tried to explore the relationship between CmADHs and the volatile organic compounds (VOCs) in oriental melon. Three different aroma types of melon were used as materials. The principle component analysis of three types of melon fruit was conducted. We also measured the CmADHs expression level and enzymatic activities of ADH and alcohol acyl-transferase (AAT) on different stages of fruit ripening. An incubation experiment was carried out to investigate the effect of substrates and inhibitor (4-MP, 4-methylpyrazole) on CmADHs expression, ADH activity, and the main compounds of oriental melon. The results illustrated that ethyl acetate, hexyl acetate (E,Z)-3,6-nonadien-1-ol and 2-ethyl-2hexen-1-ol were the four principal volatile compounds of these three types of melon. AAT activity was increasing with fruit ripening, and the AAT activity in CH were the highest, whereas ADH activity peaked on 32 DAP, 2 days before maturation, and the ADH activity in CB and CG were higher than that in CH. The expression pattern of 11 CmADH genes from 24 to 36 day after pollination (DAP) was found to vary in three melon varieties. CmADH4 was only expressed in CG and the expression levels of CmADH3 and CmADH12 in CH and CB were much higher than that in CG, and they both peaked 2 days before fruit ripening. Ethanol and 4-MP decreased the reductase activity of ADH, the expression of most CmADHs and ethyl acetate or hexyl acetate contents of CB, except for 0.1 mM 4-MP, while aldehyde improved the two acetate ester contents. In addition, we found a positive correlation between the expression of CmADH3 and CmADH12 and the key volatile compound of CB. The relationship between CmADHs and VOCs synthesis of oriental melon was discussed.

  8. Reconstructing Histories of Complex Gene Clusters on a Phylogeny

    NASA Astrophysics Data System (ADS)

    Vinař, Tomáš; Brejová, Broňa; Song, Giltae; Siepel, Adam

    Clusters of genes that have evolved by repeated segmental duplication present difficult challenges throughout genomic analysis, from sequence assembly to functional analysis. These clusters are one of the major sources of evolutionary innovation, and they are linked to multiple diseases, including HIV and a variety of cancers. Understanding their evolutionary histories is a key to the application of comparative genomics methods in these regions of the genome. We propose a probabilistic model of gene cluster evolution on a phylogeny, and an MCMC algorithm for reconstruction of duplication histories from genomic sequences in multiple species. Several projects are underway to obtain high quality BAC-based assemblies of duplicated clusters in multiple species, and we anticipate use of our methods in their analysis. Supplementary materials are located at http://compbio.fmph.uniba.sk/suppl/09recombcg/

  9. Transcriptional analysis of exopolysaccharides biosynthesis gene clusters in Lactobacillus plantarum.

    PubMed

    Vastano, Valeria; Perrone, Filomena; Marasco, Rosangela; Sacco, Margherita; Muscariello, Lidia

    2016-04-01

    Exopolysaccharides (EPS) from lactic acid bacteria contribute to specific rheology and texture of fermented milk products and find applications also in non-dairy foods and in therapeutics. Recently, four clusters of genes (cps) associated with surface polysaccharide production have been identified in Lactobacillus plantarum WCFS1, a probiotic and food-associated lactobacillus. These clusters are involved in cell surface architecture and probably in release and/or exposure of immunomodulating bacterial molecules. Here we show a transcriptional analysis of these clusters. Indeed, RT-PCR experiments revealed that the cps loci are organized in five operons. Moreover, by reverse transcription-qPCR analysis performed on L. plantarum WCFS1 (wild type) and WCFS1-2 (ΔccpA), we demonstrated that expression of three cps clusters is under the control of the global regulator CcpA. These results, together with the identification of putative CcpA target sequences (catabolite responsive element CRE) in the regulatory region of four out of five transcriptional units, strongly suggest for the first time a role of the master regulator CcpA in EPS gene transcription among lactobacilli.

  10. Meta-Analyses of ALDH2 and ADH1B with Alcohol Dependence in Asians

    ERIC Educational Resources Information Center

    Luczak, Susan E.; Glatt, Stephen J.; Wall, Tamara J.

    2006-01-01

    Meta-analyses were conducted to determine the magnitude of relationships between polymorphisms in 2 genes, ALDH2 and ADH1B, with alcohol dependence in Asians. For each gene, possession of 1 variant [asterisk]2 allele was protective against alcohol dependence, and possession of a 2nd [asterisk]2 allele did not offer significant additional…

  11. Analysis of lamprey clustered Fox genes: insight into Fox gene evolution and expression in vertebrates.

    PubMed

    Wotton, Karl R; Shimeld, Sebastian M

    2011-12-01

    In the human genome, members of the FoxC, FoxF, FoxL1, and FoxQ1 gene families are found in two paralagous clusters. One cluster contains the genes FOXQ1, FOXF2, FOXC1 and the second consists of FOXF1, FOXC2, and FOXL1. In jawed vertebrates these genes are known to be expressed in different pharyngeal tissues and all, except FoxQ1, are involved in patterning the early embryonic mesoderm. We have previously traced the evolution of this cluster in the bony vertebrates, and the gene content is identical in the dogfish, a member of the most basally branching lineage of the jawed vertebrates. Here we extend these analyses to jawless vertebrates. Using genomic searches and molecular approaches we have identified homologues of these genes from lampreys. We identify two FoxC genes, two FoxF genes, two FoxQ1 genes and single FoxL1 gene. We examine the embryonic expression of one predominantly mesodermally expressed gene family, FoxC, and the endodermally expressed member of the cluster, FoxQ1. We identified FoxQ1 transcripts in the pharyngeal endoderm, while the two FoxC genes are differentially expressed in the pharyngeal mesenchyme and ectoderm. Furthermore we identify conserved expression of lamprey FoxC genes in the paraxial and intermediate mesoderms. We interpret our results through a chordate-wide comparison of expression patterns and discuss gene content in the context of theories on the evolution of the vertebrate genome.

  12. Metabolic diversification--independent assembly of operon-like gene clusters in different plants.

    PubMed

    Field, Ben; Osbourn, Anne E

    2008-04-25

    Operons are clusters of unrelated genes with related functions that are a feature of prokaryotic genomes. Here, we report on an operon-like gene cluster in the plant Arabidopsis thaliana that is required for triterpene synthesis (the thalianol pathway). The clustered genes are coexpressed, as in bacterial operons. However, despite the resemblance to a bacterial operon, this gene cluster has been assembled from plant genes by gene duplication, neofunctionalization, and genome reorganization, rather than by horizontal gene transfer from bacteria. Furthermore, recent assembly of operon-like gene clusters for triterpene synthesis has occurred independently in divergent plant lineages (Arabidopsis and oat). Thus, selection pressure may act during the formation of certain plant metabolic pathways to drive gene clustering.

  13. Metabolic diversification--independent assembly of operon-like gene clusters in different plants.

    PubMed

    Field, Ben; Osbourn, Anne E

    2008-04-25

    Operons are clusters of unrelated genes with related functions that are a feature of prokaryotic genomes. Here, we report on an operon-like gene cluster in the plant Arabidopsis thaliana that is required for triterpene synthesis (the thalianol pathway). The clustered genes are coexpressed, as in bacterial operons. However, despite the resemblance to a bacterial operon, this gene cluster has been assembled from plant genes by gene duplication, neofunctionalization, and genome reorganization, rather than by horizontal gene transfer from bacteria. Furthermore, recent assembly of operon-like gene clusters for triterpene synthesis has occurred independently in divergent plant lineages (Arabidopsis and oat). Thus, selection pressure may act during the formation of certain plant metabolic pathways to drive gene clustering. PMID:18356490

  14. Bacillus subtilis Gene Cluster Involved in Calcium Carbonate Biomineralization▿

    PubMed Central

    Barabesi, Chiara; Galizzi, Alessandro; Mastromei, Giorgio; Rossi, Mila; Tamburini, Elena; Perito, Brunella

    2007-01-01

    Calcium carbonate precipitation, a widespread phenomenon among bacteria, has been investigated due to its wide range of scientific and technological implications. Nevertheless, little is known of the molecular mechanisms by which bacteria foster calcium carbonate mineralization. In our laboratory, we are studying calcite formation by Bacillus subtilis, in order to identify genes involved in the biomineralization process. A previous screening of UV mutants and of more than one thousand mutants obtained from the European B. subtilis Functional Analysis project allowed us to isolate strains altered in the precipitation phenotype. Starting from these results, we focused our attention on a cluster of five genes (lcfA, ysiA, ysiB, etfB, and etfA) called the lcfA operon. By insertional mutagenesis, mutant strains carrying each of the five genes were produced. All of them, with the exception of the strain carrying the mutated lcfA operon, were unable to form calcite crystals. By placing transcription under IPTG (isopropyl-β-d-thiogalactopyranoside) control, the last gene, etfA, was identified as essential for the precipitation process. To verify cotranscription in the lcfA operon, reverse transcription-PCR experiments were performed and overlapping retrocotranscripts were found comprising three adjacent genes. The genes have putative functions linked to fatty acid metabolism. A link between calcium precipitation and fatty acid metabolism is suggested. PMID:17085570

  15. Using endophenotypes for pathway clusters to map complex disease genes.

    PubMed

    Pan, Wen-Harn; Lynn, Ke-Shiuan; Chen, Chun-Houh; Wu, Yi-Lin; Lin, Chung-Yen; Chang, Hsing-Yi

    2006-02-01

    Nature determines the complexity of disease etiology and the likelihood of revealing disease genes. While culprit genes for many monogenic diseases have been successfully unraveled, efforts to map major complex disease genes have not been as productive as hoped. The conceptual framework currently adopted to deal with the heterogeneous nature of complex diseases focuses on using homogeneous internal features of the disease phenotype for mapping. However, phenotypic homogeneity does not equal genotypic homogeneity. In this report, we advocate working with well-measured phenotypes portrayed by amounts of transcripts and activities of gene products or their metabolites, which are pertinent to relatively small pathway clusters. Reliable and controlled measures for oligogenic traits resulting from proper dissection efforts may enhance statistical power. The large amounts of information obtained on gene and protein expression from technological advances can add to the power of gene finding, particularly for diseases with unclear etiology. Data-mining tools for dimension reduction can assist biologists to reveal novel molecular endophenotypes. However, there are still hurdles to overcome, including high cost, relatively poor reproducibility and comparability among platforms, the cross-sectional nature of the information, and the accessibility of human tissues. Concerted efforts are required to carry out large-scale prospective studies that are integrated at the levels of phenotype characterization, high throughput experimental techniques, data analyses, and beyond.

  16. Discovery of a widely distributed toxin biosynthetic gene cluster

    PubMed Central

    Lee, Shaun W.; Mitchell, Douglas A.; Markley, Andrew L.; Hensler, Mary E.; Gonzalez, David; Wohlrab, Aaron; Dorrestein, Pieter C.; Nizet, Victor; Dixon, Jack E.

    2008-01-01

    Bacteriocins represent a large family of ribosomally produced peptide antibiotics. Here we describe the discovery of a widely conserved biosynthetic gene cluster for the synthesis of thiazole and oxazole heterocycles on ribosomally produced peptides. These clusters encode a toxin precursor and all necessary proteins for toxin maturation and export. Using the toxin precursor peptide and heterocycle-forming synthetase proteins from the human pathogen Streptococcus pyogenes, we demonstrate the in vitro reconstitution of streptolysin S activity. We provide evidence that the synthetase enzymes, as predicted from our bioinformatics analysis, introduce heterocycles onto precursor peptides, thereby providing molecular insight into the chemical structure of streptolysin S. Furthermore, our studies reveal that the synthetase exhibits relaxed substrate specificity and modifies toxin precursors from both related and distant species. Given our findings, it is likely that the discovery of similar peptidic toxins will rapidly expand to existing and emerging genomes. PMID:18375757

  17. Cloning and characterization of the biosynthetic gene cluster for kutznerides

    PubMed Central

    Fujimori, Danica Galonić; Hrvatin, Siniša; Neumann, Christopher S.; Strieker, Matthias; Marahiel, Mohamed A.; Walsh, Christopher T.

    2007-01-01

    Kutznerides, actinomycete-derived cyclic depsipetides, consist of six nonproteinogenic residues, including a highly oxygenated tricyclic hexahydropyrroloindole, a chlorinated piperazic acid, 2-(1-methylcyclopropyl)-glycine, a β-branched-hydroxy acid, and 3-hydroxy glutamic acid, for which biosynthetic logic has not been elucidated. Herein we describe the biosynthetic gene cluster for the kutzneride family, identified by degenerate primer PCR for halogenating enzymes postulated to be involved in biosyntheses of these unusual monomers. The 56-kb gene cluster encodes a series of six nonribosomal peptide synthetase (NRPS) modules distributed over three proteins and a variety of tailoring enzymes, including both mononuclear nonheme iron and two flavin-dependent halogenases, and an array of oxygen transfer catalysts. The sequence and organization of NRPS genes support incorporation of the unusual monomer units into the densely functionalized scaffold of kutznerides. Our work provides insight into the formation of this intriguing class of compounds and provides a foundation for elucidating the timing and mechanisms of their biosynthesis. PMID:17940045

  18. From hormones to secondary metabolism: the emergence of metabolic gene clusters in plants.

    PubMed

    Chu, Hoi Yee; Wegel, Eva; Osbourn, Anne

    2011-04-01

    Gene clusters for the synthesis of secondary metabolites are a common feature of microbial genomes. Well-known examples include clusters for the synthesis of antibiotics in actinomycetes, and also for the synthesis of antibiotics and toxins in filamentous fungi. Until recently it was thought that genes for plant metabolic pathways were not clustered, and this is certainly true in many cases; however, five plant secondary metabolic gene clusters have now been discovered, all of them implicated in synthesis of defence compounds. An obvious assumption might be that these eukaryotic gene clusters have arisen by horizontal gene transfer from microbes, but there is compelling evidence to indicate that this is not the case. This raises intriguing questions about how widespread such clusters are, what the significance of clustering is, why genes for some metabolic pathways are clustered and those for others are not, and how these clusters form. In answering these questions we may hope to learn more about mechanisms of genome plasticity and adaptive evolution in plants. It is noteworthy that for the five plant secondary metabolic gene clusters reported so far, the enzymes for the first committed steps all appear to have been recruited directly or indirectly from primary metabolic pathways involved in hormone synthesis. This may or may not turn out to be a common feature of plant secondary metabolic gene clusters as new clusters emerge.

  19. Molecular characterization of neurally expressing genes in the para sodium channel gene cluster of Drosophila

    SciTech Connect

    Hong, Chang-Sook; Ganetzky, B.

    1996-03-01

    To elucidate the mechanisms regulating expression of para, which encodes the major class of sodium channels in the Drosophila nervous system, we have tried to locate upstream cis-acting regulatory elements by mapping the transcriptional start site and analyzing the region immediately upstream of para in region 14D of the polytene chromosomes. From these studies, we have discovered that the region contains a cluster of neurally expressing genes. Here we report the molecular characterization of the genomic organization of the 14D region and the genes within this region, which are: calnexin (Cnx), actin related protein 14D (Arp14D), calcineurin A 14D (CnnA14D), and chromosome associated protein (Cap). The tight clustering of these genes, their neuronal expression patterns, and their potential functions related to expression, modulation, or regulation of sodium channels raise the possibility that these genes represent a functionally related group sharing some coordinate regulatory mechanism. 76 refs., 11 figs.

  20. Molecular Characterization of Neurally Expressing Genes in the Para Sodium Channel Gene Cluster of Drosophila

    PubMed Central

    Hong, C. S.; Ganetzky, B.

    1996-01-01

    To elucidate the mechanisms regulating expression of para, which encodes the major class of sodium channels in the Drosophila nervous system, we have tried to locate upstream cis-acting regulatory elements by mapping the transcriptional start site and analyzing the region immediately upstream of para in region 14D of the polytene chromosomes. From these studies, we have discovered that the region contains a cluster of neurally expressing genes. Here we report the molecular characterization of the genomic organization of the 14D region and the genes within this region, which are: calnexin (Cnx), actin related protein 14D (Arp14D), calcineurin A 14D (CnnA14D), and chromosome associated protein (Cap). The tight clustering of these genes, their neuronal expression patterns, and their potential functions related to expression, modulation, or regulation of sodium channels raise the possibility that these genes represent a functionally related group sharing some coordinate regulatory mechanism. PMID:8849894

  1. Distribution and Genetic Diversity of Bacteriocin Gene Clusters in Rumen Microbial Genomes

    PubMed Central

    Azevedo, Analice C.; Bento, Cláudia B. P.; Ruiz, Jeronimo C.; Queiroz, Marisa V.

    2015-01-01

    Some species of ruminal bacteria are known to produce antimicrobial peptides, but the screening procedures have mostly been based on in vitro assays using standardized methods. Recent sequencing efforts have made available the genome sequences of hundreds of ruminal microorganisms. In this work, we performed genome mining of the complete and partial genome sequences of 224 ruminal bacteria and 5 ruminal archaea to determine the distribution and diversity of bacteriocin gene clusters. A total of 46 bacteriocin gene clusters were identified in 33 strains of ruminal bacteria. Twenty gene clusters were related to lanthipeptide biosynthesis, while 11 gene clusters were associated with sactipeptide production, 7 gene clusters were associated with class II bacteriocin production, and 8 gene clusters were associated with class III bacteriocin production. The frequency of strains whose genomes encode putative antimicrobial peptide precursors was 14.4%. Clusters related to the production of sactipeptides were identified for the first time among ruminal bacteria. BLAST analysis indicated that the majority of the gene clusters (88%) encoding putative lanthipeptides contained all the essential genes required for lanthipeptide biosynthesis. Most strains of Streptococcus (66.6%) harbored complete lanthipeptide gene clusters, in addition to an open reading frame encoding a putative class II bacteriocin. Albusin B-like proteins were found in 100% of the Ruminococcus albus strains screened in this study. The in silico analysis provided evidence of novel biosynthetic gene clusters in bacterial species not previously related to bacteriocin production, suggesting that the rumen microbiota represents an underexplored source of antimicrobial peptides. PMID:26253660

  2. Distribution and Genetic Diversity of Bacteriocin Gene Clusters in Rumen Microbial Genomes.

    PubMed

    Azevedo, Analice C; Bento, Cláudia B P; Ruiz, Jeronimo C; Queiroz, Marisa V; Mantovani, Hilário C

    2015-10-01

    Some species of ruminal bacteria are known to produce antimicrobial peptides, but the screening procedures have mostly been based on in vitro assays using standardized methods. Recent sequencing efforts have made available the genome sequences of hundreds of ruminal microorganisms. In this work, we performed genome mining of the complete and partial genome sequences of 224 ruminal bacteria and 5 ruminal archaea to determine the distribution and diversity of bacteriocin gene clusters. A total of 46 bacteriocin gene clusters were identified in 33 strains of ruminal bacteria. Twenty gene clusters were related to lanthipeptide biosynthesis, while 11 gene clusters were associated with sactipeptide production, 7 gene clusters were associated with class II bacteriocin production, and 8 gene clusters were associated with class III bacteriocin production. The frequency of strains whose genomes encode putative antimicrobial peptide precursors was 14.4%. Clusters related to the production of sactipeptides were identified for the first time among ruminal bacteria. BLAST analysis indicated that the majority of the gene clusters (88%) encoding putative lanthipeptides contained all the essential genes required for lanthipeptide biosynthesis. Most strains of Streptococcus (66.6%) harbored complete lanthipeptide gene clusters, in addition to an open reading frame encoding a putative class II bacteriocin. Albusin B-like proteins were found in 100% of the Ruminococcus albus strains screened in this study. The in silico analysis provided evidence of novel biosynthetic gene clusters in bacterial species not previously related to bacteriocin production, suggesting that the rumen microbiota represents an underexplored source of antimicrobial peptides.

  3. Distribution and Genetic Diversity of Bacteriocin Gene Clusters in Rumen Microbial Genomes.

    PubMed

    Azevedo, Analice C; Bento, Cláudia B P; Ruiz, Jeronimo C; Queiroz, Marisa V; Mantovani, Hilário C

    2015-10-01

    Some species of ruminal bacteria are known to produce antimicrobial peptides, but the screening procedures have mostly been based on in vitro assays using standardized methods. Recent sequencing efforts have made available the genome sequences of hundreds of ruminal microorganisms. In this work, we performed genome mining of the complete and partial genome sequences of 224 ruminal bacteria and 5 ruminal archaea to determine the distribution and diversity of bacteriocin gene clusters. A total of 46 bacteriocin gene clusters were identified in 33 strains of ruminal bacteria. Twenty gene clusters were related to lanthipeptide biosynthesis, while 11 gene clusters were associated with sactipeptide production, 7 gene clusters were associated with class II bacteriocin production, and 8 gene clusters were associated with class III bacteriocin production. The frequency of strains whose genomes encode putative antimicrobial peptide precursors was 14.4%. Clusters related to the production of sactipeptides were identified for the first time among ruminal bacteria. BLAST analysis indicated that the majority of the gene clusters (88%) encoding putative lanthipeptides contained all the essential genes required for lanthipeptide biosynthesis. Most strains of Streptococcus (66.6%) harbored complete lanthipeptide gene clusters, in addition to an open reading frame encoding a putative class II bacteriocin. Albusin B-like proteins were found in 100% of the Ruminococcus albus strains screened in this study. The in silico analysis provided evidence of novel biosynthetic gene clusters in bacterial species not previously related to bacteriocin production, suggesting that the rumen microbiota represents an underexplored source of antimicrobial peptides. PMID:26253660

  4. Regulation of Drosophila Adh promoter switching by an initiator-targeted repression mechanism.

    PubMed Central

    Ren, B; Maniatis, T

    1998-01-01

    The stage-specific expression of the Drosophila alcohol dehydrogenase (Adh) gene is achieved through the alternate activation of two tandem promoters. The proximal promoter is active primarily during late embryonic development and early larval stages, while the distal promoter is active in late third instar larvae and adults. Here, we provide evidence that this Adh promoter switch is regulated by a zinc finger repressor protein (AEF-1) that is expressed predominantly in adult flies and targets the initiator region of the proximal promoter. We propose that AEF-1 plays a critical role in Adh promoter switching by blocking interactions between a component of the general transcription machinery and the initiator region of the proximal promoter. PMID:9463385

  5. Sequencing, characterization, and gene expression analysis of the histidine decarboxylase gene cluster of Morganella morganii.

    PubMed

    Ferrario, Chiara; Borgo, Francesca; de Las Rivas, Blanca; Muñoz, Rosario; Ricci, Giovanni; Fortina, Maria Grazia

    2014-03-01

    The histidine decarboxylase gene cluster of Morganella morganii DSM30146(T) was sequenced, and four open reading frames, named hdcT1, hdc, hdcT2, and hisRS were identified. Two putative histidine/histamine antiporters (hdcT1 and hdcT2) were located upstream and downstream the hdc gene, codifying a pyridoxal-P dependent histidine decarboxylase, and followed by hisRS gene encoding a histidyl-tRNA synthetase. This organization was comparable with the gene cluster of other known Gram negative bacteria, particularly with that of Klebsiella oxytoca. Recombinant Escherichia coli strains harboring plasmids carrying the M. morganii hdc gene were shown to overproduce histidine decarboxylase, after IPTG induction at 37 °C for 4 h. Quantitative RT-PCR experiments revealed the hdc and hisRS genes were highly induced under acidic and histidine-rich conditions. This work represents the first description and identification of the hdc-related genes in M. morganii. Results support the hypothesis that the histidine decarboxylation reaction in this prolific histamine producing species may play a role in acid survival. The knowledge of the role and the regulation of genes involved in histidine decarboxylation should improve the design of rational strategies to avoid toxic histamine production in foods.

  6. Gravitation field algorithm and its application in gene cluster

    PubMed Central

    2010-01-01

    Background Searching optima is one of the most challenging tasks in clustering genes from available experimental data or given functions. SA, GA, PSO and other similar efficient global optimization methods are used by biotechnologists. All these algorithms are based on the imitation of natural phenomena. Results This paper proposes a novel searching optimization algorithm called Gravitation Field Algorithm (GFA) which is derived from the famous astronomy theory Solar Nebular Disk Model (SNDM) of planetary formation. GFA simulates the Gravitation field and outperforms GA and SA in some multimodal functions optimization problem. And GFA also can be used in the forms of unimodal functions. GFA clusters the dataset well from the Gene Expression Omnibus. Conclusions The mathematical proof demonstrates that GFA could be convergent in the global optimum by probability 1 in three conditions for one independent variable mass functions. In addition to these results, the fundamental optimization concept in this paper is used to analyze how SA and GA affect the global search and the inherent defects in SA and GA. Some results and source code (in Matlab) are publicly available at http://ccst.jlu.edu.cn/CSBG/GFA. PMID:20854683

  7. Arrangement of the Clostridium baratii F7 Toxin Gene Cluster with Identification of a σ Factor That Recognizes the Botulinum Toxin Gene Cluster Promoters

    SciTech Connect

    Dover, Nir; Barash, Jason R.; Burke, Julianne N.; Hill, Karen K.; Detter, John C.; Arnon, Stephen S.

    2014-05-22

    Botulinum neurotoxin (BoNT) is the most poisonous substances known and its eight toxin types (A to H) are distinguished by the inability of polyclonal antibodies that neutralize one toxin type to neutralize any of the other seven toxin types. Infant botulism, an intestinal toxemia orphan disease, is the most common form of human botulism in the United States. It results from swallowed spores of Clostridium botulinum (or rarely, neurotoxigenic Clostridium butyricum or Clostridium baratii) that germinate and temporarily colonize the lumen of the large intestine, where, as vegetative cells, they produce botulinum toxin. Botulinum neurotoxin is encoded by the bont gene that is part of a toxin gene cluster that includes several accessory genes. In this paper, we sequenced for the first time the complete botulinum neurotoxin gene cluster of nonproteolytic C. baratii type F7. Like the type E and the nonproteolytic type F6 botulinum toxin gene clusters, the C. baratii type F7 had an orfX toxin gene cluster that lacked the regulatory botR gene which is found in proteolytic C. botulinum strains and codes for an alternative σ factor. In the absence of botR, we identified a putative alternative regulatory gene located upstream of the C. baratii type F7 toxin gene cluster. This putative regulatory gene codes for a predicted σ factor that contains DNA-binding-domain homologues to the DNA-binding domains both of BotR and of other members of the TcdR-related group 5 of the σ70 family that are involved in the regulation of toxin gene expression in clostridia. We showed that this TcdR-related protein in association with RNA polymerase core enzyme specifically binds to the C. baratii type F7 botulinum toxin gene cluster promoters. Finally, this TcdR-related protein may therefore be involved in regulating the expression of the genes of the botulinum toxin gene cluster in neurotoxigenic C. baratii.

  8. Arrangement of the Clostridium baratii F7 toxin gene cluster with identification of a σ factor that recognizes the botulinum toxin gene cluster promoters.

    PubMed

    Dover, Nir; Barash, Jason R; Burke, Julianne N; Hill, Karen K; Detter, John C; Arnon, Stephen S

    2014-01-01

    Botulinum neurotoxin (BoNT) is the most poisonous substances known and its eight toxin types (A to H) are distinguished by the inability of polyclonal antibodies that neutralize one toxin type to neutralize any of the other seven toxin types. Infant botulism, an intestinal toxemia orphan disease, is the most common form of human botulism in the United States. It results from swallowed spores of Clostridium botulinum (or rarely, neurotoxigenic Clostridium butyricum or Clostridium baratii) that germinate and temporarily colonize the lumen of the large intestine, where, as vegetative cells, they produce botulinum toxin. Botulinum neurotoxin is encoded by the bont gene that is part of a toxin gene cluster that includes several accessory genes. We sequenced for the first time the complete botulinum neurotoxin gene cluster of nonproteolytic C. baratii type F7. Like the type E and the nonproteolytic type F6 botulinum toxin gene clusters, the C. baratii type F7 had an orfX toxin gene cluster that lacked the regulatory botR gene which is found in proteolytic C. botulinum strains and codes for an alternative σ factor. In the absence of botR, we identified a putative alternative regulatory gene located upstream of the C. baratii type F7 toxin gene cluster. This putative regulatory gene codes for a predicted σ factor that contains DNA-binding-domain homologues to the DNA-binding domains both of BotR and of other members of the TcdR-related group 5 of the σ70 family that are involved in the regulation of toxin gene expression in clostridia. We showed that this TcdR-related protein in association with RNA polymerase core enzyme specifically binds to the C. baratii type F7 botulinum toxin gene cluster promoters. This TcdR-related protein may therefore be involved in regulating the expression of the genes of the botulinum toxin gene cluster in neurotoxigenic C. baratii.

  9. Parallel evolutionary events in the haptoglobin gene clusters of rhesus monkey and human

    SciTech Connect

    Erickson, L.M.; Maeda, N.

    1994-08-01

    Parallel occurrences of evolutionary events in the haptoglobin gene clusters of rhesus monkeys and humans were studied. We found six different haplotypes among 11 individuals from two rhesus monkey families. The six haplotypes include two types of haptoglobin gene clusters: one type with a single gene and the other with two genes. DNA sequence analysis indicates that the one-gene and the two-gene clusters were both formed by unequal homologous crossovers between two genes of an ancestral three-gene cluster, near exon 5, the longest exon of the gene. This exon is also the location where a separate unequal homologous crossover occured in the human lineage, forming the human two-gene haptoglobin gene cluster from an ancestral three-gene cluster. The occurrence of independent homologous unequal crossovers in rhesus monkey and in human within the same region of DNA suggests that the evolutionary history of the haptoglobin gene cluster in primates is the consequence of frequent homologous pairings facilitated by the longest and most conserved exon of the gene. 27 refs., 7 figs., 1 tab.

  10. Recurrent adenylation domain replacement in the microcystin synthetase gene cluster

    PubMed Central

    Fewer, David P; Rouhiainen, Leo; Jokela, Jouni; Wahlsten, Matti; Laakso, Kati; Wang, Hao; Sivonen, Kaarina

    2007-01-01

    Background Microcystins are small cyclic heptapeptide toxins produced by a range of distantly related cyanobacteria. Microcystins are synthesized on large NRPS-PKS enzyme complexes. Many structural variants of microcystins are produced simulatenously. A recombination event between the first module of mcyB (mcyB1) and mcyC in the microcystin synthetase gene cluster is linked to the simultaneous production of microcystin variants in strains of the genus Microcystis. Results Here we undertook a phylogenetic study to investigate the order and timing of recombination between the mcyB1 and mcyC genes in a diverse selection of microcystin producing cyanobacteria. Our results provide support for complex evolutionary processes taking place at the mcyB1 and mcyC adenylation domains which recognize and activate the amino acids found at X and Z positions. We find evidence for recent recombination between mcyB1 and mcyC in strains of the genera Anabaena, Microcystis, and Hapalosiphon. We also find clear evidence for independent adenylation domain conversion of mcyB1 by unrelated peptide synthetase modules in strains of the genera Nostoc and Microcystis. The recombination events replace only the adenylation domain in each case and the condensation domains of mcyB1 and mcyC are not transferred together with the adenylation domain. Our findings demonstrate that the mcyB1 and mcyC adenylation domains are recombination hotspots in the microcystin synthetase gene cluster. Conclusion Recombination is thought to be one of the main mechanisms driving the diversification of NRPSs. However, there is very little information on how recombination takes place in nature. This study demonstrates that functional peptide synthetases are created in nature through transfer of adenylation domains without the concomitant transfer of condensation domains. PMID:17908306

  11. A Hybrid Distance Measure for Clustering Expressed Sequence Tags Originating from the Same Gene Family

    PubMed Central

    Ng, Keng-Hoong; Ho, Chin-Kuan; Phon-Amnuaisuk, Somnuk

    2012-01-01

    Background Clustering is a key step in the processing of Expressed Sequence Tags (ESTs). The primary goal of clustering is to put ESTs from the same transcript of a single gene into a unique cluster. Recent EST clustering algorithms mostly adopt the alignment-free distance measures, where they tend to yield acceptable clustering accuracies with reasonable computational time. Despite the fact that these clustering methods work satisfactorily on a majority of the EST datasets, they have a common weakness. They are prone to deliver unsatisfactory clustering results when dealing with ESTs from the genes derived from the same family. The root cause is the distance measures applied on them are not sensitive enough to separate these closely related genes. Methodology/Principal Findings We propose a hybrid distance measure that combines the global and local features extracted from ESTs, with the aim to address the clustering problem faced by ESTs derived from the same gene family. The clustering process is implemented using the DBSCAN algorithm. We test the hybrid distance measure on the ten EST datasets, and the clustering results are compared with the two alignment-free EST clustering tools, i.e. wcd and PEACE. The clustering results indicate that the proposed hybrid distance measure performs relatively better (in terms of clustering accuracy) than both EST clustering tools. Conclusions/Significance The clustering results provide support for the effectiveness of the proposed hybrid distance measure in solving the clustering problem for ESTs that originate from the same gene family. The improvement of clustering accuracies on the experimental datasets has supported the claim that the sensitivity of the hybrid distance measure is sufficient to solve the clustering problem. PMID:23071763

  12. Mutational and Phylogenetic Analyses of the Mycobacterial mbt Gene Cluster ▿§

    PubMed Central

    Chavadi, Sivagami Sundaram; Stirrett, Karen L.; Edupuganti, Uthamaphani R.; Vergnolle, Olivia; Sadhanandan, Gigani; Marchiano, Emily; Martin, Che; Qiu, Wei-Gang; Soll, Clifford E.; Quadri, Luis E. N.

    2011-01-01

    The mycobactin siderophore system is present in many Mycobacterium species, including M. tuberculosis and other clinically relevant mycobacteria. This siderophore system is believed to be utilized by both pathogenic and nonpathogenic mycobacteria for iron acquisition in both in vivo and ex vivo iron-limiting environments, respectively. Several M. tuberculosis genes located in a so-called mbt gene cluster have been predicted to be required for the biosynthesis of the core scaffold of mycobactin based on sequence analysis. A systematic and controlled mutational analysis probing the hypothesized essential nature of each of these genes for mycobactin production has been lacking. The degree of conservation of mbt gene cluster orthologs remains to be investigated as well. In this study, we sought to conclusively establish whether each of nine mbt genes was required for mycobactin production and to examine the conservation of gene clusters orthologous to the M. tuberculosis mbt gene cluster in other bacteria. We report a systematic mutational analysis of the mbt gene cluster ortholog found in Mycobacterium smegmatis. This mutational analysis demonstrates that eight of the nine mbt genes investigated are essential for mycobactin production. Our genome mining and phylogenetic analyses reveal the presence of orthologous mbt gene clusters in several bacterial species. These gene clusters display significant organizational differences originating from an intricate evolutionary path that might have included horizontal gene transfers. Altogether, the findings reported herein advance our understanding of the genetic requirements for the biosynthesis of an important mycobacterial secondary metabolite with relevance to virulence. PMID:21873494

  13. A Special Local Clustering Algorithm for Identifying the Genes Associated With Alzheimer’s Disease

    PubMed Central

    Pang, Chao-Yang; Hu, Wei; Hu, Ben-Qiong; Shi, Ying; Vanderburg, Charles R.; Rogers, Jack T.

    2010-01-01

    Clustering is the grouping of similar objects into a class. Local clustering feature refers to the phenomenon whereby one group of data is separated from another, and the data from these different groups are clustered locally. A compact class is defined as one cluster in which all similar elements cluster tightly within the cluster. Herein, the essence of the local clustering feature, revealed by mathematical manipulation, results in a novel clustering algorithm termed as the special local clustering (SLC) algorithm that was used to process gene microarray data related to Alzheimer’s disease (AD). SLC algorithm was able to group together genes with similar expression patterns and identify significantly varied gene expression values as isolated points. If a gene belongs to a compact class in control data and appears as an isolated point in incipient, moderate and/or severe AD gene microarray data, this gene is possibly associated with AD. Application of a clustering algorithm in disease-associated gene identification such as in AD is rarely reported. PMID:20089478

  14. Translating biosynthetic gene clusters into fungal armor and weaponry

    PubMed Central

    Keller, Nancy P

    2015-01-01

    Filamentous fungi are renowned for the production of a diverse array of secondary metabolites (SMs) where the genetic material required for synthesis of a SM is typically arrayed in a biosynthetic gene cluster (BGC). These natural products are valued for their bioactive properties stemming from their functions in fungal biology, key among those protection from abiotic and biotic stress and establishment of a secure niche. The producing fungus must not only avoid self-harm from endogenous SMs but also deliver specific SMs at the right time to the right tissue requiring biochemical aid. This review highlights functions of BGCs beyond the enzymatic assembly of SMs, considering the timing and location of SM production and other proteins in the clusters that control SM activity. Specifically, self-protection is provided by both BGC-encoded mechanisms and non-BGC subcellular containment of toxic SM precursors; delivery and timing is orchestrated through cellular trafficking patterns and stress- and developmental-responsive transcriptional programs. PMID:26284674

  15. The Xenopus alcohol dehydrogenase gene family: characterization and comparative analysis incorporating amphibian and reptilian genomes

    PubMed Central

    2014-01-01

    Background The alcohol dehydrogenase (ADH) gene family uniquely illustrates the concept of enzymogenesis. In vertebrates, tandem duplications gave rise to a multiplicity of forms that have been classified in eight enzyme classes, according to primary structure and function. Some of these classes appear to be exclusive of particular organisms, such as the frog ADH8, a unique NADP+-dependent ADH enzyme. This work describes the ADH system of Xenopus, as a model organism, and explores the first amphibian and reptilian genomes released in order to contribute towards a better knowledge of the vertebrate ADH gene family. Results Xenopus cDNA and genomic sequences along with expressed sequence tags (ESTs) were used in phylogenetic analyses and structure-function correlations of amphibian ADHs. Novel ADH sequences identified in the genomes of Anolis carolinensis (anole lizard) and Pelodiscus sinensis (turtle) were also included in these studies. Tissue and stage-specific libraries provided expression data, which has been supported by mRNA detection in Xenopus laevis tissues and regulatory elements in promoter regions. Exon-intron boundaries, position and orientation of ADH genes were deduced from the amphibian and reptilian genome assemblies, thus revealing syntenic regions and gene rearrangements with respect to the human genome. Our results reveal the high complexity of the ADH system in amphibians, with eleven genes, coding for seven enzyme classes in Xenopus tropicalis. Frogs possess the amphibian-specific ADH8 and the novel ADH1-derived forms ADH9 and ADH10. In addition, they exhibit ADH1, ADH2, ADH3 and ADH7, also present in reptiles and birds. Class-specific signatures have been assigned to ADH7, and ancestral ADH2 is predicted to be a mixed-class as the ostrich enzyme, structurally close to mammalian ADH2 but with class-I kinetic properties. Remarkably, many ADH1 and ADH7 forms are observed in the lizard, probably due to lineage-specific duplications. ADH4 is not

  16. Deletion and Gene Expression Analyses Define the Paxilline Biosynthetic Gene Cluster in Penicillium paxilli

    PubMed Central

    Scott, Barry; Young, Carolyn A.; Saikia, Sanjay; McMillan, Lisa K.; Monahan, Brendon J.; Koulman, Albert; Astin, Jonathan; Eaton, Carla J.; Bryant, Andrea; Wrenn, Ruth E.; Finch, Sarah C.; Tapper, Brian A.; Parker, Emily J.; Jameson, Geoffrey B.

    2013-01-01

    The indole-diterpene paxilline is an abundant secondary metabolite synthesized by Penicillium paxilli. In total, 21 genes have been identified at the PAX locus of which six have been previously confirmed to have a functional role in paxilline biosynthesis. A combination of bioinformatics, gene expression and targeted gene replacement analyses were used to define the boundaries of the PAX gene cluster. Targeted gene replacement identified seven genes, paxG, paxA, paxM, paxB, paxC, paxP and paxQ that were all required for paxilline production, with one additional gene, paxD, required for regular prenylation of the indole ring post paxilline synthesis. The two putative transcription factors, PP104 and PP105, were not co-regulated with the pax genes and based on targeted gene replacement, including the double knockout, did not have a role in paxilline production. The relationship of indole dimethylallyl transferases involved in prenylation of indole-diterpenes such as paxilline or lolitrem B, can be found as two disparate clades, not supported by prenylation type (e.g., regular or reverse). This paper provides insight into the P. paxilli indole-diterpene locus and reviews the recent advances identified in paxilline biosynthesis. PMID:23949005

  17. The gsdf gene locus harbors evolutionary conserved and clustered genes preferentially expressed in fish previtellogenic oocytes.

    PubMed

    Gautier, Aude; Le Gac, Florence; Lareyre, Jean-Jacques

    2011-02-01

    display a different cellular localization compared to that of the gsdf gene indicating that the later gene is not co-regulated. Interestingly, our study identifies new clustered genes that are specifically expressed in previtellogenic oocytes (nup54, aff1, klhl8, sdad1).

  18. Identification and Functional Analysis of the Nocardithiocin Gene Cluster in Nocardia pseudobrasiliensis

    PubMed Central

    Sakai, Kanae; Komaki, Hisayuki; Gonoi, Tohru

    2015-01-01

    Nocardithiocin is a thiopeptide compound isolated from the opportunistic pathogen Nocardia pseudobrasiliensis. It shows a strong activity against acid-fast bacteria and is also active against rifampicin-resistant Mycobacterium tuberculosis. Here, we report the identification of the nocardithiocin gene cluster in N. pseudobrasiliensis IFM 0761 based on conserved thiopeptide biosynthesis gene sequence and the whole genome sequence. The predicted gene cluster was confirmed by gene disruption and complementation. As expected, strains containing the disrupted gene did not produce nocardithiocin while gene complementation restored nocardithiocin production in these strains. The predicted cluster was further analyzed using RNA-seq which showed that the nocardithiocin gene cluster contains 12 genes within a 15.2-kb region. This finding will promote the improvement of nocardithiocin productivity and its derivatives production. PMID:26588225

  19. Epigenetic regulation of the RHOX homeobox gene cluster and its association with human male infertility.

    PubMed

    Richardson, Marcy E; Bleiziffer, Andreas; Tüttelmann, Frank; Gromoll, Jörg; Wilkinson, Miles F

    2014-01-01

    The X-linked RHOX cluster encodes a set of homeobox genes that are selectively expressed in the reproductive tract. Members of the RHOX cluster regulate target genes important for spermatogenesis promote male fertility in mice. Studies show that demethylating agents strongly upregulate the expression of mouse Rhox genes, suggesting that they are regulated by DNA methylation. However, whether this extends to human RHOX genes, whether DNA methylation directly regulates RHOX gene transcription and how this relates to human male infertility are unknown. To address these issues, we first defined the promoter regions of human RHOX genes and performed gain- and loss-of-function experiments to determine whether human RHOX gene transcription is regulated by DNA methylation. Our results indicated that DNA methylation is necessary and sufficient to silence human RHOX gene expression. To determine whether RHOX cluster methylation associates with male infertility, we evaluated the methylation status of RHOX genes in sperm from a large cohort of infertility patients. Linear regression analysis revealed a strong association between RHOX gene cluster hypermethylation and three independent types of semen abnormalities. Hypermethylation was restricted specifically to the RHOX cluster; we did not observe it in genes immediately adjacent to it on the X chromosome. Our results strongly suggest that human RHOX homeobox genes are under an epigenetic control mechanism that is aberrantly regulated in infertility patients. We propose that hypermethylation of the RHOX gene cluster serves as a marker for idiopathic infertility and that it is a candidate to exert a causal role in male infertility.

  20. An Ergot Alkaloid Biosynthesis Gene and Clustered Hypothetical Genes from Aspergillus fumigatus†

    PubMed Central

    Coyle, Christine M.; Panaccione, Daniel G.

    2005-01-01

    The ergot alkaloids are a family of indole-derived mycotoxins with a variety of significant biological activities. Aspergillus fumigatus, a common airborne fungus and opportunistic human pathogen, and several fungi in the relatively distant taxon Clavicipitaceae (clavicipitaceous fungi) produce different sets of ergot alkaloids. The ergot alkaloids of these divergent fungi share a four-member ergoline ring but differ in the number, type, and position of the side chains. Several genes required for ergot alkaloid production are known in the clavicipitaceous fungi, and these genes are clustered in the genome of the ergot fungus Claviceps purpurea. We investigated whether the ergot alkaloids of A. fumigatus have a common biosynthetic and genetic origin with those of the clavicipitaceous fungi. A homolog of dmaW, the gene controlling the determinant step in the ergot alkaloid pathway of clavicipitaceous fungi, was identified in the A. fumigatus genome. Knockout of dmaW eliminated all known ergot alkaloids from A. fumigatus, and complementation of the mutation restored ergot alkaloid production. Clustered with dmaW in the A. fumigatus genome are sequences corresponding to five genes previously proposed to encode steps in the ergot alkaloid pathway of C. purpurea, as well as additional sequences whose deduced protein products are consistent with their involvement in the ergot alkaloid pathway. The corresponding genes have similarities in their nucleotide sequences, but the orientations and positions within the cluster of several of these genes differ. The data indicate that the ergot alkaloid biosynthetic capabilities in A. fumigatus and the clavicipitaceous fungi had a common origin. PMID:15933009

  1. Activation and Characterization of a Cryptic Polycyclic Tetramate Macrolactam Biosynthetic Gene Cluster

    PubMed Central

    Luo, Yunzi; Huang, Hua; Liang, Jing; Wang, Meng; Lu, Lu; Shao, Zengyi; Cobb, Ryan E.; Zhao, Huimin

    2014-01-01

    Polycyclic tetramate macrolactams (PTMs) are a widely distributed class of natural products with important biological activities. However, many of them have not been characterized. Here we apply a plug and play synthetic biology strategy to activate a cryptic PTM biosynthetic gene cluster SGR810-815 from Streptomyces griseus and discover three potential PTMs. This gene cluster is highly conserved in phylogenetically diverse bacterial strains and contains an unusual hybrid polyketide synthase-nonribosomal peptide synthetase (PKS-NRPS) which resembles iterative PKSs known in fungi. To further characterize this gene cluster, we use the same synthetic biology approach to create a series of gene deletion constructs and elucidate the biosynthetic steps for the formation of the polycyclic system. The strategy we employ bypasses the traditional laborious processes to elicit gene cluster expression and should be generally applicable to many other silent or cryptic gene clusters for discovery and characterization of new natural products. PMID:24305602

  2. High presence/absence gene variability in defense-related gene clusters of Cucumis melo

    PubMed Central

    2013-01-01

    Background Changes in the copy number of DNA sequences are one of the main mechanisms generating genome variability in eukaryotes. These changes are often related to phenotypic effects such as genetic disorders or novel pathogen resistance. The increasing availability of genome sequences through the application of next-generation massive sequencing technologies has allowed the study of genomic polymorphisms at both the interspecific and intraspecific levels, thus helping to understand how species adapt to changing environments through genome variability. Results Data on gene presence/absence variation (PAV) in melon was obtained by resequencing a cultivated accession and an old-relative melon variety, and using previously obtained resequencing data from three other melon cultivars, among them DHL92, on which the current draft melon genome sequence is based. A total of 1,697 PAV events were detected, involving 4.4% of the predicted melon gene complement. In all, an average 1.5% of genes were absent from each analyzed cultivar as compared to the DHL92 reference genome. The most populated functional category among the 304 PAV genes of known function was that of stress response proteins (30% of all classified PAVs). Our results suggest that genes from multi-copy families are five times more likely to be affected by PAV than singleton genes. Also, the chance of genes present in the genome in tandem arrays being affected by PAV is double that of isolated genes, with PAV genes tending to be in longer clusters. The highest concentration of PAV events detected in the melon genome was found in a 1.1 Mb region of linkage group V, which also shows the highest density of melon stress-response genes. In particular, this region contains the longest continuous gene-containing PAV sequence so far identified in melon. Conclusions The first genome-wide report of PAV variation among several melon cultivars is presented here. Multi-copy and clustered genes, especially those with

  3. Phenotype-based clustering of glycosylation-related genes by RNAi-mediated gene silencing.

    PubMed

    Yamamoto-Hino, Miki; Yoshida, Hideki; Ichimiya, Tomomi; Sakamura, Sho; Maeda, Megumi; Kimura, Yoshinobu; Sasaki, Norihiko; Aoki-Kinoshita, Kiyoko F; Kinoshita-Toyoda, Akiko; Toyoda, Hidenao; Ueda, Ryu; Nishihara, Shoko; Goto, Satoshi

    2015-06-01

    Glycan structures are synthesized by a series of reactions conducted by glycosylation-related (GR) proteins such as glycosyltransferases, glycan-modifying enzymes, and nucleotide-sugar transporters. For example, the common core region of glycosaminoglycans (GAGs) is sequentially synthesized by peptide-O-xylosyltransferase, β1,4-galactosyltransferase I, β1,3-galactosyltransferase II, and β1,3-glucuronyltransferase. This raises the possibility that functional impairment of GR proteins involved in synthesis of the same glycan might result in the same phenotypic abnormality. To examine this possibility, comprehensive silencing of genes encoding GR and proteoglycan core proteins was conducted in Drosophila. Drosophila GR candidate genes (125) were classified into five functional groups for synthesis of GAGs, N-linked, O-linked, Notch-related, and unknown glycans. Spatiotemporally regulated silencing caused a range of malformed phenotypes that fell into three types: extra veins, thick veins, and depigmentation. The clustered phenotypes reflected the biosynthetic pathways of GAGs, Fringe-dependent glycan on Notch, and glycans placed at or near nonreducing ends (herein termed terminal domains of glycans). Based on the phenotypic clustering, CG33145 was predicted to be involved in formation of terminal domains. Our further analysis showed that CG33145 exhibited galactosyltransferase activity in synthesis of terminal N-linked glycans. Phenotypic clustering, therefore, has potential for the functional prediction of novel GR genes. PMID:25940448

  4. Phenotype-based clustering of glycosylation-related genes by RNAi-mediated gene silencing

    PubMed Central

    Yamamoto-Hino, Miki; Yoshida, Hideki; Ichimiya, Tomomi; Sakamura, Sho; Maeda, Megumi; Kimura, Yoshinobu; Sasaki, Norihiko; Aoki-Kinoshita, Kiyoko F; Kinoshita-Toyoda, Akiko; Toyoda, Hidenao; Ueda, Ryu; Nishihara, Shoko; Goto, Satoshi

    2015-01-01

    Glycan structures are synthesized by a series of reactions conducted by glycosylation-related (GR) proteins such as glycosyltransferases, glycan-modifying enzymes, and nucleotide-sugar transporters. For example, the common core region of glycosaminoglycans (GAGs) is sequentially synthesized by peptide-O-xylosyltransferase, β1,4-galactosyltransferase I, β1,3-galactosyltransferase II, and β1,3-glucuronyltransferase. This raises the possibility that functional impairment of GR proteins involved in synthesis of the same glycan might result in the same phenotypic abnormality. To examine this possibility, comprehensive silencing of genes encoding GR and proteoglycan core proteins was conducted in Drosophila. Drosophila GR candidate genes (125) were classified into five functional groups for synthesis of GAGs, N-linked, O-linked, Notch-related, and unknown glycans. Spatiotemporally regulated silencing caused a range of malformed phenotypes that fell into three types: extra veins, thick veins, and depigmentation. The clustered phenotypes reflected the biosynthetic pathways of GAGs, Fringe-dependent glycan on Notch, and glycans placed at or near nonreducing ends (herein termed terminal domains of glycans). Based on the phenotypic clustering, CG33145 was predicted to be involved in formation of terminal domains. Our further analysis showed that CG33145 exhibited galactosyltransferase activity in synthesis of terminal N-linked glycans. Phenotypic clustering, therefore, has potential for the functional prediction of novel GR genes. PMID:25940448

  5. A tripartite clustering analysis on microRNA, gene and disease model.

    PubMed

    Shen, Chengcheng; Liu, Ying

    2012-02-01

    Alteration of gene expression in response to regulatory molecules or mutations could lead to different diseases. MicroRNAs (miRNAs) have been discovered to be involved in regulation of gene expression and a wide variety of diseases. In a tripartite biological network of human miRNAs, their predicted target genes and the diseases caused by altered expressions of these genes, valuable knowledge about the pathogenicity of miRNAs, involved genes and related disease classes can be revealed by co-clustering miRNAs, target genes and diseases simultaneously. Tripartite co-clustering can lead to more informative results than traditional co-clustering with only two kinds of members and pass the hidden relational information along the relation chain by considering multi-type members. Here we report a spectral co-clustering algorithm for k-partite graph to find clusters with heterogeneous members. We use the method to explore the potential relationships among miRNAs, genes and diseases. The clusters obtained from the algorithm have significantly higher density than randomly selected clusters, which means members in the same cluster are more likely to have common connections. Results also show that miRNAs in the same family based on the hairpin sequences tend to belong to the same cluster. We also validate the clustering results by checking the correlation of enriched gene functions and disease classes in the same cluster. Finally, widely studied miR-17-92 and its paralogs are analyzed as a case study to reveal that genes and diseases co-clustered with the miRNAs are in accordance with current research findings. PMID:22809308

  6. A tripartite clustering analysis on microRNA, gene and disease model.

    PubMed

    Shen, Chengcheng; Liu, Ying

    2012-02-01

    Alteration of gene expression in response to regulatory molecules or mutations could lead to different diseases. MicroRNAs (miRNAs) have been discovered to be involved in regulation of gene expression and a wide variety of diseases. In a tripartite biological network of human miRNAs, their predicted target genes and the diseases caused by altered expressions of these genes, valuable knowledge about the pathogenicity of miRNAs, involved genes and related disease classes can be revealed by co-clustering miRNAs, target genes and diseases simultaneously. Tripartite co-clustering can lead to more informative results than traditional co-clustering with only two kinds of members and pass the hidden relational information along the relation chain by considering multi-type members. Here we report a spectral co-clustering algorithm for k-partite graph to find clusters with heterogeneous members. We use the method to explore the potential relationships among miRNAs, genes and diseases. The clusters obtained from the algorithm have significantly higher density than randomly selected clusters, which means members in the same cluster are more likely to have common connections. Results also show that miRNAs in the same family based on the hairpin sequences tend to belong to the same cluster. We also validate the clustering results by checking the correlation of enriched gene functions and disease classes in the same cluster. Finally, widely studied miR-17-92 and its paralogs are analyzed as a case study to reveal that genes and diseases co-clustered with the miRNAs are in accordance with current research findings.

  7. [Molecular evidences of non-ADH pathway in alcohol metabolism and Class III alcohol dehydrogenase (ADH3)].

    PubMed

    Haseba, Takeshi

    2014-06-01

    Class I alcohol dehydrogenase (ADH1), a key enzyme of alcohol metabolism, contributes around 70% to the systemic alcohol metabolism and also to the acceleration of the metabolism due to chronic alcohol consumption by increasing its liver content, if the liver damage or disease is not apparent. However, the contribution of ADH1 to alcohol metabolism decreases in case of acute alcohol poisoning or chronic alcohol consumption inducing liver damage or disease. On the contrary, non-ADH pathway, which is independent of ADH1, increases the contribution to alcohol metabolism in these cases, by complementing the reduced role of ADH1. The molecular substantiality of non-ADH pathway has been still unknown in spite of the long and hot controversy between two candidates of microsomal ethanol oxidizing system (MEOS) and catalase. This research history suggests the existence of other candidates. Among ADH isozymes, Class III (ADH3) has the highest Km for ethanol and the highest resistance to pyrazole reagents of specific ADH inhibitors. This ADH3 was demonstrated to increase the contribution to alcohol metabolism in vivo dose-dependently, therefore, is a potent candidate of non-ADH pathway. Moreover, ADH3 is considered to increase the contribution to alcohol metabolism in case of alcoholic liver diseases, because the enzyme content increases in damaged tissues with increased hydrophobicity or the activity of the liver correlates with the accumulated alcohol consumptions of patients with alcoholic liver diseases. Such adaptation of ADH3 to alcohol metabolism in these pathological conditions makes patients possible to keep drinking a lot in spite of decrease of ADH1 activity and develops alcoholism seriously.

  8. Conservation of Hox gene clusters in the self-fertilizing fish Kryptolebias marmoratus (Cyprinodontiformes; Rivulidae).

    PubMed

    Kim, B-M; Lee, B-Y; Lee, J-H; Rhee, J-S; Lee, J-S

    2016-03-01

    In this study, whole Hox gene clusters in the self-fertilizing mangrove killifish Kryptolebias marmoratus (Cyprinodontiformes; Rivulidae), a unique hermaphroditic vertebrate in which both sex organs are functional at the same time, were identified from whole genome and transcriptome sequences. The aim was to increase the understanding of the evolutionary status of conservation of this Hox gene cluster across fish species. PMID:26822496

  9. Identification and analysis of a highly conserved chemotaxis gene cluster in Shewanella species.

    SciTech Connect

    Li, J.; Romine, Margaret F.; Ward, M.

    2007-08-01

    A conserved cluster of chemotaxis genes was identified from the genome sequences of fifteen Shewanella species. An in-frame deletion of the cheA-3 gene, which is located in this cluster, was created in S. oneidensis MR-1 and the gene shown to be essential for chemotactic responses to anaerobic electron acceptors. The CheA-3 protein showed strong similarity to Vibrio cholerae CheA-2 and P. aeruginosa CheA-1, two proteins that are also essential for chemotaxis. The genes encoding these proteins were shown to be located in chemotaxis gene clusters closely related to the cheA-3-containing cluster in Shewanella species. The results of this study suggest that a combination of gene neighborhood and homology analyses may be used to predict which cheA genes are essential for chemotaxis in groups of closely related microorganisms.

  10. A phylogenomic gene cluster resource: The phylogeneticallyinferred groups (PhlGs) database

    SciTech Connect

    Dehal, Paramvir S.; Boore, Jeffrey L.

    2005-08-25

    We present here the PhIGs database, a phylogenomic resource for sequenced genomes. Although many methods exist for clustering gene families, very few attempt to create truly orthologous clusters sharing descent from a single ancestral gene across a range of evolutionary depths. Although these non-phylogenetic gene family clusters have been used broadly for gene annotation, errors are known to be introduced by the artifactual association of slowly evolving paralogs and lack of annotation for those more rapidly evolving. A full phylogenetic framework is necessary for accurate inference of function and for many studies that address pattern and mechanism of the evolution of the genome. The automated generation of evolutionary gene clusters, creation of gene trees, determination of orthology and paralogy relationships, and the correlation of this information with gene annotations, expression information, and genomic context is an important resource to the scientific community.

  11. Base J represses genes at the end of polycistronic gene clusters in Leishmania major by promoting RNAP II termination.

    PubMed

    Reynolds, David L; Hofmeister, Brigitte T; Cliffe, Laura; Siegel, T Nicolai; Anderson, Britta A; Beverley, Stephen M; Schmitz, Robert J; Sabatini, Robert

    2016-08-01

    The genomes of kinetoplastids are organized into polycistronic gene clusters that are flanked by the modified DNA base J. Previous work has established a role of base J in promoting RNA polymerase II termination in Leishmania spp. where the loss of J leads to termination defects and transcription into adjacent gene clusters. It remains unclear whether these termination defects affect gene expression and whether read through transcription is detrimental to cell growth, thus explaining the essential nature of J. We now demonstrate that reduction of base J at specific sites within polycistronic gene clusters in L. major leads to read through transcription and increased expression of downstream genes in the cluster. Interestingly, subsequent transcription into the opposing polycistronic gene cluster does not lead to downregulation of sense mRNAs. These findings indicate a conserved role for J regulating transcription termination and expression of genes within polycistronic gene clusters in trypanosomatids. In contrast to the expectations often attributed to opposing transcription, the essential nature of J in Leishmania spp. is related to its role in gene repression rather than preventing transcriptional interference resulting from read through and dual strand transcription.

  12. Base J represses genes at the end of polycistronic gene clusters in Leishmania major by promoting RNAP II termination

    PubMed Central

    Reynolds, David L.; Hofmeister, Brigitte T.; Cliffe, Laura; Siegel, T. Nicolai; Anderson, Britta A.; Beverley, Stephen M.; Schmitz, Robert J.; Sabatini, Robert

    2016-01-01

    Summary The genomes of kinetoplastids are organized into polycistronic gene clusters that are flanked by the modified DNA base J. Previous work has established a role of base J in promoting RNA polymerase II termination in Leishmania spp. where the loss of J leads to termination defects and transcription into adjacent gene clusters. It remains unclear whether these termination defects affect gene expression and whether read through transcription is detrimental to cell growth, thus explaining the essential nature of J. We now demonstrate that reduction of base J at specific sites within polycistronic gene clusters in L. major leads to read through transcription and increased expression of downstream genes in the cluster. Interestingly, subsequent transcription into the opposing polycistronic gene cluster does not lead to downregulation of sense mRNAs. These findings indicate a conserved role for J regulating transcription termination and expression of genes within polycistronic gene clusters in trypanosomatids. In contrast to the expectations often attributed to opposing transcription, the essential nature of J in Leishmania spp. is related to its role in gene repression rather than preventing transcriptional interference resulting from read through and dual strand transcription. PMID:27125778

  13. The genomisotopic approach: a systematic method to isolate products of orphan biosynthetic gene clusters.

    PubMed

    Gross, Harald; Stockwell, Virginia O; Henkels, Marcella D; Nowak-Thompson, Brian; Loper, Joyce E; Gerwick, William H

    2007-01-01

    With the increasing number of genomes sequenced and available in the public domain, a large number of orphan gene clusters, for which the encoded natural product is unknown, have been identified. These orphan gene clusters represent a tremendous source of novel and possibly bioactive compounds. Here, we describe a "genomisotopic approach," which employs a combination of genomic sequence analysis and isotope-guided fractionation to identify unknown compounds synthesized from orphan gene clusters containing nonribosomal peptide synthetases. Analysis of the Pseudomonas fluorescens Pf-5 genome led to the identification of an orphan gene cluster predicted to code for the biosynthesis of a lipopeptide natural product. Application of the genomisotopic approach to isolate the product of this gene cluster resulted in the discovery of orfamide A, founder of a group of bioactive cyclic lipopeptides.

  14. Identification and Characterization of a Novel Diterpene Gene Cluster in Aspergillus nidulans

    PubMed Central

    Bromann, Kirsi; Toivari, Mervi; Viljanen, Kaarina; Vuoristo, Anu; Ruohonen, Laura; Nakari-Setälä, Tiina

    2012-01-01

    Fungal secondary metabolites are a rich source of medically useful compounds due to their pharmaceutical and toxic properties. Sequencing of fungal genomes has revealed numerous secondary metabolite gene clusters, yet products of many of these biosynthetic pathways are unknown since the expression of the clustered genes usually remains silent in normal laboratory conditions. Therefore, to discover new metabolites, it is important to find ways to induce the expression of genes in these otherwise silent biosynthetic clusters. We discovered a novel secondary metabolite in Aspergillus nidulans by predicting a biosynthetic gene cluster with genomic mining. A Zn(II)2Cys6–type transcription factor, PbcR, was identified, and its role as a pathway-specific activator for the predicted gene cluster was demonstrated. Overexpression of pbcR upregulated the transcription of seven genes in the identified cluster and led to the production of a diterpene compound, which was characterized with GC/MS as ent-pimara-8(14),15-diene. A change in morphology was also observed in the strains overexpressing pbcR. The activation of a cryptic gene cluster by overexpression of its putative Zn(II)2Cys6–type transcription factor led to discovery of a novel secondary metabolite in Aspergillus nidulans. Quantitative real-time PCR and DNA array analysis allowed us to predict the borders of the biosynthetic gene cluster. Furthermore, we identified a novel fungal pimaradiene cyclase gene as well as genes encoding 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase and a geranylgeranyl pyrophosphate (GGPP) synthase. None of these genes have been previously implicated in the biosynthesis of terpenes in Aspergillus nidulans. These results identify the first Aspergillus nidulans diterpene gene cluster and suggest a biosynthetic pathway for ent-pimara-8(14),15-diene. PMID:22506079

  15. Identification and characterization of a novel diterpene gene cluster in Aspergillus nidulans.

    PubMed

    Bromann, Kirsi; Toivari, Mervi; Viljanen, Kaarina; Vuoristo, Anu; Ruohonen, Laura; Nakari-Setälä, Tiina

    2012-01-01

    Fungal secondary metabolites are a rich source of medically useful compounds due to their pharmaceutical and toxic properties. Sequencing of fungal genomes has revealed numerous secondary metabolite gene clusters, yet products of many of these biosynthetic pathways are unknown since the expression of the clustered genes usually remains silent in normal laboratory conditions. Therefore, to discover new metabolites, it is important to find ways to induce the expression of genes in these otherwise silent biosynthetic clusters. We discovered a novel secondary metabolite in Aspergillus nidulans by predicting a biosynthetic gene cluster with genomic mining. A Zn(II)(2)Cys(6)-type transcription factor, PbcR, was identified, and its role as a pathway-specific activator for the predicted gene cluster was demonstrated. Overexpression of pbcR upregulated the transcription of seven genes in the identified cluster and led to the production of a diterpene compound, which was characterized with GC/MS as ent-pimara-8(14),15-diene. A change in morphology was also observed in the strains overexpressing pbcR. The activation of a cryptic gene cluster by overexpression of its putative Zn(II)(2)Cys(6)-type transcription factor led to discovery of a novel secondary metabolite in Aspergillus nidulans. Quantitative real-time PCR and DNA array analysis allowed us to predict the borders of the biosynthetic gene cluster. Furthermore, we identified a novel fungal pimaradiene cyclase gene as well as genes encoding 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase and a geranylgeranyl pyrophosphate (GGPP) synthase. None of these genes have been previously implicated in the biosynthesis of terpenes in Aspergillus nidulans. These results identify the first Aspergillus nidulans diterpene gene cluster and suggest a biosynthetic pathway for ent-pimara-8(14),15-diene.

  16. Dose-Dependent Change in Elimination Kinetics of Ethanol due to Shift of Dominant Metabolizing Enzyme from ADH 1 (Class I) to ADH 3 (Class III) in Mouse.

    PubMed

    Haseba, Takeshi; Kameyama, Kouji; Mashimo, Keiko; Ohno, Youkichi

    2012-01-01

    ADH 1 and ADH 3 are major two ADH isozymes in the liver, which participate in systemic alcohol metabolism, mainly distributing in parenchymal and in sinusoidal endothelial cells of the liver, respectively. We investigated how these two ADHs contribute to the elimination kinetics of blood ethanol by administering ethanol to mice at various doses, and by measuring liver ADH activity and liver contents of both ADHs. The normalized AUC (AUC/dose) showed a concave increase with an increase in ethanol dose, inversely correlating with β. CL(T) (dose/AUC) linearly correlated with liver ADH activity and also with both the ADH-1 and -3 contents (mg/kg B.W.). When ADH-1 activity was calculated by multiplying ADH-1 content by its V(max⁡)/mg (4.0) and normalized by the ratio of liver ADH activity of each ethanol dose to that of the control, the theoretical ADH-1 activity decreased dose-dependently, correlating with β. On the other hand, the theoretical ADH-3 activity, which was calculated by subtracting ADH-1 activity from liver ADH activity and normalized, increased dose-dependently, correlating with the normalized AUC. These results suggested that the elimination kinetics of blood ethanol in mice was dose-dependently changed, accompanied by a shift of the dominant metabolizing enzyme from ADH 1 to ADH 3.

  17. Dose-Dependent Change in Elimination Kinetics of Ethanol due to Shift of Dominant Metabolizing Enzyme from ADH 1 (Class I) to ADH 3 (Class III) in Mouse

    PubMed Central

    Haseba, Takeshi; Kameyama, Kouji; Mashimo, Keiko; Ohno, Youkichi

    2012-01-01

    ADH 1 and ADH 3 are major two ADH isozymes in the liver, which participate in systemic alcohol metabolism, mainly distributing in parenchymal and in sinusoidal endothelial cells of the liver, respectively. We investigated how these two ADHs contribute to the elimination kinetics of blood ethanol by administering ethanol to mice at various doses, and by measuring liver ADH activity and liver contents of both ADHs. The normalized AUC (AUC/dose) showed a concave increase with an increase in ethanol dose, inversely correlating with β. CLT (dose/AUC) linearly correlated with liver ADH activity and also with both the ADH-1 and -3 contents (mg/kg B.W.). When ADH-1 activity was calculated by multiplying ADH-1 content by its Vmax⁡/mg (4.0) and normalized by the ratio of liver ADH activity of each ethanol dose to that of the control, the theoretical ADH-1 activity decreased dose-dependently, correlating with β. On the other hand, the theoretical ADH-3 activity, which was calculated by subtracting ADH-1 activity from liver ADH activity and normalized, increased dose-dependently, correlating with the normalized AUC. These results suggested that the elimination kinetics of blood ethanol in mice was dose-dependently changed, accompanied by a shift of the dominant metabolizing enzyme from ADH 1 to ADH 3. PMID:22164338

  18. Calcilytic Ameliorates Abnormalities of Mutant Calcium-Sensing Receptor (CaSR) Knock-In Mice Mimicking Autosomal Dominant Hypocalcemia (ADH).

    PubMed

    Dong, Bingzi; Endo, Itsuro; Ohnishi, Yukiyo; Kondo, Takeshi; Hasegawa, Tomoka; Amizuka, Norio; Kiyonari, Hiroshi; Shioi, Go; Abe, Masahiro; Fukumoto, Seiji; Matsumoto, Toshio

    2015-11-01

    Activating mutations of calcium-sensing receptor (CaSR) cause autosomal dominant hypocalcemia (ADH). ADH patients develop hypocalcemia, hyperphosphatemia, and hypercalciuria, similar to the clinical features of hypoparathyroidism. The current treatment of ADH is similar to the other forms of hypoparathyroidism, using active vitamin D3 or parathyroid hormone (PTH). However, these treatments aggravate hypercalciuria and renal calcification. Thus, new therapeutic strategies for ADH are needed. Calcilytics are allosteric antagonists of CaSR, and may be effective for the treatment of ADH caused by activating mutations of CaSR. In order to examine the effect of calcilytic JTT-305/MK-5442 on CaSR harboring activating mutations in the extracellular and transmembrane domains in vitro, we first transfected a mutated CaSR gene into HEK cells. JTT-305/MK-5442 suppressed the hypersensitivity to extracellular Ca(2+) of HEK cells transfected with the CaSR gene with activating mutations in the extracellular and transmembrane domains. We then selected two activating mutations locating in the extracellular (C129S) and transmembrane (A843E) domains, and generated two strains of CaSR knock-in mice to build an ADH mouse model. Both mutant mice mimicked almost all the clinical features of human ADH. JTT-305/MK-5442 treatment in vivo increased urinary cAMP excretion, improved serum and urinary calcium and phosphate levels by stimulating endogenous PTH secretion, and prevented renal calcification. In contrast, PTH(1-34) treatment normalized serum calcium and phosphate but could not reduce hypercalciuria or renal calcification. CaSR knock-in mice exhibited low bone turnover due to the deficiency of PTH, and JTT-305/MK-5442 as well as PTH(1-34) increased bone turnover and bone mineral density (BMD) in these mice. These results demonstrate that calcilytics can reverse almost all the phenotypes of ADH including hypercalciuria and renal calcification, and suggest that calcilytics can become a

  19. Identifying Subspace Gene Clusters from Microarray Data Using Low-Rank Representation

    PubMed Central

    Cui, Yan; Zheng, Chun-Hou; Yang, Jian

    2013-01-01

    Identifying subspace gene clusters from the gene expression data is useful for discovering novel functional gene interactions. In this paper, we propose to use low-rank representation (LRR) to identify the subspace gene clusters from microarray data. LRR seeks the lowest-rank representation among all the candidates that can represent the genes as linear combinations of the bases in the dataset. The clusters can be extracted based on the block diagonal representation matrix obtained using LRR, and they can well capture the intrinsic patterns of genes with similar functions. Meanwhile, the parameter of LRR can balance the effect of noise so that the method is capable of extracting useful information from the data with high level of background noise. Compared with traditional methods, our approach can identify genes with similar functions yet without similar expression profiles. Also, it could assign one gene into different clusters. Moreover, our method is robust to the noise and can identify more biologically relevant gene clusters. When applied to three public datasets, the results show that the LRR based method is superior to existing methods for identifying subspace gene clusters. PMID:23527177

  20. Arrangement of the Clostridium baratii F7 Toxin Gene Cluster with Identification of a σ Factor That Recognizes the Botulinum Toxin Gene Cluster Promoters

    DOE PAGES

    Dover, Nir; Barash, Jason R.; Burke, Julianne N.; Hill, Karen K.; Detter, John C.; Arnon, Stephen S.

    2014-05-22

    Botulinum neurotoxin (BoNT) is the most poisonous substances known and its eight toxin types (A to H) are distinguished by the inability of polyclonal antibodies that neutralize one toxin type to neutralize any of the other seven toxin types. Infant botulism, an intestinal toxemia orphan disease, is the most common form of human botulism in the United States. It results from swallowed spores of Clostridium botulinum (or rarely, neurotoxigenic Clostridium butyricum or Clostridium baratii) that germinate and temporarily colonize the lumen of the large intestine, where, as vegetative cells, they produce botulinum toxin. Botulinum neurotoxin is encoded by the bontmore » gene that is part of a toxin gene cluster that includes several accessory genes. In this paper, we sequenced for the first time the complete botulinum neurotoxin gene cluster of nonproteolytic C. baratii type F7. Like the type E and the nonproteolytic type F6 botulinum toxin gene clusters, the C. baratii type F7 had an orfX toxin gene cluster that lacked the regulatory botR gene which is found in proteolytic C. botulinum strains and codes for an alternative σ factor. In the absence of botR, we identified a putative alternative regulatory gene located upstream of the C. baratii type F7 toxin gene cluster. This putative regulatory gene codes for a predicted σ factor that contains DNA-binding-domain homologues to the DNA-binding domains both of BotR and of other members of the TcdR-related group 5 of the σ70 family that are involved in the regulation of toxin gene expression in clostridia. We showed that this TcdR-related protein in association with RNA polymerase core enzyme specifically binds to the C. baratii type F7 botulinum toxin gene cluster promoters. Finally, this TcdR-related protein may therefore be involved in regulating the expression of the genes of the botulinum toxin gene cluster in neurotoxigenic C. baratii.« less

  1. A recently transferred cluster of bacterial genes in Trichomonas vaginalis - lateral gene transfer and the fate of acquired genes

    PubMed Central

    2014-01-01

    Background Lateral Gene Transfer (LGT) has recently gained recognition as an important contributor to some eukaryote proteomes, but the mechanisms of acquisition and fixation in eukaryotic genomes are still uncertain. A previously defined norm for LGTs in microbial eukaryotes states that the majority are genes involved in metabolism, the LGTs are typically localized one by one, surrounded by vertically inherited genes on the chromosome, and phylogenetics shows that a broad collection of bacterial lineages have contributed to the transferome. Results A unique 34 kbp long fragment with 27 clustered genes (TvLF) of prokaryote origin was identified in the sequenced genome of the protozoan parasite Trichomonas vaginalis. Using a PCR based approach we confirmed the presence of the orthologous fragment in four additional T. vaginalis strains. Detailed sequence analyses unambiguously suggest that TvLF is the result of one single, recent LGT event. The proposed donor is a close relative to the firmicute bacterium Peptoniphilus harei. High nucleotide sequence similarity between T. vaginalis strains, as well as to P. harei, and the absence of homologs in other Trichomonas species, suggests that the transfer event took place after the radiation of the genus Trichomonas. Some genes have undergone pseudogenization and degradation, indicating that they may not be retained in the future. Functional annotations reveal that genes involved in informational processes are particularly prone to degradation. Conclusions We conclude that, although the majority of eukaryote LGTs are single gene occurrences, they may be acquired in clusters of several genes that are subsequently cleansed of evolutionarily less advantageous genes. PMID:24898731

  2. Combination of ADH1B*2/ALDH2*2 polymorphisms alters acetaldehyde-derived DNA damage in the blood of Japanese alcoholics.

    PubMed

    Yukawa, Yoshiyuki; Muto, Manabu; Hori, Kimiko; Nagayoshi, Haruna; Yokoyama, Akira; Chiba, Tsutomu; Matsuda, Tomonari

    2012-09-01

    The acetaldehyde associated with alcoholic beverages is an evident carcinogen for the esophagus. Genetic polymorphisms of the alcohol dehydrogenase 1B (ADH1B) and aldehyde dehydrogenase 2 (ALDH2) genes are associated with the risk of esophageal cancer. However, the exact mechanism via which these genetic polymorphisms affect esophageal carcinogenesis has not been elucidated. ADH1B*2 is involved in overproduction of acetaldehyde due to increased ethanol metabolism into acetaldehyde, and ALDH2*2 is involved in accumulation of acetaldehyde due to the deficiency of acetaldehyde metabolism. Acetaldehyde can interact with DNA and form DNA adducts, resulting in DNA damage. N(2)-ethylidene-2'-deoxyguanosine (N(2)-ethylidene-dG) is the most abundant DNA adduct derived from acetaldehyde. Therefore, we quantified N(2)-ethylidene-dG levels in blood samples from 66 Japanese alcoholic patients using liquid chromatography/electrospray tandem mass spectrometry, and investigated the relationship between N(2)-ethylidene-dG levels and ADH1B and ALDH2 genotypes. The median N(2)-ethylidene-dG levels (25th percentile, 75th percentile) in patients with ADH1B*1/*1 plus ALDH2*1/*1, ADH1B*2 carrier plus ALDH2*1/*1, ADH1B*1/*1 plus ALDH2*1/*2, and ADH1B*2 carrier plus ALDH2*1/*2 were 2.14 (0.97, 2.37)/10(7) bases, 2.38 (1.18, 2.98)/10(7) bases, 5.38 (3.19, 6.52)/10(7) bases, and 21.04 (12.75, 34.80)/10(7) bases, respectively. In the ALDH2*1/*2 group, N(2)-ethylidene-dG levels were significantly higher in ADH1B*2 carriers than in the ADH1B*1/*1 group (P < 0.01). N(2)-ethylidene-dG levels were significantly higher in the ALDH2*1/*2 group than in the ALDH2*1/*1 group, regardless of ADH1B genotype (ADH1B*1/*1, P < 0.05; ADH1B*2 carriers, P < 0.01) N(2)-ethylidene-dG levels in blood DNA of the alcoholics was remarkably higher in individuals with a combination of the ADH1B*2 and ALDH2*2 alleles. These results provide a new perspective on the carcinogenicity of the acetaldehyde associated with

  3. Clustering of Drosophila melanogaster Immune Genes in Interplay with Recombination Rate

    PubMed Central

    Wegner, K. Mathias

    2008-01-01

    Background Gene order in eukaryotic chromosomes is not random and has been linked to coordination of gene expression, chromatin structure and also recombination rate. The evolution of recombination rate is especially relevant for genes involved in immunity because host-parasite co-evolution could select for increased recombination rate (Red Queen hypothesis). To identify patterns left by the intimate interaction between hosts and parasites, I analysed the genomic parameters of the immune genes from 24 gene families/groups of Drosophila melanogaster. Principal Findings Immune genes that directly interact with the pathogen (i.e. recognition and effector genes) clustered in regions of higher recombination rates. Out of these, clustered effector genes were transcribed fastest indicating that transcriptional control might be one major cause for cluster formation. The relative position of clusters to each other, on the other hand, cannot be explained by transcriptional control per se. Drosophila immune genes that show epistatic interactions can be found at an average distance of 15.44±2.98 cM, which is considerably closer than genes that do not interact (30.64±1.95 cM). Conclusions Epistatically interacting genes rarely belong to the same cluster, which supports recent models of optimal recombination rates between interacting genes in antagonistic host-parasite co-evolution. These patterns suggest that formation of local clusters might be a result of transcriptional control, but that in the condensed genome of D. melanogaster relative position of these clusters may be a result of selection for optimal rather than maximal recombination rates between these clusters. PMID:18665272

  4. Applying Robust Directional Similarity based Clustering approach RDSC to classification of gene expression data.

    PubMed

    Li, H X; Wang, Shitong; Xiu, Yu

    2006-06-01

    Despite the fact that the classification of gene expression data from a cDNA microarrays has been extensively studied, nowadays a robust clustering method, which can estimate an appropriate number of clusters and be insensitive to its initialization has not yet been developed. In this work, a novel Robust Clustering approach, RDSC, based on the new Directional Similarity measure is presented. This new approach RDSC, which integrates the Directional Similarity based Clustering Algorithm, DSC, with the Agglomerative Hierarchical Clustering Algorithm, AHC, exhibits its robustness to initialization and its capability to determine the appropriate number of clusters reasonably. RDSC has been successfully employed to both artificial and benchmarking gene expression datasets. Our experimental results demonstrate its distinctive superiority over the conventional method Kmeans and the two typical directional clustering algorithms SPKmeans and moVMF.

  5. High-throughput platform for the discovery of elicitors of silent bacterial gene clusters.

    PubMed

    Seyedsayamdost, Mohammad R

    2014-05-20

    Over the past decade, bacterial genome sequences have revealed an immense reservoir of biosynthetic gene clusters, sets of contiguous genes that have the potential to produce drugs or drug-like molecules. However, the majority of these gene clusters appear to be inactive for unknown reasons prompting terms such as "cryptic" or "silent" to describe them. Because natural products have been a major source of therapeutic molecules, methods that rationally activate these silent clusters would have a profound impact on drug discovery. Herein, a new strategy is outlined for awakening silent gene clusters using small molecule elicitors. In this method, a genetic reporter construct affords a facile read-out for activation of the silent cluster of interest, while high-throughput screening of small molecule libraries provides potential inducers. This approach was applied to two cryptic gene clusters in the pathogenic model Burkholderia thailandensis. The results not only demonstrate a prominent activation of these two clusters, but also reveal that the majority of elicitors are themselves antibiotics, most in common clinical use. Antibiotics, which kill B. thailandensis at high concentrations, act as inducers of secondary metabolism at low concentrations. One of these antibiotics, trimethoprim, served as a global activator of secondary metabolism by inducing at least five biosynthetic pathways. Further application of this strategy promises to uncover the regulatory networks that activate silent gene clusters while at the same time providing access to the vast array of cryptic molecules found in bacteria.

  6. Locus Adh of Drosophila melanogaster under selection for delayed senescence

    SciTech Connect

    Khaustova, N.D.

    1995-05-01

    Dynamics of the Adh activity and frequencies of alleles Adh{sup F} and Adh{sup S} were analyzed under selection for delayed senescence. The experiments were performed on Drosophila melanogaster. Lines Adh{sup S}cn and Adh{sup F}vg and experimental populations cn` and vg`, selected for an increased duration of reproductive period (late oviposition) were used. Analysis of fertility, longevity, viability and resistance to starvation showed that selection for late oviposition resulted in delayed senescence of flies of the experimental populations. Genetic structure of population vg` changed considerably with regard to the Adh locus. This was confirmed by parameters of activity, thermostability, and electrophoretic mobility of the enzyme isolated from flies after 30 generations of selection. Analysis of frequencies of the Adh alleles showed that in both selected populations, which initially had different genetic composition, accumulated allele Adh{sup S}, which encodes the isozyme that is less active but more resistant to inactivation. Genetic mechanism of delayed senescence in Drosophila is assumed to involve selection at vitally important enzyme loci, including Adh. 18 refs., 2 tabs., 4 figs.

  7. Sphingolipids regulate telomere clustering by affecting the transcription of genes involved in telomere homeostasis.

    PubMed

    Ikeda, Atsuko; Muneoka, Tetsuya; Murakami, Suguru; Hirota, Ayaka; Yabuki, Yukari; Karashima, Takefumi; Nakazono, Kota; Tsuruno, Masahiro; Pichler, Harald; Shirahige, Katsuhiko; Kodama, Yukiko; Shimamoto, Toshi; Mizuta, Keiko; Funato, Kouichi

    2015-07-15

    In eukaryotic organisms, including mammals, nematodes and yeasts, the ends of chromosomes, telomeres are clustered at the nuclear periphery. Telomere clustering is assumed to be functionally important because proper organization of chromosomes is necessary for proper genome function and stability. However, the mechanisms and physiological roles of telomere clustering remain poorly understood. In this study, we demonstrate a role for sphingolipids in telomere clustering in the budding yeast Saccharomyces cerevisiae. Because abnormal sphingolipid metabolism causes downregulation of expression levels of genes involved in telomere organization, sphingolipids appear to control telomere clustering at the transcriptional level. In addition, the data presented here provide evidence that telomere clustering is required to protect chromosome ends from DNA-damage checkpoint signaling. As sphingolipids are found in all eukaryotes, we speculate that sphingolipid-based regulation of telomere clustering and the protective role of telomere clusters in maintaining genome stability might be conserved in eukaryotes.

  8. Birth of Four Chimeric Plastid Gene Clusters in Japanese Umbrella Pine.

    PubMed

    Hsu, Chih-Yao; Wu, Chung-Shien; Chaw, Shu-Miaw

    2016-01-01

    Many genes in the plastid genomes (plastomes) of plants are organized as gene clusters, in which genes are co-transcribed, resembling bacterial operons. These plastid operons are highly conserved, even among conifers, whose plastomes are highly rearranged relative to other seed plants. We have determined the complete plastome sequence of Sciadopitys verticillata (Japanese umbrella pine), the sole member of Sciadopityaceae. The Sciadopitys plastome is characterized by extensive inversions, pseudogenization of four tRNA genes after tandem duplications, and a unique pair of 370-bp inverted repeats involved in the formation of isomeric plastomes. We showed that plastomic inversions in Sciadopitys have led to shuffling of the remote conserved operons, resulting in the birth of four chimeric gene clusters. Our data also demonstrated that the relocated genes can be co-transcribed in these chimeric gene clusters. The plastome of Sciadopitys advances our current understanding of how the conifer plastomes have evolved toward increased diversity and complexity. PMID:27269365

  9. Birth of Four Chimeric Plastid Gene Clusters in Japanese Umbrella Pine

    PubMed Central

    Hsu, Chih-Yao; Wu, Chung-Shien; Chaw, Shu-Miaw

    2016-01-01

    Many genes in the plastid genomes (plastomes) of plants are organized as gene clusters, in which genes are co-transcribed, resembling bacterial operons. These plastid operons are highly conserved, even among conifers, whose plastomes are highly rearranged relative to other seed plants. We have determined the complete plastome sequence of Sciadopitys verticillata (Japanese umbrella pine), the sole member of Sciadopityaceae. The Sciadopitys plastome is characterized by extensive inversions, pseudogenization of four tRNA genes after tandem duplications, and a unique pair of 370-bp inverted repeats involved in the formation of isomeric plastomes. We showed that plastomic inversions in Sciadopitys have led to shuffling of the remote conserved operons, resulting in the birth of four chimeric gene clusters. Our data also demonstrated that the relocated genes can be co-transcribed in these chimeric gene clusters. The plastome of Sciadopitys advances our current understanding of how the conifer plastomes have evolved toward increased diversity and complexity. PMID:27269365

  10. Transcriptome Analysis of Aspergillus flavus Reveals veA-Dependent Regulation of Secondary Metabolite Gene Clusters, Including the Novel Aflavarin Cluster

    PubMed Central

    Cary, J. W.; Han, Z.; Yin, Y.; Lohmar, J. M.; Shantappa, S.; Harris-Coward, P. Y.; Mack, B.; Ehrlich, K. C.; Wei, Q.; Arroyo-Manzanares, N.; Uka, V.; Vanhaecke, L.; Bhatnagar, D.; Yu, J.; Nierman, W. C.; Johns, M. A.; Sorensen, D.; Shen, H.; De Saeger, S.; Diana Di Mavungu, J.

    2015-01-01

    The global regulatory veA gene governs development and secondary metabolism in numerous fungal species, including Aspergillus flavus. This is especially relevant since A. flavus infects crops of agricultural importance worldwide, contaminating them with potent mycotoxins. The most well-known are aflatoxins, which are cytotoxic and carcinogenic polyketide compounds. The production of aflatoxins and the expression of genes implicated in the production of these mycotoxins are veA dependent. The genes responsible for the synthesis of aflatoxins are clustered, a signature common for genes involved in fungal secondary metabolism. Studies of the A. flavus genome revealed many gene clusters possibly connected to the synthesis of secondary metabolites. Many of these metabolites are still unknown, or the association between a known metabolite and a particular gene cluster has not yet been established. In the present transcriptome study, we show that veA is necessary for the expression of a large number of genes. Twenty-eight out of the predicted 56 secondary metabolite gene clusters include at least one gene that is differentially expressed depending on presence or absence of veA. One of the clusters under the influence of veA is cluster 39. The absence of veA results in a downregulation of the five genes found within this cluster. Interestingly, our results indicate that the cluster is expressed mainly in sclerotia. Chemical analysis of sclerotial extracts revealed that cluster 39 is responsible for the production of aflavarin. PMID:26209694

  11. A Cluster of Genes Involved in Polysaccharide Biosynthesis from Enterococcus faecalis OG1RF

    PubMed Central

    Xu, Yi; Murray, Barbara E.; Weinstock, George M.

    1998-01-01

    Our previous work identified a cosmid clone containing a 43-kb insert from Enterococcus faecalis OG1RF that produced a nonprotein antigen in Escherichia coli. In the present work, we studied this clone in detail. Periodate treatment of lysates of the clone confirmed that the antigen was carbohydrate in nature. Analysis of DNA sequences and transposon insertion mutants suggested that the insert contained a multicistronic gene cluster. Database comparison showed that the cluster contained genes similar to genes involved in the biosynthesis of dTDP-rhamnose, glycosyltransferases, and ABC transporters involved in the export of sugar polymers from both gram-positive and gram-negative organisms. Insertions in several genes within the cluster abolished the immunoreactivity of the clone. This is the first report on a gene cluster of E. faecalis involved in the biosynthesis of an antigenic polysaccharide. PMID:9712783

  12. The clustering of functionally related genes contributes to CNV-mediated disease

    PubMed Central

    Andrews, Tallulah; Honti, Frantisek; Pfundt, Rolph; de Leeuw, Nicole; Hehir-Kwa, Jayne; Vulto-van Silfhout, Anneke; de Vries, Bert; Webber, Caleb

    2015-01-01

    Clusters of functionally related genes can be disrupted by a single copy number variant (CNV). We demonstrate that the simultaneous disruption of multiple functionally related genes is a frequent and significant characteristic of de novo CNVs in patients with developmental disorders (P = 1 × 10−3). Using three different functional networks, we identified unexpectedly large numbers of functionally related genes within de novo CNVs from two large independent cohorts of individuals with developmental disorders. The presence of multiple functionally related genes was a significant predictor of a CNV's pathogenicity when compared to CNVs from apparently healthy individuals and a better predictor than the presence of known disease or haploinsufficient genes for larger CNVs. The functionally related genes found in the de novo CNVs belonged to 70% of all clusters of functionally related genes found across the genome. De novo CNVs were more likely to affect functional clusters and affect them to a greater extent than benign CNVs (P = 6 × 10−4). Furthermore, such clusters of functionally related genes are phenotypically informative: Different patients possessing CNVs that affect the same cluster of functionally related genes exhibit more similar phenotypes than expected (P < 0.05). The spanning of multiple functionally similar genes by single CNVs contributes substantially to how these variants exert their pathogenic effects. PMID:25887030

  13. Comparative and genetic analyses of the putative Vibrio cholerae lipopolysaccharide core oligosaccharide biosynthesis (wav) gene cluster.

    PubMed

    Nesper, Jutta; Kraiss, Anita; Schild, Stefan; Blass, Julia; Klose, Karl E; Bockemühl, Jochen; Reidl, Joachim

    2002-05-01

    We identified five different putative wav gene cluster types, which are responsible for the synthesis of the core oligosaccharide (OS) region of Vibrio cholerae lipopolysaccharide. Preliminary evidence that the genes encoded by this cluster are involved in core OS biosynthesis came from analysis of the recently released O1 El Tor V. cholerae genome sequence and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of O1 El Tor mutant strains defective in three genes (waaF, waaL, and wavB). Investigations of 38 different V. cholerae strains by Southern blotting, PCR, and sequencing analyses showed that the O1 El Tor wav gene cluster type is prevalent among clinical isolates of different serogroups associated with cholera and environmental O1 strains. In contrast, we found differences in the wav gene contents of 19 unrelated non-O1, non-O139 environmental and human isolates not associated with cholera. These strains contained four new wav gene cluster types that differ from each other in distinct gene loci, providing evidence for horizontal transfer of wav genes and for limited structural diversity of the core OS among V. cholerae isolates. Our results show genetic diversity in the core OS biosynthesis gene cluster and predominance of the type 1 wav gene locus in strains associated with clinical cholera, suggesting that a specific core OS structure could contribute to V. cholerae virulence.

  14. Comparative and Genetic Analyses of the Putative Vibrio cholerae Lipopolysaccharide Core Oligosaccharide Biosynthesis (wav) Gene Cluster

    PubMed Central

    Nesper, Jutta; Kraiß, Anita; Schild, Stefan; Blaβ, Julia; Klose, Karl E.; Bockemühl, Jochen; Reidl, Joachim

    2002-01-01

    We identified five different putative wav gene cluster types, which are responsible for the synthesis of the core oligosaccharide (OS) region of Vibrio cholerae lipopolysaccharide. Preliminary evidence that the genes encoded by this cluster are involved in core OS biosynthesis came from analysis of the recently released O1 El Tor V. cholerae genome sequence and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of O1 El Tor mutant strains defective in three genes (waaF, waaL, and wavB). Investigations of 38 different V. cholerae strains by Southern blotting, PCR, and sequencing analyses showed that the O1 El Tor wav gene cluster type is prevalent among clinical isolates of different serogroups associated with cholera and environmental O1 strains. In contrast, we found differences in the wav gene contents of 19 unrelated non-O1, non-O139 environmental and human isolates not associated with cholera. These strains contained four new wav gene cluster types that differ from each other in distinct gene loci, providing evidence for horizontal transfer of wav genes and for limited structural diversity of the core OS among V. cholerae isolates. Our results show genetic diversity in the core OS biosynthesis gene cluster and predominance of the type 1 wav gene locus in strains associated with clinical cholera, suggesting that a specific core OS structure could contribute to V. cholerae virulence. PMID:11953379

  15. Isolation and Characterization of the Gibberellin Biosynthetic Gene Cluster in Sphaceloma manihoticola▿ †

    PubMed Central

    Bömke, Christiane; Rojas, Maria Cecilia; Gong, Fan; Hedden, Peter; Tudzynski, Bettina

    2008-01-01

    Gibberellins (GAs) are tetracyclic diterpenoid phytohormones that were first identified as secondary metabolites of the fungus Fusarium fujikuroi (teleomorph, Gibberella fujikuroi). GAs were also found in the cassava pathogen Sphaceloma manihoticola, but the spectrum of GAs differed from that in F. fujikuroi. In contrast to F. fujikuroi, the GA biosynthetic pathway has not been studied in detail in S. manihoticola, and none of the GA biosynthetic genes have been cloned from the species. Here, we present the identification of the GA biosynthetic gene cluster from S. manihoticola consisting of five genes encoding a bifunctional ent-copalyl/ent-kaurene synthase (CPS/KS), a pathway-specific geranylgeranyl diphosphate synthase (GGS2), and three cytochrome P450 monooxygenases. The functions of all of the genes were analyzed either by a gene replacement approach or by complementing the corresponding F. fujikuroi mutants. The cluster organization and gene functions are similar to those in F. fujikuroi. However, the two border genes in the Fusarium cluster encoding the GA4 desaturase (DES) and the 13-hydroxylase (P450-3) are absent in the S. manihoticola GA gene cluster, consistent with the spectrum of GAs produced by this fungus. The close similarity between the two GA gene clusters, the identical gene functions, and the conserved intron positions suggest a common evolutionary origin despite the distant relatedness of the two fungi. PMID:18567680

  16. Identification of gene-gene and gene-environment interactions within the fibrinogen gene cluster for fibrinogen levels in three ethnically diverse populations.

    PubMed

    Jeff, Janina M; Brown-Gentry, Kristin; Crawford, Dana C

    2015-01-01

    Elevated levels of plasma fibrinogen are associated with clot formation in the absence of inflammation or injury and is a biomarker for arterial clotting, the leading cause of cardiovascular disease. Fibrinogen levels are heritable with >50% attributed to genetic factors, however little is known about possible genetic modifiers that might explain the missing heritability. The fibrinogen gene cluster is comprised of three genes (FGA, FGB, and FGG) that make up the fibrinogen polypeptide essential for fibrinogen production in the blood. Given the known interaction with these genes, we tested 25 variants in the fibrinogen gene cluster for gene x gene and gene x environment interactions in 620 non-Hispanic blacks, 1,385 non-Hispanic whites, and 664 Mexican Americans from a cross-sectional dataset enriched with environmental data, the Third National Health and Nutrition Examination Survey (NHANES III). Using a multiplicative approach, we added cross product terms (gene x gene or gene x environment) to a linear regression model and declared significance at p < 0.05. We identified 19 unique gene x gene and 13 unique gene x environment interactions that impact fibrinogen levels in at least one population at p < 0.05. Over 90% of the gene x gene interactions identified include a variant in the rate-limiting gene, FGB that is essential for the formation of the fibrinogen polypeptide. We also detected gene x environment interactions with fibrinogen variants and sex, smoking, and body mass index. These findings highlight the potential for the discovery of genetic modifiers for complex phenotypes in multiple populations and give a better understanding of the interaction between genes and/or the environment for fibrinogen levels. The need for more powerful and robust methods to identify genetic modifiers is still warranted. PMID:25592583

  17. IDENTIFICATION OF GENE-GENE AND GENE-ENVIRONMENT INTERACTIONS WITHIN THE FIBRINOGEN GENE CLUSTER FOR FIBRINOGEN LEVELS IN THREE ETHNICALLY DIVERSE POPULATIONS

    PubMed Central

    Jeff, Janina M.; Brown-Gentry, Kristin; Crawford, Dana C.

    2014-01-01

    Elevated levels of plasma fibrinogen are associated with clot formation in the absence of inflammation or injury and is a biomarker for arterial clotting, the leading cause of cardiovascular disease. Fibrinogen levels are heritable with >50% attributed to genetic factors, however little is known about possible genetic modifiers that might explain the missing heritability. The fibrinogen gene cluster is comprised of three genes (FGA, FGB, and FGG) that make up the fibrinogen polypeptide essential for fibrinogen production in the blood. Given the known interaction with these genes, we tested 25 variants in the fibrinogen gene cluster for gene × gene and gene × environment interactions in 620 non-Hispanic blacks, 1,385 non-Hispanic whites, and 664 Mexican Americans from a cross-sectional dataset enriched with environmental data, the Third National Health and Nutrition Examination Survey (NHANES III). Using a multiplicative approach, we added cross product terms (gene × gene or gene × environment) to a linear regression model and declared significance at p < 0.05. We identified 19 unique gene × gene and 13 unique gene × environment interactions that impact fibrinogen levels in at least one population at p <0.05. Over 90% of the gene × gene interactions identified include a variant in the rate-limiting gene, FGB that is essential for the formation of the fibrinogen polypeptide. We also detected gene × environment interactions with fibrinogen variants and sex, smoking, and body mass index. These findings highlight the potential for the discovery of genetic modifiers for complex phenotypes in multiple populations and give a better understanding of the interaction between genes and/or the environment for fibrinogen levels. The need for more powerful and robust methods to identify genetic modifiers is still warranted. PMID:25592583

  18. Identification of gene-gene and gene-environment interactions within the fibrinogen gene cluster for fibrinogen levels in three ethnically diverse populations.

    PubMed

    Jeff, Janina M; Brown-Gentry, Kristin; Crawford, Dana C

    2015-01-01

    Elevated levels of plasma fibrinogen are associated with clot formation in the absence of inflammation or injury and is a biomarker for arterial clotting, the leading cause of cardiovascular disease. Fibrinogen levels are heritable with >50% attributed to genetic factors, however little is known about possible genetic modifiers that might explain the missing heritability. The fibrinogen gene cluster is comprised of three genes (FGA, FGB, and FGG) that make up the fibrinogen polypeptide essential for fibrinogen production in the blood. Given the known interaction with these genes, we tested 25 variants in the fibrinogen gene cluster for gene x gene and gene x environment interactions in 620 non-Hispanic blacks, 1,385 non-Hispanic whites, and 664 Mexican Americans from a cross-sectional dataset enriched with environmental data, the Third National Health and Nutrition Examination Survey (NHANES III). Using a multiplicative approach, we added cross product terms (gene x gene or gene x environment) to a linear regression model and declared significance at p < 0.05. We identified 19 unique gene x gene and 13 unique gene x environment interactions that impact fibrinogen levels in at least one population at p < 0.05. Over 90% of the gene x gene interactions identified include a variant in the rate-limiting gene, FGB that is essential for the formation of the fibrinogen polypeptide. We also detected gene x environment interactions with fibrinogen variants and sex, smoking, and body mass index. These findings highlight the potential for the discovery of genetic modifiers for complex phenotypes in multiple populations and give a better understanding of the interaction between genes and/or the environment for fibrinogen levels. The need for more powerful and robust methods to identify genetic modifiers is still warranted.

  19. Hox gene clusters of early vertebrates: do they serve as reliable markers for genome evolution?

    PubMed

    Kuraku, Shigehiro

    2011-06-01

    Hox genes, responsible for regional specification along the anteroposterior axis in embryogenesis, are found as clusters in most eumetazoan genomes sequenced to date. Invertebrates possess a single Hox gene cluster with some exceptions of secondary cluster breakages, while osteichthyans (bony vertebrates) have multiple Hox clusters. In tetrapods, four Hox clusters, derived from the so-called two-round whole genome duplications (2R-WGDs), are observed. Overall, the number of Hox gene clusters has been regarded as a reliable marker of ploidy levels in animal genomes. In fact, this scheme also fits the situations in teleost fishes that experienced an additional WGD. In this review, I focus on cyclostomes and cartilaginous fishes as lineages that would fill the gap between invertebrates and osteichthyans. A recent study highlighted a possible loss of the HoxC cluster in the galeomorph shark lineage, while other aspects of cartilaginous fish Hox clusters usually mark their conserved nature. In contrast, existing resources suggest that the cyclostomes exhibit a different mode of Hox cluster organization. For this group of species, whose genomes could have differently responded to the 2R-WGDs from jawed vertebrates, therefore the number of Hox clusters may not serve as a good indicator of their ploidy level. PMID:21802046

  20. Cloning and Heterologous Expression of the Thioviridamide Biosynthesis Gene Cluster from Streptomyces olivoviridis

    PubMed Central

    Izawa, Masumi; Kawasaki, Takashi

    2013-01-01

    Thioviridamide is a unique peptide antibiotic containing five thioamide bonds from Streptomyces olivoviridis. Draft genome sequencing revealed a gene (the tvaA gene) encoding the thioviridamide precursor peptide. The thioviridamide biosynthesis gene cluster was identified by heterologous production of thioviridamide in Streptomyces lividans. PMID:23995943

  1. Leveraging long sequencing reads to investigate R-gene clustering and variation in sugar beet

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Host-pathogen interactions are of prime importance to modern agriculture. Plants utilize various types of resistance genes to mitigate pathogen damage. Identification of the specific gene responsible for a specific resistance can be difficult due to duplication and clustering within R-gene families....

  2. Minisatellite instability at the Adh locus reveals somatic polymorphism in amphioxus

    PubMed Central

    Cañestro, Cristian; Gonzàlez-Duarte, Roser; Albalat, Ricard

    2002-01-01

    Amphioxus (subphylum Cephalochordata) is the closest living relative to vertebrates and widely used for phylogenetic analyses of vertebrate gene evolution. Amphioxus genes are highly polymorphic, but the origin and nature of this variability is unknown. We have analyzed the alcohol dehydrogenase locus (Adh3) in two amphioxus species (Branchiostoma lanceolatum and Branchiostoma floridae) and found that genetic variation is related to repetitive DNA sequences, mainly minisatellites. Small pool-PCR assays indicated that allelic variants are generated by minisatellite instability. We conclude that the generation of new forms was not preferentially linked to germline processes but rather to somatic events leading to mosaic adult animals. Furthermore, most Adh minisatellites belong to a novel class, which we have named mirages. Their distinctive feature is that the repeat subunit spans the exon–intron boundaries and generates potential duplications of the splice sites. However, splicing may not be compromised as no aberrant mRNA variants were detected. PMID:12087171

  3. Prediction of operon-like gene clusters in the Arabidopsis thaliana genome based on co-expression analysis of neighboring genes.

    PubMed

    Wada, Masayoshi; Takahashi, Hiroki; Altaf-Ul-Amin, Md; Nakamura, Kensuke; Hirai, Masami Y; Ohta, Daisaku; Kanaya, Shigehiko

    2012-07-15

    Operon-like arrangements of genes occur in eukaryotes ranging from yeasts and filamentous fungi to nematodes, plants, and mammals. In plants, several examples of operon-like gene clusters involved in metabolic pathways have recently been characterized, e.g. the cyclic hydroxamic acid pathways in maize, the avenacin biosynthesis gene clusters in oat, the thalianol pathway in Arabidopsis thaliana, and the diterpenoid momilactone cluster in rice. Such operon-like gene clusters are defined by their co-regulation or neighboring positions within immediate vicinity of chromosomal regions. A comprehensive analysis of the expression of neighboring genes therefore accounts a crucial step to reveal the complete set of operon-like gene clusters within a genome. Genome-wide prediction of operon-like gene clusters should contribute to functional annotation efforts and provide novel insight into evolutionary aspects acquiring certain biological functions as well. We predicted co-expressed gene clusters by comparing the Pearson correlation coefficient of neighboring genes and randomly selected gene pairs, based on a statistical method that takes false discovery rate (FDR) into consideration for 1469 microarray gene expression datasets of A. thaliana. We estimated that A. thaliana contains 100 operon-like gene clusters in total. We predicted 34 statistically significant gene clusters consisting of 3 to 22 genes each, based on a stringent FDR threshold of 0.1. Functional relationships among genes in individual clusters were estimated by sequence similarity and functional annotation of genes. Duplicated gene pairs (determined based on BLAST with a cutoff of E<10(-5)) are included in 27 clusters. Five clusters are associated with metabolism, containing P450 genes restricted to the Brassica family and predicted to be involved in secondary metabolism. Operon-like clusters tend to include genes encoding bio-machinery associated with ribosomes, the ubiquitin/proteasome system, secondary

  4. Influence of microarrays experiments missing values on the stability of gene groups by hierarchical clustering

    PubMed Central

    de Brevern, Alexandre G; Hazout, Serge; Malpertuy, Alain

    2004-01-01

    Background Microarray technologies produced large amount of data. The hierarchical clustering is commonly used to identify clusters of co-expressed genes. However, microarray datasets often contain missing values (MVs) representing a major drawback for the use of the clustering methods. Usually the MVs are not treated, or replaced by zero or estimated by the k-Nearest Neighbor (kNN) approach. The topic of the paper is to study the stability of gene clusters, defined by various hierarchical clustering algorithms, of microarrays experiments including or not MVs. Results In this study, we show that the MVs have important effects on the stability of the gene clusters. Moreover, the magnitude of the gene misallocations is depending on the aggregation algorithm. The most appropriate aggregation methods (e.g. complete-linkage and Ward) are highly sensitive to MVs, and surprisingly, for a very tiny proportion of MVs (e.g. 1%). In most of the case, the MVs must be replaced by expected values. The MVs replacement by the kNN approach clearly improves the identification of co-expressed gene clusters. Nevertheless, we observe that kNN approach is less suitable for the extreme values of gene expression. Conclusion The presence of MVs (even at a low rate) is a major factor of gene cluster instability. In addition, the impact depends on the hierarchical clustering algorithm used. Some methods should be used carefully. Nevertheless, the kNN approach constitutes one efficient method for restoring the missing expression gene values, with a low error level. Our study highlights the need of statistical treatments in microarray data to avoid misinterpretation. PMID:15324460

  5. A gene cluster for amylovoran synthesis in Erwinia amylovora: characterization and relationship to cps genes in Erwinia stewartii.

    PubMed

    Bernhard, F; Coplin, D L; Geider, K

    1993-05-01

    A large ams gene cluster required for production of the acidic extracellular polysaccharide (EPS) amylovoran by the fire blight pathogen Erwinia amylovora was cloned. Tn5 mutagenesis and gene replacement were used to construct chromosomal ams mutants. Five complementation groups, essential for amylovoran synthesis and virulence in E. amylovora, were identified and designated ams A-E. The ams gene cluster is about 7 kb in size and functionally equivalent to the cps gene cluster involved in EPS synthesis by the related pathogen Erwinia stewartii. Mucoidy and virulence were restored to E. stewartii mutants in four cps complementation groups by the cloned E. amylovora ams genes. Conversely, the E. stewartii cps gene cluster was able to complement mutations in E. amylovora ams genes. Correspondence was found between the amsA-E complementation groups and the cpsB-D region, but the arrangement of the genes appears to be different. EPS production and virulence were also restored to E. amylovora amsE and E. stewartii cpsD mutants by clones containing the Rhizobium meliloti exo A gene.

  6. Epigenetic regulation of the RHOX homeobox gene cluster and its association with human male infertility

    PubMed Central

    Richardson, Marcy E.; Bleiziffer, Andreas; Tüttelmann, Frank; Gromoll, Jörg; Wilkinson, Miles F.

    2014-01-01

    The X-linked RHOX cluster encodes a set of homeobox genes that are selectively expressed in the reproductive tract. Members of the RHOX cluster regulate target genes important for spermatogenesis promote male fertility in mice. Studies show that demethylating agents strongly upregulate the expression of mouse Rhox genes, suggesting that they are regulated by DNA methylation. However, whether this extends to human RHOX genes, whether DNA methylation directly regulates RHOX gene transcription and how this relates to human male infertility are unknown. To address these issues, we first defined the promoter regions of human RHOX genes and performed gain- and loss-of-function experiments to determine whether human RHOX gene transcription is regulated by DNA methylation. Our results indicated that DNA methylation is necessary and sufficient to silence human RHOX gene expression. To determine whether RHOX cluster methylation associates with male infertility, we evaluated the methylation status of RHOX genes in sperm from a large cohort of infertility patients. Linear regression analysis revealed a strong association between RHOX gene cluster hypermethylation and three independent types of semen abnormalities. Hypermethylation was restricted specifically to the RHOX cluster; we did not observe it in genes immediately adjacent to it on the X chromosome. Our results strongly suggest that human RHOX homeobox genes are under an epigenetic control mechanism that is aberrantly regulated in infertility patients. We propose that hypermethylation of the RHOX gene cluster serves as a marker for idiopathic infertility and that it is a candidate to exert a causal role in male infertility. PMID:23943794

  7. A cross-species bi-clustering approach to identifying conserved co-regulated genes

    PubMed Central

    Sun, Jiangwen; Jiang, Zongliang; Tian, Xiuchun; Bi, Jinbo

    2016-01-01

    Motivation: A growing number of studies have explored the process of pre-implantation embryonic development of multiple mammalian species. However, the conservation and variation among different species in their developmental programming are poorly defined due to the lack of effective computational methods for detecting co-regularized genes that are conserved across species. The most sophisticated method to date for identifying conserved co-regulated genes is a two-step approach. This approach first identifies gene clusters for each species by a cluster analysis of gene expression data, and subsequently computes the overlaps of clusters identified from different species to reveal common subgroups. This approach is ineffective to deal with the noise in the expression data introduced by the complicated procedures in quantifying gene expression. Furthermore, due to the sequential nature of the approach, the gene clusters identified in the first step may have little overlap among different species in the second step, thus difficult to detect conserved co-regulated genes. Results: We propose a cross-species bi-clustering approach which first denoises the gene expression data of each species into a data matrix. The rows of the data matrices of different species represent the same set of genes that are characterized by their expression patterns over the developmental stages of each species as columns. A novel bi-clustering method is then developed to cluster genes into subgroups by a joint sparse rank-one factorization of all the data matrices. This method decomposes a data matrix into a product of a column vector and a row vector where the column vector is a consistent indicator across the matrices (species) to identify the same gene cluster and the row vector specifies for each species the developmental stages that the clustered genes co-regulate. Efficient optimization algorithm has been developed with convergence analysis. This approach was first validated on

  8. Molecular evolution of Adh and LEAFY and the phylogenetic utility of their introns in Pyrus (Rosaceae)

    PubMed Central

    2011-01-01

    Background The genus Pyrus belongs to the tribe Pyreae (the former subfamily Maloideae) of the family Rosaceae, and includes one of the most important commercial fruit crops, pear. The phylogeny of Pyrus has not been definitively reconstructed. In our previous efforts, the internal transcribed spacer region (ITS) revealed a poorly resolved phylogeny due to non-concerted evolution of nrDNA arrays. Therefore, introns of low copy nuclear genes (LCNG) are explored here for improved resolution. However, paralogs and lineage sorting are still two challenges for applying LCNGs in phylogenetic studies, and at least two independent nuclear loci should be compared. In this work the second intron of LEAFY and the alcohol dehydrogenase gene (Adh) were selected to investigate their molecular evolution and phylogenetic utility. Results DNA sequence analyses revealed a complex ortholog and paralog structure of Adh genes in Pyrus and Malus, the pears and apples. Comparisons between sequences from RT-PCR and genomic PCR indicate that some Adh homologs are putatively nonfunctional. A partial region of Adh1 was sequenced for 18 Pyrus species and three subparalogs representing Adh1-1 were identified. These led to poorly resolved phylogenies due to low sequence divergence and the inclusion of putative recombinants. For the second intron of LEAFY, multiple inparalogs were discovered for both LFY1int2 and LFY2int2. LFY1int2 is inadequate for phylogenetic analysis due to lineage sorting of two inparalogs. LFY2int2-N, however, showed a relatively high sequence divergence and led to the best-resolved phylogeny. This study documents the coexistence of outparalogs and inparalogs, and lineage sorting of these paralogs and orthologous copies. It reveals putative recombinants that can lead to incorrect phylogenetic inferences, and presents an improved phylogenetic resolution of Pyrus using LFY2int2-N. Conclusions Our study represents the first phylogenetic analyses based on LCNGs in Pyrus

  9. An adaptive strategy for single- and multi-cluster gene assignment.

    PubMed

    Garg, Sanjeev; Hansen, Marc F; Rowe, David W; Achenie, Luke E K

    2003-01-01

    Strict assignment of genes to one class, dimensionality reduction, a priori specification of the number of classes, the need for a training set, nonunique solution, and complex learning mechanisms are some of the inadequacies of current clustering algorithms. Existing algorithms cluster genes on the basis of high positive correlations between their expression patterns. However, genes with strong negative correlations can also have similar functions and are most likely to have a role in the same pathways. To address some of these issues, we propose the adaptive centroid algorithm (ACA), which employs an analysis of variance (ANOVA)-based performance criterion. The ACA also uses Euclidian distances, the center-of-mass principle for heterogeneously distributed mass elements, and the given data set to give unique solutions. The proposed approach involves three stages. In the first stage a two-way ANOVA of the gene expression matrix is performed. The two factors in the ANOVA are gene expression and experimental condition. The residual mean squared error (MSE) from the ANOVA is used as a performance criterion in the ACA. Finally, correlated clusters are found based on the Pearson correlation coefficients. To validate the proposed approach, a two-way ANOVA is again performed on the discovered clusters. The results from this last step indicate that MSEs of the clusters are significantly lower compared to that of the fibroblast-serum gene expression matrix. The ACA is employed in this study for single- as well as multi-cluster gene assignments.

  10. Identification of Nocobactin NA Biosynthetic Gene Clusters in Nocardia farcinica▿ §

    PubMed Central

    Hoshino, Yasutaka; Chiba, Kazuhiro; Ishino, Keiko; Fukai, Toshio; Igarashi, Yasuhiro; Yazawa, Katsukiyo; Mikami, Yuzuru; Ishikawa, Jun

    2011-01-01

    We identified the biosynthetic gene clusters of the siderophore nocobactin NA. The nbt clusters, which were discovered as genes highly homologous to the mycobactin biosynthesis genes by the genomic sequencing of Nocardia farcinica IFM 10152, consist of 10 genes separately located at two genomic regions. The gene organization of the nbt clusters and the predicted functions of the nbt genes, particularly the cyclization and epimerization domains, were in good agreement with the chemical structure of nocobactin NA. Disruptions of the nbtA and nbtE genes, respectively, reduced and abolished the productivity of nocobactin NA. The heterologous expression of the nbtS gene revealed that this gene encoded a salicylate synthase. These results indicate that the nbt clusters are responsible for the biosynthesis of nocobactin NA. We also found putative IdeR-binding sequences upstream of the nbtA, -G, -H, -S, and -T genes, whose expression was more than 10-fold higher in the low-iron condition than in the high-iron condition. These results suggest that nbt genes are regulated coordinately by IdeR protein in an iron-dependent manner. The ΔnbtE mutant was found to be impaired in cytotoxicity against J774A.1 cells, suggesting that nocobactin NA production is required for virulence of N. farcinica. PMID:21097631

  11. The nonribosomal peptide and polyketide synthetic gene clusters in two strains of entomopathogenic fungi in Cordyceps.

    PubMed

    Wang, Wen-Jing; Vogel, Heiko; Yao, Yi-Jian; Ping, Liyan

    2012-11-01

    Species of Cordyceps Fr. are entomopathogenic fungi that parasitize the larvae or pupae of lepidopteran insects. The secondary metabolites, nonribosomal peptides and polyketides are well-known mediators of pathogenesis. The biosynthetic gene clusters of these compounds in two fungal strains (1630 and DSM 1153) formerly known as Cordyceps militaris were screened using polymerase chain reaction with degenerate primers. Two nonribosomal peptide synthetase genes, one polyketide synthetase gene and one hybrid gene cluster were identified, and certain characteristics of the structures of their potential products were predicted. All four genes were actively expressed under laboratory conditions but at markedly different levels. The gene clusters from the two fungal strains were structurally and functionally unrelated, suggesting different evolutionary origins and physiological functions. Phylogenetic and biochemical analyses confirmed that the two fungal strains are not conspecific as currently assigned. PMID:22889355

  12. Identification of a recent recombination event within the human beta-globin gene cluster.

    PubMed Central

    Gerhard, D S; Kidd, K K; Kidd, J R; Egeland, J A; Housman, D E

    1984-01-01

    In a detailed study of inheritance of DNA sequence polymorphism in a large reference pedigree, an individual was identified with an apparent genetic recombination event within the human beta-globin gene cluster. Analysis of the haplotypes of relevant individuals within this pedigree suggested that the meiotic crossing-over event is likely to have occurred within a 19.8-kilobase-pair region of the beta-globin gene cluster. Analysis of other DNA markers closely linked to the beta-globin gene cluster--segment 12 of chromosome 11 (D11S12) and loci for insulin, the cellular oncogene c-Ha-ras, and preproparathyroid hormone--confirmed that a crossover event must have occurred within the region of chromosome 11 between D11S12 and the beta-globin gene cluster. It is suggested that the event observed has occurred within a DNA region compatible with recombinational "hot spots" suggested by population studies. PMID:6096866

  13. Neuronal plasticity after spinal cord injury: identification of a gene cluster driving neurite outgrowth.

    PubMed

    Di Giovanni, Simone; Faden, Alan I; Yakovlev, Alexander; Duke-Cohan, Jonathan S; Finn, Tom; Thouin, Melissa; Knoblach, Susan; De Biase, Andrea; Bregman, Barbara S; Hoffman, Eric P

    2005-01-01

    Functional recovery after spinal cord injury (SCI) may result in part from axon outgrowth and related plasticity through coordinated changes at the molecular level. We employed microarray analysis to identify a subset of genes the expression patterns of which were temporally coregulated and correlated to functional recovery after SCI. Steady-state mRNA levels of this synchronously regulated gene cluster were depressed in both ventral and dorsal horn neurons within 24 h after injury, followed by strong re-induction during the following 2 wk, which paralleled functional recovery. The identified cluster includes neuritin, attractin, microtubule-associated protein 1a, and myelin oligodendrocyte protein genes. Transcriptional and protein regulation of this novel gene cluster was also evaluated in spinal cord tissue and in single neurons and was shown to play a role in axonal plasticity. Finally, in vitro transfection experiments in primary dorsal root ganglion cells showed that cluster members act synergistically to drive neurite outgrowth. PMID:15522907

  14. Characterization of the fumonisin B2 biosynthetic gene cluster in Aspergillus niger and A. awamori.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus niger and A. awamori strains isolated from grapes cultivated in Mediterranean basin were examined for fumonisin B2 (FB2) production and presence/absence of sequences within the fumonisin biosynthetic gene (fum) cluster. Presence of 13 regions in the fum cluster was evaluated by PCR assay...

  15. Effects of endogenous antidiuretic hormone (ADH) on macrophage phagocytosis

    SciTech Connect

    Fernandez-Repollet, E.; Opava-Stitzer, S.; Tiffany, S.; Schwartz, A.

    1983-07-01

    Although several studies have indicated that antidiuretic hormone (ADH) enhances the phagocytic function of the reticuloendothelial system (RES) in shock syndromes, it remains unknown what influence ADH exerts upon the individual phagocytic components of this system. The present investigation was designed to evaluate the effects of endogenous ADH on the phagocytic activity of peritoneal macrophage cells. As a phagocytic stimuli, fluorescent methacrylate microbeads were injected intraperitoneally into Brattleboro (ADH deficient) and normal Long Evans rats in the presence and absence of exogenous ADH. Peritoneal cells were harvested 19-22 hr after the administration of the microbeads and the percent phagocytosis was determined in macrophage cells using a fluorescence-activated cell sorter (FACS II). Our results indicate that the percentage of peritoneal macrophages ingesting the fluorescent methacrylate microbeads was significantly reduced in the absence of ADH (Brattleboro rats: 5.4 +/- 0.6% versus Long Evans rats: 16.8 +/- 2.3%; p less than 0.001). In addition, our data demonstrate that exogenous administration of ADH significantly enhanced macrophage phagocytosis in Brattleboro (14.7 +/- 2.2%) and normal Long Evans (49.6 +/- 4.5%) rats. These data suggest, for the first time, that endogenous ADH might play a modulatory role in the phagocytic activity of a specific component of the RES, namely, the macrophage cell.

  16. Clusters of antibiotic resistance genes enriched together stay together in swine agriculture

    DOE PAGES

    Johnson, Timothy A.; Stedtfeld, Robert D.; Wang, Qiong; Cole, James R.; Hashsham, Syed A.; Looft, Torey; Zhu, Yong -Guan; Tiedje, James M.

    2016-04-12

    Antibiotic resistance is a worldwide health risk, but the influence of animal agriculture on the genetic context and enrichment of individual antibiotic resistance alleles remains unclear. Using quantitative PCR followed by amplicon sequencing, we quantified and sequenced 44 genes related to antibiotic resistance, mobile genetic elements, and bacterial phylogeny in microbiomes from U.S. laboratory swine and from swine farms from three Chinese regions. We identified highly abundant resistance clusters: groups of resistance and mobile genetic element alleles that cooccur. For example, the abundance of genes conferring resistance to six classes of antibiotics together with class 1 integrase and the abundancemore » of IS6100-type transposons in three Chinese regions are directly correlated. These resistance cluster genes likely colocalize in microbial genomes in the farms. Resistance cluster alleles were dramatically enriched (up to 1 to 10% as abundant as 16S rRNA) and indicate that multidrug-resistant bacteria are likely the norm rather than an exception in these communities. This enrichment largely occurred independently of phylogenetic composition; thus, resistance clusters are likely present in many bacterial taxa. Furthermore, resistance clusters contain resistance genes that confer resistance to antibiotics independently of their particular use on the farms. Selection for these clusters is likely due to the use of only a subset of the broad range of chemicals to which the clusters confer resistance. The scale of animal agriculture and its wastes, the enrichment and horizontal gene transfer potential of the clusters, and the vicinity of large human populations suggest that managing this resistance reservoir is important for minimizing human risk.Agricultural antibiotic use results in clusters of cooccurring resistance genes that together confer resistance to multiple antibiotics. The use of a single antibiotic could select for an entire suite of resistance

  17. The Biosynthetic Gene Cluster of Zorbamycin, a Member of the Bleomycin Family of Antitumor Antibiotics, from Streptomyces flavoviridis ATCC 21892

    PubMed Central

    Galm, Ute; Wendt-Pienkowski, Evelyn; Wang, Liyan; George, Nicholas P.; Oh, Tae-Jin; Yi, Fan; Tao, Meifeng; Coughlin, Jane M.; Shen, Ben

    2011-01-01

    The biosynthetic gene cluster for the glycopeptide-derived antitumor antibiotic zorbamycin (ZBM) was cloned by screening a cosmid library of Streptomyces flavoviridis ATCC 21892. Sequence analysis revealed 40 ORFs belonging to the ZBM biosynthetic gene cluster. However, only 23 and 22 ORFs showed striking similarities to the biosynthetic gene clusters for the bleomycins (BLMs) and tallysomycins (TLMs), respectively; the remaining ORFs do not show significant homology to ORFs from the related BLM and TLM clusters. The ZBM gene cluster consists of 16 nonribosomal peptide synthetase (NRPS) genes encoding eight complete NRPS modules, three incomplete didomain NRPS modules, and eight freestanding single NRPS domains or associated enzymes, a polyketide synthase (PKS) gene encoding one PKS module, six sugar biosynthesis genes, as well as genes encoding other biosynthesis and resistance proteins. A genetic system using Escherichia coli-Streptomyces flavoviridis intergeneric conjugation was developed to enable ZBM gene cluster boundary determinations and biosynthetic pathway manipulations. PMID:19081934

  18. Integrating Data Clustering and Visualization for the Analysis of 3D Gene Expression Data

    SciTech Connect

    Data Analysis and Visualization and the Department of Computer Science, University of California, Davis, One Shields Avenue, Davis CA 95616, USA,; nternational Research Training Group ``Visualization of Large and Unstructured Data Sets,'' University of Kaiserslautern, Germany; Computational Research Division, Lawrence Berkeley National Laboratory, One Cyclotron Road, Berkeley, CA 94720, USA; Genomics Division, Lawrence Berkeley National Laboratory, One Cyclotron Road, Berkeley CA 94720, USA; Life Sciences Division, Lawrence Berkeley National Laboratory, One Cyclotron Road, Berkeley CA 94720, USA,; Computer Science Division,University of California, Berkeley, CA, USA,; Computer Science Department, University of California, Irvine, CA, USA,; All authors are with the Berkeley Drosophila Transcription Network Project, Lawrence Berkeley National Laboratory,; Rubel, Oliver; Weber, Gunther H.; Huang, Min-Yu; Bethel, E. Wes; Biggin, Mark D.; Fowlkes, Charless C.; Hendriks, Cris L. Luengo; Keranen, Soile V. E.; Eisen, Michael B.; Knowles, David W.; Malik, Jitendra; Hagen, Hans; Hamann, Bernd

    2008-05-12

    The recent development of methods for extracting precise measurements of spatial gene expression patterns from three-dimensional (3D) image data opens the way for new analyses of the complex gene regulatory networks controlling animal development. We present an integrated visualization and analysis framework that supports user-guided data clustering to aid exploration of these new complex datasets. The interplay of data visualization and clustering-based data classification leads to improved visualization and enables a more detailed analysis than previously possible. We discuss (i) integration of data clustering and visualization into one framework; (ii) application of data clustering to 3D gene expression data; (iii) evaluation of the number of clusters k in the context of 3D gene expression clustering; and (iv) improvement of overall analysis quality via dedicated post-processing of clustering results based on visualization. We discuss the use of this framework to objectively define spatial pattern boundaries and temporal profiles of genes and to analyze how mRNA patterns are controlled by their regulatory transcription factors.

  19. Regulation of transcription of cell division genes in the Escherichia coli dcw cluster.

    PubMed

    Vicente, M; Gomez, M J; Ayala, J A

    1998-04-01

    The Escherichia coli dcw cluster contains cell division genes, such as the phylogenetically ubiquitous ftsZ, and genes involved in peptidoglycan synthesis. Transcription in the cluster proceeds in the same direction as the progress of the replication fork along the chromosome. Regulation is exerted at the transcriptional and post-transcriptional levels. The absence of transcriptional termination signals may, in principle, allow extension of the transcripts initiated at the up-stream promoter (mraZ1p) even to the furthest down-stream gene (envA). Complementation tests suggest that they extend into ftsW in the central part of the cluster. In addition, the cluster contains other promoters individually regulated by cis- and trans-acting signals. Dissociation of the expression of the ftsZ gene, located after ftsQ and A near the 3' end of the cluster, from its natural regulatory signals leads to an alteration in the physiology of cell division. The complexities observed in the regulation of gene expression in the cluster may then have an important biological role. Among them, LexA-binding SOS boxes have been found at the 5' end of the cluster, preceding promoters which direct the expression of ftsI (coding for PBP3, the penicillin-binding protein involved in septum formation). A gearbox promoter, ftsQ1p, forms part of the signals regulating the transcription of ftsQ, A and Z. It is an inversely growth-dependent mechanism driven by RNA polymerase containing sigma s, the factor involved in the expression of stationary phase-specific genes. Although the dcw cluster is conserved to a different extent in a variety of bacteria, the regulation of gene expression, the presence or absence of individual genes, and even the essentiality of some of them, show variations in the phylogenetic scale which may reflect adaptation to specific life cycles.

  20. A Stationary Wavelet Entropy-Based Clustering Approach Accurately Predicts Gene Expression

    PubMed Central

    Nguyen, Nha; Vo, An; Choi, Inchan

    2015-01-01

    Abstract Studying epigenetic landscapes is important to understand the condition for gene regulation. Clustering is a useful approach to study epigenetic landscapes by grouping genes based on their epigenetic conditions. However, classical clustering approaches that often use a representative value of the signals in a fixed-sized window do not fully use the information written in the epigenetic landscapes. Clustering approaches to maximize the information of the epigenetic signals are necessary for better understanding gene regulatory environments. For effective clustering of multidimensional epigenetic signals, we developed a method called Dewer, which uses the entropy of stationary wavelet of epigenetic signals inside enriched regions for gene clustering. Interestingly, the gene expression levels were highly correlated with the entropy levels of epigenetic signals. Dewer separates genes better than a window-based approach in the assessment using gene expression and achieved a correlation coefficient above 0.9 without using any training procedure. Our results show that the changes of the epigenetic signals are useful to study gene regulation. PMID:25383910

  1. Yeast homologous recombination-based promoter engineering for the activation of silent natural product biosynthetic gene clusters.

    PubMed

    Montiel, Daniel; Kang, Hahk-Soo; Chang, Fang-Yuan; Charlop-Powers, Zachary; Brady, Sean F

    2015-07-21

    Large-scale sequencing of prokaryotic (meta)genomic DNA suggests that most bacterial natural product gene clusters are not expressed under common laboratory culture conditions. Silent gene clusters represent a promising resource for natural product discovery and the development of a new generation of therapeutics. Unfortunately, the characterization of molecules encoded by these clusters is hampered owing to our inability to express these gene clusters in the laboratory. To address this bottleneck, we have developed a promoter-engineering platform to transcriptionally activate silent gene clusters in a model heterologous host. Our approach uses yeast homologous recombination, an auxotrophy complementation-based yeast selection system and sequence orthogonal promoter cassettes to exchange all native promoters in silent gene clusters with constitutively active promoters. As part of this platform, we constructed and validated a set of bidirectional promoter cassettes consisting of orthogonal promoter sequences, Streptomyces ribosome binding sites, and yeast selectable marker genes. Using these tools we demonstrate the ability to simultaneously insert multiple promoter cassettes into a gene cluster, thereby expediting the reengineering process. We apply this method to model active and silent gene clusters (rebeccamycin and tetarimycin) and to the silent, cryptic pseudogene-containing, environmental DNA-derived Lzr gene cluster. Complete promoter refactoring and targeted gene exchange in this "dead" cluster led to the discovery of potent indolotryptoline antiproliferative agents, lazarimides A and B. This potentially scalable and cost-effective promoter reengineering platform should streamline the discovery of natural products from silent natural product biosynthetic gene clusters. PMID:26150486

  2. Yeast homologous recombination-based promoter engineering for the activation of silent natural product biosynthetic gene clusters.

    PubMed

    Montiel, Daniel; Kang, Hahk-Soo; Chang, Fang-Yuan; Charlop-Powers, Zachary; Brady, Sean F

    2015-07-21

    Large-scale sequencing of prokaryotic (meta)genomic DNA suggests that most bacterial natural product gene clusters are not expressed under common laboratory culture conditions. Silent gene clusters represent a promising resource for natural product discovery and the development of a new generation of therapeutics. Unfortunately, the characterization of molecules encoded by these clusters is hampered owing to our inability to express these gene clusters in the laboratory. To address this bottleneck, we have developed a promoter-engineering platform to transcriptionally activate silent gene clusters in a model heterologous host. Our approach uses yeast homologous recombination, an auxotrophy complementation-based yeast selection system and sequence orthogonal promoter cassettes to exchange all native promoters in silent gene clusters with constitutively active promoters. As part of this platform, we constructed and validated a set of bidirectional promoter cassettes consisting of orthogonal promoter sequences, Streptomyces ribosome binding sites, and yeast selectable marker genes. Using these tools we demonstrate the ability to simultaneously insert multiple promoter cassettes into a gene cluster, thereby expediting the reengineering process. We apply this method to model active and silent gene clusters (rebeccamycin and tetarimycin) and to the silent, cryptic pseudogene-containing, environmental DNA-derived Lzr gene cluster. Complete promoter refactoring and targeted gene exchange in this "dead" cluster led to the discovery of potent indolotryptoline antiproliferative agents, lazarimides A and B. This potentially scalable and cost-effective promoter reengineering platform should streamline the discovery of natural products from silent natural product biosynthetic gene clusters.

  3. Yeast homologous recombination-based promoter engineering for the activation of silent natural product biosynthetic gene clusters

    PubMed Central

    Montiel, Daniel; Kang, Hahk-Soo; Chang, Fang-Yuan; Charlop-Powers, Zachary; Brady, Sean F.

    2015-01-01

    Large-scale sequencing of prokaryotic (meta)genomic DNA suggests that most bacterial natural product gene clusters are not expressed under common laboratory culture conditions. Silent gene clusters represent a promising resource for natural product discovery and the development of a new generation of therapeutics. Unfortunately, the characterization of molecules encoded by these clusters is hampered owing to our inability to express these gene clusters in the laboratory. To address this bottleneck, we have developed a promoter-engineering platform to transcriptionally activate silent gene clusters in a model heterologous host. Our approach uses yeast homologous recombination, an auxotrophy complementation-based yeast selection system and sequence orthogonal promoter cassettes to exchange all native promoters in silent gene clusters with constitutively active promoters. As part of this platform, we constructed and validated a set of bidirectional promoter cassettes consisting of orthogonal promoter sequences, Streptomyces ribosome binding sites, and yeast selectable marker genes. Using these tools we demonstrate the ability to simultaneously insert multiple promoter cassettes into a gene cluster, thereby expediting the reengineering process. We apply this method to model active and silent gene clusters (rebeccamycin and tetarimycin) and to the silent, cryptic pseudogene-containing, environmental DNA-derived Lzr gene cluster. Complete promoter refactoring and targeted gene exchange in this “dead” cluster led to the discovery of potent indolotryptoline antiproliferative agents, lazarimides A and B. This potentially scalable and cost-effective promoter reengineering platform should streamline the discovery of natural products from silent natural product biosynthetic gene clusters. PMID:26150486

  4. The naphthalene catabolic (nag) genes of Polaromonas naphthalenivorans CJ2: Evolutionary implications for two gene clusters and novel regulatory control

    SciTech Connect

    Jeon, C.O.; Park, M.; Ro, H.S.; Park, W.; Madsen, E.L.

    2006-02-15

    Polaromonas naphthalenivorans CJ2, found to be responsible for the degradation of naphthalene in situ at a coal tar waste-contaminated site, is able to grow on mineral salts agar media with naphthalene as the sole carbon source. Beginning from a 484-bp nagAc-like region, we used a genome walking strategy to sequence genes encoding the entire naphthalene degradation pathway and additional flanking regions. We found that the naphthalene catabolic genes in P. naphthalenivorans CJ2 were divided into one large and one small gene cluster, separated by an unknown distance. The large gene cluster is bounded by a LysR-type regulator (nagR). The small cluster is bounded by a MarR-type regulator (nagR2). The catabolic genes of P. naphthalenivorans CJ2 were homologous to many of those of Ralstonia U2, which uses the gentisate pathway to convert naphthalene to central metabolites. However, three open reading frames (nagY, nagM, and nagN), present in Ralstonia U2, were absent. Also, P. naphthalenivorans carries two copies of gentisate dioxygenase (nagI) with 77.4% DNA sequence identity to one another and 82% amino acid identity to their homologue in Ralstonia sp. strain U2. Investigation of the operons using reverse transcription PCR showed that each cluster was controlled independently by its respective promoter. Insertional inactivation and lacZ reporter assays showed that nagR2 is a negative regulator and that expression of the small cluster is not induced by naphthalene, salicylate, or gentisate. Association of two putative Azoarcus-related transposases with the large cluster and one Azoarcus-related putative salicylate 5-hydroxylase gene (ORF2) in the small cluster suggests that mobile genetic elements were likely involved in creating the novel arrangement of catabolic and regulatory genes in P. naphthalenivorans.

  5. Unusual Gene Order and Organization of the Sea Urchin Hox Cluster

    SciTech Connect

    Cameron, R A; Rowen, L; Nesbitt, R; Bloom, S; Rast, J P; Berney, K; Arenas-Mena, C; Martinez, P; Lucas, S; Richardson, P M; Davidson, E H; Peterson, K J; Hood, L

    2005-10-11

    The highly consistent gene order and axial colinear expression patterns found in vertebrate hox gene clusters are less well conserved across the rest of bilaterians. We report the first deuterostome instance of an intact hox cluster with a unique gene order where the paralog groups are not expressed in a sequential manner. The finished sequence from BAC clones from the genome of the sea urchin, Strongylocentrotus purpuratus, reveals a gene order wherein the anterior genes (Hox1, Hox2 and Hox3) lie nearest the posterior genes in the cluster such that the most 3 gene is Hox5. (The gene order is : 5-Hox1, 2, 3, 11/13c, 11/13b, 11/13a, 9/10, 8, 7, 6, 5 - 3). The finished sequence result is corroborated by restriction mapping evidence and BAC-end scaffold analyses. Comparisons with a putative ancestral deuterostome Hox gene cluster suggest that the rearrangements leading to the sea urchin gene order were many and complex.

  6. Unusual Gene Order and Organization of the Sea Urchin HoxCluster

    SciTech Connect

    Richardson, Paul M.; Lucas, Susan; Cameron, R. Andrew; Rowen,Lee; Nesbitt, Ryan; Bloom, Scott; Rast, Jonathan P.; Berney, Kevin; Arenas-Mena, Cesar; Martinez, Pedro; Davidson, Eric H.; Peterson, KevinJ.; Hood, Leroy

    2005-05-10

    The highly consistent gene order and axial colinear expression patterns found in vertebrate hox gene clusters are less well conserved across the rest of bilaterians. We report the first deuterostome instance of an intact hox cluster with a unique gene order where the paralog groups are not expressed in a sequential manner. The finished sequence from BAC clones from the genome of the sea urchin, Strongylocentrotus purpuratus, reveals a gene order wherein the anterior genes (Hox1, Hox2 and Hox3) lie nearest the posterior genes in the cluster such that the most 3' gene is Hox5. (The gene order is : 5'-Hox1,2, 3, 11/13c, 11/13b, '11/13a, 9/10, 8, 7, 6, 5 - 3)'. The finished sequence result is corroborated by restriction mapping evidence and BAC-end scaffold analyses. Comparisons with a putative ancestral deuterostome Hox gene cluster suggest that the rearrangements leading to the sea urchin gene order were many and complex.

  7. Visualization of mappings between the gene ontology and cluster trees

    NASA Astrophysics Data System (ADS)

    Jusufi, Ilir; Kerren, Andreas; Aleksakhin, Vladyslav; Schreiber, Falk

    2012-01-01

    Ontologies and hierarchical clustering are both important tools in biology and medicine to study high-throughput data such as transcriptomics and metabolomics data. Enrichment of ontology terms in the data is used to identify statistically overrepresented ontology terms, giving insight into relevant biological processes or functional modules. Hierarchical clustering is a standard method to analyze and visualize data to find relatively homogeneous clusters of experimental data points. Both methods support the analysis of the same data set, but are usually considered independently. However, often a combined view is desired: visualizing a large data set in the context of an ontology under consideration of a clustering of the data. This paper proposes a new visualization method for this task.

  8. Nanoscale spatial organization of the HoxD gene cluster in distinct transcriptional states.

    PubMed

    Fabre, Pierre J; Benke, Alexander; Joye, Elisabeth; Nguyen Huynh, Thi Hanh; Manley, Suliana; Duboule, Denis

    2015-11-10

    Chromatin condensation plays an important role in the regulation of gene expression. Recently, it was shown that the transcriptional activation of Hoxd genes during vertebrate digit development involves modifications in 3D interactions within and around the HoxD gene cluster. This reorganization follows a global transition from one set of regulatory contacts to another, between two topologically associating domains (TADs) located on either side of the HoxD locus. Here, we use 3D DNA FISH to assess the spatial organization of chromatin at and around the HoxD gene cluster and report that although the two TADs are tightly associated, they appear as spatially distinct units. We measured the relative position of genes within the cluster and found that they segregate over long distances, suggesting that a physical elongation of the HoxD cluster can occur. We analyzed this possibility by super-resolution imaging (STORM) and found that tissues with distinct transcriptional activity exhibit differing degrees of elongation. We also observed that the morphological change of the HoxD cluster in developing digits is associated with its position at the boundary between the two TADs. Such variations in the fine-scale architecture of the gene cluster suggest causal links among its spatial configuration, transcriptional activation, and the flanking chromatin context. PMID:26504220

  9. Engineering Streptomyces coelicolor for heterologous expression of secondary metabolite gene clusters.

    PubMed

    Gomez-Escribano, Juan Pablo; Bibb, Mervyn J

    2011-03-01

    We have constructed derivatives of Streptomyces coelicolor M145 as hosts for the heterologous expression of secondary metabolite gene clusters. To remove potentially competitive sinks of carbon and nitrogen, and to provide a host devoid of antibiotic activity, we deleted four endogenous secondary metabolite gene clusters from S. coelicolor M145--those for actinorhodin, prodiginine, CPK and CDA biosynthesis. We then introduced point mutations into rpoB and rpsL to pleiotropically increase the level of secondary metabolite production. Introduction of the native actinorhodin gene cluster and of gene clusters for the heterologous production of chloramphenicol and congocidine revealed dramatic increases in antibiotic production compared with the parental strain. In addition to lacking antibacterial activity, the engineered strains possess relatively simple extracellular metabolite profiles. When combined with liquid chromatography and mass spectrometry, we believe that these genetically engineered strains will markedly facilitate the discovery of new compounds by heterologous expression of cloned gene clusters, particularly the numerous cryptic secondary metabolic gene clusters that are prevalent within actinomycete genome sequences.

  10. The gene cluster of aureocyclicin 4185: the first cyclic bacteriocin of Staphylococcus aureus.

    PubMed

    Potter, Amina; Ceotto, Hilana; Coelho, Marcus Lívio Varella; Guimarães, Allan J; Bastos, Maria do Carmo de Freire

    2014-05-01

    Staphylococcus aureus 4185 was previously shown to produce at least two bacteriocins. One of them is encoded by pRJ101. To detect the bacteriocin-encoding gene cluster, an ~9160 kb region of pRJ101 was sequenced. In silico analyses identified 10 genes (aclX, aclB, aclI, aclT, aclC, aclD, aclA, aclF, aclG and aclH) that might be involved in the production of a novel cyclic bacteriocin named aureocyclicin 4185. The organization of these genes was quite similar to that of the gene cluster responsible for carnocyclin A production and immunity. Four putative proteins encoded by these genes (AclT, AclC, AclD and AclA) also exhibited similarity to proteins encoded by cyclic bacteriocin gene clusters. Mutants derived from insertion of Tn917-lac into aclC, aclF, aclH and aclX were affected in bacteriocin production and growth. AclX is a 205 aa putative protein not encoded by the gene clusters of other cyclic bacteriocins. AclX exhibits 50 % similarity to a permease and has five putative membrane-spanning domains. Transcription analyses suggested that aclX is part of the aureocyclicin 4185 gene cluster, encoding a protein required for bacteriocin production. The aclA gene is the structural gene of aureocyclicin 4185, which shows 65 % similarity to garvicin ML. AclA is proposed to be cleaved off, generating a mature peptide with a predicted Mr of 5607 Da (60 aa). By homology modelling, AclA presents four α-helices, like carnocyclin A. AclA could not be found at detectable levels in the culture supernatant of a strain carrying only pRJ101. To our knowledge, this is the first report of a cyclic bacteriocin gene cluster in the genus Staphylococcus. PMID:24574434

  11. Characterization of two acetyltransferase genes in the pyripyropene biosynthetic gene cluster from Penicillium coprobium

    PubMed Central

    Hu, Jie; Furutani, Ayako; Yamamoto, Kentaro; Oyama, Kazuhiko; Mitomi, Masaaki; Anzai, Hiroyuki

    2014-01-01

    Pyripyropenes potently and selectively inhibit acyl-CoA:cholesterol acyltransferase 2 (ACAT-2). Among multiple isomers of pyripyropene (A to R), pyripyropene A (PyA) has insecticidal properties in addition to its growth inhibition properties against human umbilical vein endothelial cells. Based on the predicted biosynthetic gene cluster of pyripyropene A, two genes (ppb8 and ppb9) encoding two acetyltransferases (ATs) were separately isolated and introduced into the model fungus Aspergillus oryzae, using the protoplast–polyethylene glycol method. The bioconversion of certain predicted intermediates in the transformants revealed the manner by which acetylation occurred in the biosynthetic pathway by the products expressed by these two genes (AT-1 and AT-2). The acetylated products detected by high-performance liquid chromatography (HPLC) in the extracts from AT-1 and AT-2 transformant clones were not present in the extract from the transformant clone with an empty vector. The HLPC charts of each bioconversion study exhibited high peaks at 12, 10.5 and 9 min, respectively. Further ultraviolet absorption and mass spectrometry analyses identified the products as PyE, PyO and PyA, respectively. AT-1 acetylated the C-1 of deacetyl-pyripyropene E (deAc-PyE), while AT-2 played an active role in acetylating the C-11 of 11-deAc-PyO and C-7 of deAc-PyA at two different steps of the biosynthetic pathway. PMID:26019565

  12. Evidence that a secondary metabolic biosynthetic gene cluster has grown by gene relocation during evolution of the filamentous fungus Fusarium.

    PubMed

    Proctor, Robert H; McCormick, Susan P; Alexander, Nancy J; Desjardins, Anne E

    2009-12-01

    Trichothecenes are terpene-derived secondary metabolites produced by multiple genera of filamentous fungi, including many plant pathogenic species of Fusarium. These metabolites are of interest because they are toxic to animals and plants and can contribute to pathogenesis of Fusarium on some crop species. Fusarium graminearum and F. sporotrichioides have trichothecene biosynthetic genes (TRI) at three loci: a 12-gene TRI cluster and two smaller TRI loci that consist of one or two genes. Here, comparisons of additional Fusarium species have provided evidence that TRI loci have a complex evolutionary history that has included loss, non-functionalization and rearrangement of genes as well as trans-species polymorphism. The results also indicate that the TRI cluster has expanded in some species by relocation of two genes into it from the smaller loci. Thus, evolutionary forces have driven consolidation of TRI genes into fewer loci in some fusaria but have maintained three distinct TRI loci in others. PMID:19843228

  13. Partial mixture model for tight clustering of gene expression time-course

    PubMed Central

    Yuan, Yinyin; Li, Chang-Tsun; Wilson, Roland

    2008-01-01

    Background Tight clustering arose recently from a desire to obtain tighter and potentially more informative clusters in gene expression studies. Scattered genes with relatively loose correlations should be excluded from the clusters. However, in the literature there is little work dedicated to this area of research. On the other hand, there has been extensive use of maximum likelihood techniques for model parameter estimation. By contrast, the minimum distance estimator has been largely ignored. Results In this paper we show the inherent robustness of the minimum distance estimator that makes it a powerful tool for parameter estimation in model-based time-course clustering. To apply minimum distance estimation, a partial mixture model that can naturally incorporate replicate information and allow scattered genes is formulated. We provide experimental results of simulated data fitting, where the minimum distance estimator demonstrates superior performance to the maximum likelihood estimator. Both biological and statistical validations are conducted on a simulated dataset and two real gene expression datasets. Our proposed partial regression clustering algorithm scores top in Gene Ontology driven evaluation, in comparison with four other popular clustering algorithms. Conclusion For the first time partial mixture model is successfully extended to time-course data analysis. The robustness of our partial regression clustering algorithm proves the suitability of the combination of both partial mixture model and minimum distance estimator in this field. We show that tight clustering not only is capable to generate more profound understanding of the dataset under study well in accordance to established biological knowledge, but also presents interesting new hypotheses during interpretation of clustering results. In particular, we provide biological evidences that scattered genes can be relevant and are interesting subjects for study, in contrast to prevailing opinion

  14. Organization, structure and evolution of the CYP2 gene cluster on human chromosome 19.

    PubMed

    Hoffman, S M; Nelson, D R; Keeney, D S

    2001-11-01

    The cytochrome P450 superfamily of mixed-function oxygenases has been extensively studied due to its many critical metabolic roles, and also because it is a fascinating example of gene family evolution. The cluster of genes on human chromosome 19 from the CYP2A, 2B, and 2F subfamilies has been previously described as having a complex organization and many pseudogenes. We describe the discovery of genes from three more CYP2 subfamilies inside the cluster, and assemble a complete map of the region. We comprehensively review the organization, structure, and expression of genes from all six subfamilies. A general hypothesis for the evolution of this complex gene cluster is also presented.

  15. Selenate reductase activity in Escherichia coli requires Isc iron-sulfur cluster biosynthesis genes.

    PubMed

    Yee, Nathan; Choi, Jessica; Porter, Abigail W; Carey, Sean; Rauschenbach, Ines; Harel, Arye

    2014-12-01

    The selenate reductase in Escherichia coli is a multi-subunit enzyme predicted to bind Fe-S clusters. In this study, we examined the iron-sulfur cluster biosynthesis genes that are required for selenate reductase activity. Mutants devoid of either the iscU or hscB gene in the Isc iron-sulfur cluster biosynthesis pathway lost the ability to reduce selenate. Genetic complementation by the wild-type sequences restored selenate reductase activity. The results indicate the Isc biosynthetic system plays a key role in selenate reductase Fe-S cofactor assembly and is essential for enzyme activity.

  16. Identification of two gene clusters involved in cyclohexanone oxidation in Brevibacterium epidermidis strain HCU.

    PubMed

    Brzostowicz, P C; Blasko, M S; Rouvière, P E

    2002-05-01

    Brevibacterium epidermidis HCU can grow on cyclic ketones and alcohols as a sole carbon source. We have previously reported the identification of two cyclohexanone-induced Bayer-Villiger monooxygenase genes by mRNA differential display. Using the related technique of Out-PCR, we have amplified large DNA fragments flanking the two monooxygenase genes. Two large gene clusters were sequenced. Several ORFs in each gene cluster encoded proteins homologous to cyclohexanol and cyclohexanone oxidation enzymes from Acinetobacter. However, the structure of these two gene clusters differs significantly from that of Acinetobacter, where the complete pathway has been described. To assess activity of these genes, they were cloned and expressed in Escherichia coli. In vivo and in vitro assays enabled us to assign functions to the expressed ORFs. These ORFs included a cyclohexanol dehydrogenase, two different epsilon-caprolactone hydrolases and two 6-hydroxyhexanoate dehydrogenases belonging to different enzyme families. Because this environmental isolate is difficult to manipulate, we cannot determine at this time which cluster is involved in the degradation of cyclohexanone under physiological conditions. However, the original differential display experiments and some of the experiments reported here suggest the involvement of both gene clusters in the oxidation of cyclic ketones.

  17. Two Horizontally Transferred Xenobiotic Resistance Gene Clusters Associated with Detoxification of Benzoxazolinones by Fusarium Species

    PubMed Central

    Glenn, Anthony E.; Davis, C. Britton; Gao, Minglu; Gold, Scott E.; Mitchell, Trevor R.; Proctor, Robert H.; Stewart, Jane E.; Snook, Maurice E.

    2016-01-01

    Microbes encounter a broad spectrum of antimicrobial compounds in their environments and often possess metabolic strategies to detoxify such xenobiotics. We have previously shown that Fusarium verticillioides, a fungal pathogen of maize known for its production of fumonisin mycotoxins, possesses two unlinked loci, FDB1 and FDB2, necessary for detoxification of antimicrobial compounds produced by maize, including the γ-lactam 2-benzoxazolinone (BOA). In support of these earlier studies, microarray analysis of F. verticillioides exposed to BOA identified the induction of multiple genes at FDB1 and FDB2, indicating the loci consist of gene clusters. One of the FDB1 cluster genes encoded a protein having domain homology to the metallo-β-lactamase (MBL) superfamily. Deletion of this gene (MBL1) rendered F. verticillioides incapable of metabolizing BOA and thus unable to grow on BOA-amended media. Deletion of other FDB1 cluster genes, in particular AMD1 and DLH1, did not affect BOA degradation. Phylogenetic analyses and topology testing of the FDB1 and FDB2 cluster genes suggested two horizontal transfer events among fungi, one being transfer of FDB1 from Fusarium to Colletotrichum, and the second being transfer of the FDB2 cluster from Fusarium to Aspergillus. Together, the results suggest that plant-derived xenobiotics have exerted evolutionary pressure on these fungi, leading to horizontal transfer of genes that enhance fitness or virulence. PMID:26808652

  18. Two Horizontally Transferred Xenobiotic Resistance Gene Clusters Associated with Detoxification of Benzoxazolinones by Fusarium Species.

    PubMed

    Glenn, Anthony E; Davis, C Britton; Gao, Minglu; Gold, Scott E; Mitchell, Trevor R; Proctor, Robert H; Stewart, Jane E; Snook, Maurice E

    2016-01-01

    Microbes encounter a broad spectrum of antimicrobial compounds in their environments and often possess metabolic strategies to detoxify such xenobiotics. We have previously shown that Fusarium verticillioides, a fungal pathogen of maize known for its production of fumonisin mycotoxins, possesses two unlinked loci, FDB1 and FDB2, necessary for detoxification of antimicrobial compounds produced by maize, including the γ-lactam 2-benzoxazolinone (BOA). In support of these earlier studies, microarray analysis of F. verticillioides exposed to BOA identified the induction of multiple genes at FDB1 and FDB2, indicating the loci consist of gene clusters. One of the FDB1 cluster genes encoded a protein having domain homology to the metallo-β-lactamase (MBL) superfamily. Deletion of this gene (MBL1) rendered F. verticillioides incapable of metabolizing BOA and thus unable to grow on BOA-amended media. Deletion of other FDB1 cluster genes, in particular AMD1 and DLH1, did not affect BOA degradation. Phylogenetic analyses and topology testing of the FDB1 and FDB2 cluster genes suggested two horizontal transfer events among fungi, one being transfer of FDB1 from Fusarium to Colletotrichum, and the second being transfer of the FDB2 cluster from Fusarium to Aspergillus. Together, the results suggest that plant-derived xenobiotics have exerted evolutionary pressure on these fungi, leading to horizontal transfer of genes that enhance fitness or virulence.

  19. Two Horizontally Transferred Xenobiotic Resistance Gene Clusters Associated with Detoxification of Benzoxazolinones by Fusarium Species.

    PubMed

    Glenn, Anthony E; Davis, C Britton; Gao, Minglu; Gold, Scott E; Mitchell, Trevor R; Proctor, Robert H; Stewart, Jane E; Snook, Maurice E

    2016-01-01

    Microbes encounter a broad spectrum of antimicrobial compounds in their environments and often possess metabolic strategies to detoxify such xenobiotics. We have previously shown that Fusarium verticillioides, a fungal pathogen of maize known for its production of fumonisin mycotoxins, possesses two unlinked loci, FDB1 and FDB2, necessary for detoxification of antimicrobial compounds produced by maize, including the γ-lactam 2-benzoxazolinone (BOA). In support of these earlier studies, microarray analysis of F. verticillioides exposed to BOA identified the induction of multiple genes at FDB1 and FDB2, indicating the loci consist of gene clusters. One of the FDB1 cluster genes encoded a protein having domain homology to the metallo-β-lactamase (MBL) superfamily. Deletion of this gene (MBL1) rendered F. verticillioides incapable of metabolizing BOA and thus unable to grow on BOA-amended media. Deletion of other FDB1 cluster genes, in particular AMD1 and DLH1, did not affect BOA degradation. Phylogenetic analyses and topology testing of the FDB1 and FDB2 cluster genes suggested two horizontal transfer events among fungi, one being transfer of FDB1 from Fusarium to Colletotrichum, and the second being transfer of the FDB2 cluster from Fusarium to Aspergillus. Together, the results suggest that plant-derived xenobiotics have exerted evolutionary pressure on these fungi, leading to horizontal transfer of genes that enhance fitness or virulence. PMID:26808652

  20. Revealing gene clusters associated with the development of cholangiocarcinoma, based on a time series analysis.

    PubMed

    Wu, Jianyu; Xiao, Zhifu; Zhao, Xiulei; Wu, Xiangsong

    2015-05-01

    Cholangiocarcinoma (CC) is a rapidly lethal malignancy and currently is considered to be incurable. Biomarkers related to the development of CC remain unclear. The present study aimed to identify differentially expressed genes (DEGs) between normal tissue and intrahepatic CC, as well as specific gene expression patterns that changed together with the development of CC. By using a two‑way analysis of variance test, the biomarkers that could distinguish between normal tissue and intrahepatic CC dissected from different days were identified. A k‑means cluster method was used to identify gene clusters associated with the development of CC according to their changing expression pattern. Functional enrichment analysis was used to infer the function of each of the gene sets. A time series analysis was constructed to reveal gene signatures that were associated with the development of CC based on gene expression profile changes. Genes related to CC were shown to be involved in 'mitochondrion' and 'focal adhesion'. Three interesting gene groups were identified by the k‑means cluster method. Gene clusters with a unique expression pattern are related with the development of CC. The data of this study will facilitate novel discoveries regarding the genetic study of CC by further work.

  1. Histone gene number and organisation in Xenopus: Xenopus borealis has a homogeneous major cluster.

    PubMed Central

    Turner, P C; Woodland, H R

    1983-01-01

    Using a Xenopus laevis H4 cDNA clone as a probe we have determined that the numbers of H4 histone genes in Xenopus laevis and Xenopus borealis are approximately the same. These numbers are dependent on the hybridization stringency and we measure about 90 H4 genes per haploid genome after a 60 degrees C wash in 3 X SSC. Using histone probes from both Xenopus and sea urchin we have studied the genomic organization of histone genes in these two species. In all of the X.borealis individuals analyzed about 70% of the histone genes were present in a very homogeneous major cluster. These genes are present in the order H1, H2B, H2A, H4 and H3, and the minimum length of the repeated unit is 16kb. In contrast, the histone gene clusters in X.laevis showed considerable sequence variation. However two major cluster types with different gene orders seem to be present in most individuals. The differences in histone gene organization seen in species of Xenopus suggest that even in closely related vertebrates the major histone gene clusters are quite fluid structures in evolutionary terms. Images PMID:6298735

  2. The Local Maximum Clustering Method and Its Application in Microarray Gene Expression Data Analysis

    NASA Astrophysics Data System (ADS)

    Wu, Xiongwu; Chen, Yidong; Brooks, Bernard R.; Su, Yan A.

    2004-12-01

    An unsupervised data clustering method, called the local maximum clustering (LMC) method, is proposed for identifying clusters in experiment data sets based on research interest. A magnitude property is defined according to research purposes, and data sets are clustered around each local maximum of the magnitude property. By properly defining a magnitude property, this method can overcome many difficulties in microarray data clustering such as reduced projection in similarities, noises, and arbitrary gene distribution. To critically evaluate the performance of this clustering method in comparison with other methods, we designed three model data sets with known cluster distributions and applied the LMC method as well as the hierarchic clustering method, the[InlineEquation not available: see fulltext.]-mean clustering method, and the self-organized map method to these model data sets. The results show that the LMC method produces the most accurate clustering results. As an example of application, we applied the method to cluster the leukemia samples reported in the microarray study of Golub et al. (1999).

  3. Wide Distribution of O157-Antigen Biosynthesis Gene Clusters in Escherichia coli

    PubMed Central

    Seto, Kazuko; Ooka, Tadasuke; Ogura, Yoshitoshi; Hayashi, Tetsuya; Osawa, Kayo; Osawa, Ro

    2011-01-01

    Most Escherichia coli O157-serogroup strains are classified as enterohemorrhagic E. coli (EHEC), which is known as an important food-borne pathogen for humans. They usually produce Shiga toxin (Stx) 1 and/or Stx2, and express H7-flagella antigen (or nonmotile). However, O157 strains that do not produce Stxs and express H antigens different from H7 are sometimes isolated from clinical and other sources. Multilocus sequence analysis revealed that these 21 O157:non-H7 strains tested in this study belong to multiple evolutionary lineages different from that of EHEC O157:H7 strains, suggesting a wide distribution of the gene set encoding the O157-antigen biosynthesis in multiple lineages. To gain insight into the gene organization and the sequence similarity of the O157-antigen biosynthesis gene clusters, we conducted genomic comparisons of the chromosomal regions (about 59 kb in each strain) covering the O-antigen gene cluster and its flanking regions between six O157:H7/non-H7 strains. Gene organization of the O157-antigen gene cluster was identical among O157:H7/non-H7 strains, but was divided into two distinct types at the nucleotide sequence level. Interestingly, distribution of the two types did not clearly follow the evolutionary lineages of the strains, suggesting that horizontal gene transfer of both types of O157-antigen gene clusters has occurred independently among E. coli strains. Additionally, detailed sequence comparison revealed that some positions of the repetitive extragenic palindromic (REP) sequences in the regions flanking the O-antigen gene clusters were coincident with possible recombination points. From these results, we conclude that the horizontal transfer of the O157-antigen gene clusters induced the emergence of multiple O157 lineages within E. coli and speculate that REP sequences may involve one of the driving forces for exchange and evolution of O-antigen loci. PMID:21876740

  4. Automatic Summarization of Mouse Gene Information by Clustering and Sentence Extraction from MEDLINE Abstracts

    PubMed Central

    Yang, Jianji; Cohen, Aaron M.; Hersh, William

    2007-01-01

    Tools to automatically summarize gene information from the literature have the potential to help genomics researchers better interpret gene expression data and investigate biological pathways. The task of finding information on sets of genes is common for genomic researchers, and PubMed is still the first choice because the most recent and original information can only be found in the unstructured, free text biomedical literature. However, finding information on a set of genes by manually searching and scanning the literature is a time-consuming and daunting task for scientists. We built and evaluated a query-based automatic summarizer of information on mouse genes studied in microarray experiments. The system clusters a set of genes by MeSH, GO and free text features and presents summaries for each gene by ranked sentences extracted from MEDLINE abstracts. Evaluation showed that the system seems to provide meaningful clusters and informative sentences are ranked higher by the algorithm. PMID:18693953

  5. Biosilica formation in spicules of the sponge Suberites domuncula: synchronous expression of a gene cluster.

    PubMed

    Schröder, Heinz C; Perovic-Ottstadt, Sanja; Grebenjuk, Vladislav A; Engel, Sylvia; Müller, Isabel M; Müller, Werner E G

    2005-06-01

    The formation of spicules is a complicated morphogenetic process in sponges (phylum Porifera). The primmorph system was used to demonstrate that in the demosponge Suberites domuncula the synthesis of the siliceous spicules starts intracellularly and is dependent on the concentration of silicic acid. To understand spicule formation, a cluster of genes was isolated. In the center of this cluster is the silicatein gene, which codes for the enzyme that synthesizes spicules. This gene is flanked by an ankyrin repeat gene at one side and by a tumor necrosis factor receptor-associated factor and a protein kinase gene at the other side. All genes are strongly expressed in primmorphs and intact animals after exposure to silicic acid, and this expression is restricted to those areas where the spicule formation starts or where spicules are maintained in the animals. Our observations suggest that in S. domuncula a coordinated expression of physically linked genes is essential for the synthesis of the major skeletal elements.

  6. Clustering of time-course gene expression profiles using normal mixture models with autoregressive random effects

    PubMed Central

    2012-01-01

    Background Time-course gene expression data such as yeast cell cycle data may be periodically expressed. To cluster such data, currently used Fourier series approximations of periodic gene expressions have been found not to be sufficiently adequate to model the complexity of the time-course data, partly due to their ignoring the dependence between the expression measurements over time and the correlation among gene expression profiles. We further investigate the advantages and limitations of available models in the literature and propose a new mixture model with autoregressive random effects of the first order for the clustering of time-course gene-expression profiles. Some simulations and real examples are given to demonstrate the usefulness of the proposed models. Results We illustrate the applicability of our new model using synthetic and real time-course datasets. We show that our model outperforms existing models to provide more reliable and robust clustering of time-course data. Our model provides superior results when genetic profiles are correlated. It also gives comparable results when the correlation between the gene profiles is weak. In the applications to real time-course data, relevant clusters of coregulated genes are obtained, which are supported by gene-function annotation databases. Conclusions Our new model under our extension of the EMMIX-WIRE procedure is more reliable and robust for clustering time-course data because it adopts a random effects model that allows for the correlation among observations at different time points. It postulates gene-specific random effects with an autocorrelation variance structure that models coregulation within the clusters. The developed R package is flexible in its specification of the random effects through user-input parameters that enables improved modelling and consequent clustering of time-course data. PMID:23151154

  7. The Fumagillin Gene Cluster, an Example of Hundreds of Genes under veA Control in Aspergillus fumigatus

    PubMed Central

    Dhingra, Sourabh; Lind, Abigail L.; Lin, Hsiao-Ching; Tang, Yi; Rokas, Antonis; Calvo, Ana M.

    2013-01-01

    Aspergillus fumigatus is the causative agent of invasive aspergillosis, leading to infection-related mortality in immunocompromised patients. We previously showed that the conserved and unique-to-fungi veA gene affects different cell processes such as morphological development, gliotoxin biosynthesis and protease activity, suggesting a global regulatory effect on the genome of this medically relevant fungus. In this study, RNA sequencing analysis revealed that veA controls the expression of hundreds of genes in A. fumigatus, including those comprising more than a dozen known secondary metabolite gene clusters. Chemical analysis confirmed that veA controls the synthesis of other secondary metabolites in this organism in addition to gliotoxin. Among the secondary metabolite gene clusters regulated by veA is the elusive but recently identified gene cluster responsible for the biosynthesis of fumagillin, a meroterpenoid known for its anti-angiogenic activity by binding to human methionine aminopeptidase 2. The fumagillin gene cluster contains a veA-dependent regulatory gene, fumR (Afu8g00420), encoding a putative C6 type transcription factor. Deletion of fumR results in silencing of the gene cluster and elimination of fumagillin biosynthesis. We found expression of fumR to also be dependent on laeA, a gene encoding another component of the fungal velvet complex. The results in this study argue that veA is a global regulator of secondary metabolism in A. fumigatus, and that veA may be a conduit via which chemical development is coupled to morphological development and other cellular processes. PMID:24116213

  8. Insights into secondary metabolism from a global analysis of prokaryotic biosynthetic gene clusters

    PubMed Central

    Cimermancic, Peter; Medema, Marnix H.; Claesen, Jan; Kurita, Kenji; Wieland Brown, Laura C.; Mavrommatis, Konstantinos; Pati, Amrita; Godfrey, Paul A.; Koehrsen, Michael; Clardy, Jon; Birren, Bruce W.; Takano, Eriko; Sali, Andrej; Linington, Roger G.; Fischbach, Michael A.

    2014-01-01

    Summary Although biosynthetic gene clusters (BGCs) have been discovered for hundreds of bacterial metabolites, our knowledge of their diversity remains limited. Here, we used a novel algorithm to systematically identify BGCs in the extensive extant microbial sequencing data. Network analysis of the predicted BGCs revealed large gene cluster families, the vast majority uncharacterized. We experimentally characterized the most prominent family, consisting of two subfamilies of hundreds of BGCs distributed throughout the Proteobacteria; their products are aryl polyenes, lipids with an aryl head group conjugated to a polyene tail. We identified a distant relationship to a third subfamily of aryl polyene BGCs, and together the three subfamilies represent the largest known family of biosynthetic gene clusters, with more than 1,000 members. Although these clusters are widely divergent in sequence, their small molecule products are remarkably conserved, indicating for the first time the important roles these compounds play in Gram-negative cell biology. PMID:25036635

  9. Selfish Operons: Horizontal Transfer May Drive the Evolution of Gene Clusters

    PubMed Central

    Lawrence, J. G.; Roth, J. R.

    1996-01-01

    A model is presented whereby the formation of gene clusters in bacteria is mediated by transfer of DNA within and among taxa. Bacterial operons are typically composed of genes whose products contribute to a single function. If this function is subject to weak selection or to long periods with no selection, the contributing genes may accumulate mutations and be lost by genetic drift. From a cell's perspective, once several genes are lost, the function can be restored only if all missing genes were acquired simultaneously by lateral transfer. The probability of transfer of multiple genes increases when genes are physically proximate. From a gene's perspective, horizontal transfer provides a way to escape evolutionary loss by allowing colonization of organisms lacking the encoded functions. Since organisms bearing clustered genes are more likely to act as successful donors, clustered genes would spread among bacterial genomes. The physical proximity of genes may be considered a selfish property of the operon since it affects the probability of successful horizontal transfer but may provide no physiological benefit to the host. This process predicts a mosaic structure of modern genomes in which ancestral chromosomal material is interspersed with novel, horizontally transferred operons providing peripheral metabolic functions. PMID:8844169

  10. Shared Gene Structures and Clusters of Mutually Exclusive Spliced Exons within the Metazoan Muscle Myosin Heavy Chain Genes

    PubMed Central

    Kollmar, Martin; Hatje, Klas

    2014-01-01

    Multicellular animals possess two to three different types of muscle tissues. Striated muscles have considerable ultrastructural similarity and contain a core set of proteins including the muscle myosin heavy chain (Mhc) protein. The ATPase activity of this myosin motor protein largely dictates muscle performance at the molecular level. Two different solutions to adjusting myosin properties to different muscle subtypes have been identified so far: Vertebrates and nematodes contain many independent differentially expressed Mhc genes while arthropods have single Mhc genes with clusters of mutually exclusive spliced exons (MXEs). The availability of hundreds of metazoan genomes now allowed us to study whether the ancient bilateria already contained MXEs, how MXE complexity subsequently evolved, and whether additional scenarios to control contractile properties in different muscles could be proposed, By reconstructing the Mhc genes from 116 metazoans we showed that all intron positions within the motor domain coding regions are conserved in all bilateria analysed. The last common ancestor of the bilateria already contained a cluster of MXEs coding for part of the loop-2 actin-binding sequence. Subsequently the protostomes and later the arthropods gained many further clusters while MXEs got completely lost independently in several branches (vertebrates and nematodes) and species (for example the annelid Helobdella robusta and the salmon louse Lepeophtheirus salmonis). Several bilateria have been found to encode multiple Mhc genes that might all or in part contain clusters of MXEs. Notable examples are a cluster of six tandemly arrayed Mhc genes, of which two contain MXEs, in the owl limpet Lottia gigantea and four Mhc genes with three encoding MXEs in the predatory mite Metaseiulus occidentalis. Our analysis showed that similar solutions to provide different myosin isoforms (multiple genes or clusters of MXEs or both) have independently been developed several times

  11. Genomics-driven discovery of the pneumocandin biosynthetic gene cluster in the fungus Glarea lozoyensis

    PubMed Central

    2013-01-01

    Background The antifungal therapy caspofungin is a semi-synthetic derivative of pneumocandin B0, a lipohexapeptide produced by the fungus Glarea lozoyensis, and was the first member of the echinocandin class approved for human therapy. The nonribosomal peptide synthetase (NRPS)-polyketide synthases (PKS) gene cluster responsible for pneumocandin biosynthesis from G. lozoyensis has not been elucidated to date. In this study, we report the elucidation of the pneumocandin biosynthetic gene cluster by whole genome sequencing of the G. lozoyensis wild-type strain ATCC 20868. Results The pneumocandin biosynthetic gene cluster contains a NRPS (GLNRPS4) and a PKS (GLPKS4) arranged in tandem, two cytochrome P450 monooxygenases, seven other modifying enzymes, and genes for L-homotyrosine biosynthesis, a component of the peptide core. Thus, the pneumocandin biosynthetic gene cluster is significantly more autonomous and organized than that of the recently characterized echinocandin B gene cluster. Disruption mutants of GLNRPS4 and GLPKS4 no longer produced the pneumocandins (A0 and B0), and the Δglnrps4 and Δglpks4 mutants lost antifungal activity against the human pathogenic fungus Candida albicans. In addition to pneumocandins, the G. lozoyensis genome encodes a rich repertoire of natural product-encoding genes including 24 PKSs, six NRPSs, five PKS-NRPS hybrids, two dimethylallyl tryptophan synthases, and 14 terpene synthases. Conclusions Characterization of the gene cluster provides a blueprint for engineering new pneumocandin derivatives with improved pharmacological properties. Whole genome estimation of the secondary metabolite-encoding genes from G. lozoyensis provides yet another example of the huge potential for drug discovery from natural products from the fungal kingdom. PMID:23688303

  12. Degeneration of aflatoxin gene cluster in Aspergillus flavus from Africa and North America

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus flavus is the primary causal agent of food and feed contamination with the toxic fungal metabolites aflatoxins. Aflatoxin-producing potential of A. flavus is known to vary among isolates. The genes involved in aflatoxin biosynthesis are clustered together and the order of genes within th...

  13. Identification of the Herboxidiene Biosynthetic Gene Cluster in Streptomyces chromofuscus ATCC 49982

    PubMed Central

    Shao, Lei; Zi, Jiachen; Zeng, Jia

    2012-01-01

    The 53-kb biosynthetic gene cluster for the novel anticholesterol natural product herboxidiene was identified in Streptomyces chromofuscus ATCC 49982 by genome sequencing and gene inactivation. In addition to herboxidiene, a biosynthetic intermediate, 18-deoxy-herboxidiene, was also isolated from the fermentation broth of S. chromofuscus ATCC 49982 as a minor metabolite. PMID:22247174

  14. Degeneration of aflatoxin gene clusters in Aspergillus flavus from Africa and North America.

    PubMed

    Adhikari, Bishwo N; Bandyopadhyay, Ranajit; Cotty, Peter J

    2016-12-01

    Aspergillus flavus is the most common causal agent of aflatoxin contamination of food and feed. However, aflatoxin-producing potential varies widely among A. flavus genotypes with many producing no aflatoxins. Some non-aflatoxigenic genotypes are used as biocontrol agents to prevent contamination. Aflatoxin biosynthesis genes are tightly clustered in a highly conserved order. Gene deletions and presence of single nucleotide polymorphisms (SNPs) in aflatoxin biosynthesis genes are often associated with A. flavus inability to produce aflatoxins. In order to identify mechanisms of non-aflatoxigenicity in non-aflatoxigenic genotypes of value in aflatoxin biocontrol, complete cluster sequences of 35 A. flavus genotypes from Africa and North America were analyzed. Inability of some genotypes to produce aflatoxin resulted from deletion of biosynthesis genes. In other genotypes, non-aflatoxigenicity originated from SNP formation. The process of degeneration differed across the gene cluster; genes involved in early biosynthesis stages were more likely to be deleted while genes involved in later stages displayed high frequencies of SNPs. Comparative analyses of aflatoxin gene clusters provides insight into the diversity of mechanisms of non-aflatoxigenicity in A. flavus genotypes used as biological control agents. The sequences provide resources for both diagnosis of non-aflatoxigenicity and monitoring of biocontrol genotypes during biopesticide manufacture and in the environment. PMID:27576895

  15. Phage p1-derived artificial chromosomes facilitate heterologous expression of the FK506 gene cluster.

    PubMed

    Jones, Adam C; Gust, Bertolt; Kulik, Andreas; Heide, Lutz; Buttner, Mark J; Bibb, Mervyn J

    2013-01-01

    We describe a procedure for the conjugative transfer of phage P1-derived Artificial Chromosome (PAC) library clones containing large natural product gene clusters (≥70 kilobases) to Streptomyces coelicolor strains that have been engineered for improved heterologous production of natural products. This approach is demonstrated using the gene cluster for FK506 (tacrolimus), a clinically important immunosuppressant of high commercial value. The entire 83.5 kb FK506 gene cluster from Streptomyces tsukubaensis NRRL 18488 present in one 130 kb PAC clone was introduced into four different S. coelicolor derivatives and all produced FK506 and smaller amounts of the related compound FK520. FK506 yields were increased by approximately five-fold (from 1.2 mg L(-1) to 5.5 mg L(-1)) in S. coelicolor M1146 containing the FK506 PAC upon over-expression of the FK506 LuxR regulatory gene fkbN. The PAC-based gene cluster conjugation methodology described here provides a tractable means to evaluate and manipulate FK506 biosynthesis and is readily applicable to other large gene clusters encoding natural products of interest to medicine, agriculture and biotechnology.

  16. Structural variation of the ribosomal gene cluster within the class Insecta

    SciTech Connect

    Mukha, D.V.; Sidorenko, A.P.; Lazebnaya, I.V.

    1995-09-01

    General estimation of ribosomal DNA variation within the class Insecta is presented. It is shown that, using blot-hybridization, one can detect differences in the structure of the ribosomal gene cluster not only between genera within an order, but also between species within a genera, including sibling species. Structure of the ribosomal gene cluster of the Coccinellidae family (ladybirds) is analyzed. It is shown that cloned highly conservative regions of ribosomal DNA of Tetrahymena pyriformis can be used as probes for analyzing ribosomal genes in insects. 24 refs., 4 figs.

  17. Heterologous Expression of Fluostatin Gene Cluster Leads to a Bioactive Heterodimer.

    PubMed

    Yang, Chunfang; Huang, Chunshuai; Zhang, Wenjun; Zhu, Yiguang; Zhang, Changsheng

    2015-11-01

    The biosynthesis gene cluster (fls) for atypical angucycline fluostatins was identified from the marine derived Micromonospora rosaria SCSIO N160 and was confirmed by gene knockouts and the biochemical characterization of a bifunctional oxygenase FlsO2. The absolute configuration of the key biosynthetic intermediate prejadomycin was determined for the first time by Cu Kα X-ray analysis. Heterologous expression of the intact fls-gene cluster in Streptomyces coelicolor YF11 in the presence of 3% sea salts led to the isolation of two new compounds: fluostatin L (1) and difluostatin A (2). Difluostatin A (2), an unusual heterodimer, exhibited antibacterial activities.

  18. Adh enhances Actinobacillus pleuropneumoniae pathogenicity by binding to OR5M11 and activating p38 which induces apoptosis of PAMs and IL-8 release.

    PubMed

    Wang, Lei; Qin, Wanhai; Zhang, Jing; Bao, Chuntong; Zhang, Hu; Che, Yanyi; Sun, Changjiang; Gu, Jingmin; Feng, Xin; Du, Chongtao; Han, Wenyu; Richard, Paul Langford; Lei, Liancheng

    2016-04-05

    Members of the Trimeric Autotransporter Adhesin (TAA) family play a crucial role in the adhesion of Gram-negative pathogens to host cells, but the immunopathogenesis of TAAs remains unknown. Our previous studies demonstrated that Adh from Actinobacillus pleuropneumoniae (A. pleuropneumoniae) is required for full bacterial pathogenicity. Alveolar macrophages are the first line of defense against respiratory infections. This study compared the interactions between porcine alveolar macrophages (PAMs) and wild-type A. pleuropneumoniae (5b WT) or an Adh-deletion strain (5b ΔAdh) via gene microarray, immunoprecipitation and other technologies. We found that Adh was shown to interact with the PAMs membrane protein OR5M11, an olfactory receptor, resulting in the high-level secretion of IL-8 by activation of p38 MAPK signaling pathway. Subsequently, PAMs apoptosis via the activation of the Fax and Bax signaling pathways was observed, followed by activation of caspases 8, 9, and 3. The immunological pathogenic roles of Adh were also confirmed in both murine and piglets infectious models in vivo. These results identify a novel immunological strategy for TAAs to boost the pathogenicity of A. pleuropneumoniae. Together, these datas reveal the high versatility of the Adh protein as a virulence factor and provide novel insight into the immunological pathogenic role of TAAs.

  19. Adh enhances Actinobacillus pleuropneumoniae pathogenicity by binding to OR5M11 and activating p38 which induces apoptosis of PAMs and IL-8 release

    PubMed Central

    Wang, Lei; Qin, Wanhai; Zhang, Jing; Bao, Chuntong; Zhang, Hu; Che, Yanyi; Sun, Changjiang; Gu, Jingmin; Feng, Xin; Du, Chongtao; Han, Wenyu; Richard, Paul Langford; Lei, Liancheng

    2016-01-01

    Members of the Trimeric Autotransporter Adhesin (TAA) family play a crucial role in the adhesion of Gram-negative pathogens to host cells, but the immunopathogenesis of TAAs remains unknown. Our previous studies demonstrated that Adh from Actinobacillus pleuropneumoniae (A. pleuropneumoniae) is required for full bacterial pathogenicity. Alveolar macrophages are the first line of defense against respiratory infections. This study compared the interactions between porcine alveolar macrophages (PAMs) and wild-type A. pleuropneumoniae (5b WT) or an Adh-deletion strain (5b ΔAdh) via gene microarray, immunoprecipitation and other technologies. We found that Adh was shown to interact with the PAMs membrane protein OR5M11, an olfactory receptor, resulting in the high-level secretion of IL-8 by activation of p38 MAPK signaling pathway. Subsequently, PAMs apoptosis via the activation of the Fax and Bax signaling pathways was observed, followed by activation of caspases 8, 9, and 3. The immunological pathogenic roles of Adh were also confirmed in both murine and piglets infectious models in vivo. These results identify a novel immunological strategy for TAAs to boost the pathogenicity of A. pleuropneumoniae. Together, these datas reveal the high versatility of the Adh protein as a virulence factor and provide novel insight into the immunological pathogenic role of TAAs. PMID:27046446

  20. A putative greigite-type magnetosome gene cluster from the candidate phylum Latescibacteria.

    PubMed

    Lin, Wei; Pan, Yongxin

    2015-04-01

    The intracellular biomineralization of magnetite and/or greigite magnetosomes in magnetotactic bacteria (MTB) is strictly controlled by a group of conserved genes, termed magnetosome genes, which are organized as clusters (or islands) in MTB genomes. So far, all reported MTB are affiliated within the Proteobacteria phylum, the Nitrospirae phylum and the candidate division OP3. Here, we report the discovery of a putative magnetosome gene cluster structure from the draft genome of an uncultivated bacterium belonging to the candidate phylum Latescibacteria (formerly candidate division WS3) recently recovered by Rinke and colleagues, which contains 10 genes with homology to magnetosome mam genes of magnetotactic Proteobacteria and Nitrospirae. Moreover, these genes are phylogenetically closely related to greigite-type magnetosome genes that were only found from the Deltaproteobacteria MTB before, suggesting that the greigite genes may originate earlier than previously imagined. These findings indicate that some members of Latescibacteria may be capable of forming greigite magnetosomes, and thus may play previously unrecognized roles in environmental iron and sulfur cycles. The conserved genomic structure of magnetosome gene cluster in Latescibacteria phylum supports the hypothesis of horizontal transfer of these genes among distantly related bacterial groups in nature.

  1. Co-clustering phenome–genome for phenotype classification and disease gene discovery

    PubMed Central

    Hwang, TaeHyun; Atluri, Gowtham; Xie, MaoQiang; Dey, Sanjoy; Hong, Changjin; Kumar, Vipin; Kuang, Rui

    2012-01-01

    Understanding the categorization of human diseases is critical for reliably identifying disease causal genes. Recently, genome-wide studies of abnormal chromosomal locations related to diseases have mapped >2000 phenotype–gene relations, which provide valuable information for classifying diseases and identifying candidate genes as drug targets. In this article, a regularized non-negative matrix tri-factorization (R-NMTF) algorithm is introduced to co-cluster phenotypes and genes, and simultaneously detect associations between the detected phenotype clusters and gene clusters. The R-NMTF algorithm factorizes the phenotype–gene association matrix under the prior knowledge from phenotype similarity network and protein–protein interaction network, supervised by the label information from known disease classes and biological pathways. In the experiments on disease phenotype–gene associations in OMIM and KEGG disease pathways, R-NMTF significantly improved the classification of disease phenotypes and disease pathway genes compared with support vector machines and Label Propagation in cross-validation on the annotated phenotypes and genes. The newly predicted phenotypes in each disease class are highly consistent with human phenotype ontology annotations. The roles of the new member genes in the disease pathways are examined and validated in the protein–protein interaction subnetworks. Extensive literature review also confirmed many new members of the disease classes and pathways as well as the predicted associations between disease phenotype classes and pathways. PMID:22735708

  2. Structure and gene cluster of the O-antigen of Escherichia coli O137.

    PubMed

    Perepelov, Andrei V; Guo, Xi; Senchenkova, Sof'ya N; Li, Yayue; Shashkov, Alexander S; Liu, Bin; Knirel, Yuriy A

    2016-03-01

    The O-polysaccharide (O-antigen) was isolated from the lipopolysaccharide of Escherichia coli O137 and studied by sugar analysis and NMR spectroscopy. The following structure of the branched tetrasaccharide repeating unit was established: Formula: see text] Both structure and gene cluster of the E. coli O137 polysaccharide are related to those of the E. coli K40 polysaccharide (Amor et al., 1999), which lacks the side-chain glucosylation but contains serine that is amide-linked to GlcA. Functions of genes in the O137-antigen gene cluster were assigned by a comparison with those in K40 and sequences in the available databases. Particularly, predicted glycosyltransferases encoded in the gene cluster were assigned to the formation of three glycosidic linkages in the O-polysaccharide repeating unit.

  3. Identification and manipulation of the pleuromutilin gene cluster from Clitopilus passeckerianus for increased rapid antibiotic production

    NASA Astrophysics Data System (ADS)

    Bailey, Andy M.; Alberti, Fabrizio; Kilaru, Sreedhar; Collins, Catherine M.; de Mattos-Shipley, Kate; Hartley, Amanda J.; Hayes, Patrick; Griffin, Alison; Lazarus, Colin M.; Cox, Russell J.; Willis, Christine L.; O’Dwyer, Karen; Spence, David W.; Foster, Gary D.

    2016-05-01

    Semi-synthetic derivatives of the tricyclic diterpene antibiotic pleuromutilin from the basidiomycete Clitopilus passeckerianus are important in combatting bacterial infections in human and veterinary medicine. These compounds belong to the only new class of antibiotics for human applications, with novel mode of action and lack of cross-resistance, representing a class with great potential. Basidiomycete fungi, being dikaryotic, are not generally amenable to strain improvement. We report identification of the seven-gene pleuromutilin gene cluster and verify that using various targeted approaches aimed at increasing antibiotic production in C. passeckerianus, no improvement in yield was achieved. The seven-gene pleuromutilin cluster was reconstructed within Aspergillus oryzae giving production of pleuromutilin in an ascomycete, with a significant increase (2106%) in production. This is the first gene cluster from a basidiomycete to be successfully expressed in an ascomycete, and paves the way for the exploitation of a metabolically rich but traditionally overlooked group of fungi.

  4. Microbisporicin gene cluster reveals unusual features of lantibiotic biosynthesis in actinomycetes

    PubMed Central

    Foulston, Lucy C.; Bibb, Mervyn J.

    2010-01-01

    Lantibiotics are ribosomally synthesized, posttranslationally modified peptide antibiotics. The biosynthetic gene cluster for microbisporicin, a potent lantibiotic produced by the actinomycete Microbispora corallina containing chlorinated tryptophan and dihydroxyproline residues, was identified by genome scanning and isolated from an M. corallina cosmid library. Heterologous expression in Nonomuraea sp. ATCC 39727 confirmed that all of the genes required for microbisporicin biosynthesis were present in the cluster. Deletion, in M. corallina, of the gene (mibA) predicted to encode the prepropeptide abolished microbisporicin production. Further deletion analysis revealed insights into the biosynthesis of this unusual and potentially clinically useful lantibiotic, shedding light on mechanisms of regulation and self-resistance. In particular, we report an example of the involvement of a tryptophan halogenase in the modification of a ribosomally synthesized peptide and the pathway-specific regulation of an antibiotic biosynthetic gene cluster by an extracytoplasmic function σ factor–anti-σ factor complex. PMID:20628010

  5. Identification and manipulation of the pleuromutilin gene cluster from Clitopilus passeckerianus for increased rapid antibiotic production

    PubMed Central

    Bailey, Andy M.; Alberti, Fabrizio; Kilaru, Sreedhar; Collins, Catherine M.; de Mattos-Shipley, Kate; Hartley, Amanda J.; Hayes, Patrick; Griffin, Alison; Lazarus, Colin M.; Cox, Russell J.; Willis, Christine L.; O’Dwyer, Karen; Spence, David W.; Foster, Gary D.

    2016-01-01

    Semi-synthetic derivatives of the tricyclic diterpene antibiotic pleuromutilin from the basidiomycete Clitopilus passeckerianus are important in combatting bacterial infections in human and veterinary medicine. These compounds belong to the only new class of antibiotics for human applications, with novel mode of action and lack of cross-resistance, representing a class with great potential. Basidiomycete fungi, being dikaryotic, are not generally amenable to strain improvement. We report identification of the seven-gene pleuromutilin gene cluster and verify that using various targeted approaches aimed at increasing antibiotic production in C. passeckerianus, no improvement in yield was achieved. The seven-gene pleuromutilin cluster was reconstructed within Aspergillus oryzae giving production of pleuromutilin in an ascomycete, with a significant increase (2106%) in production. This is the first gene cluster from a basidiomycete to be successfully expressed in an ascomycete, and paves the way for the exploitation of a metabolically rich but traditionally overlooked group of fungi. PMID:27143514

  6. Microbisporicin gene cluster reveals unusual features of lantibiotic biosynthesis in actinomycetes.

    PubMed

    Foulston, Lucy C; Bibb, Mervyn J

    2010-07-27

    Lantibiotics are ribosomally synthesized, posttranslationally modified peptide antibiotics. The biosynthetic gene cluster for microbisporicin, a potent lantibiotic produced by the actinomycete Microbispora corallina containing chlorinated tryptophan and dihydroxyproline residues, was identified by genome scanning and isolated from an M. corallina cosmid library. Heterologous expression in Nonomuraea sp. ATCC 39727 confirmed that all of the genes required for microbisporicin biosynthesis were present in the cluster. Deletion, in M. corallina, of the gene (mibA) predicted to encode the prepropeptide abolished microbisporicin production. Further deletion analysis revealed insights into the biosynthesis of this unusual and potentially clinically useful lantibiotic, shedding light on mechanisms of regulation and self-resistance. In particular, we report an example of the involvement of a tryptophan halogenase in the modification of a ribosomally synthesized peptide and the pathway-specific regulation of an antibiotic biosynthetic gene cluster by an extracytoplasmic function sigma factor-anti-sigma factor complex.

  7. Ethylene-responsive transcription factors interact with promoters of ADH and PDC involved in persimmon (Diospyros kaki) fruit de-astringency.

    PubMed

    Min, Ting; Yin, Xue-ren; Shi, Yan-na; Luo, Zheng-rong; Yao, Yun-cong; Grierson, Donald; Ferguson, Ian B; Chen, Kun-song

    2012-11-01

    The persimmon fruit is a particularly good model for studying fruit response to hypoxia, in particular, the hypoxia-response ERF (HRE) genes. An anaerobic environment reduces fruit astringency by converting soluble condensed tannins (SCTs) into an insoluble form. Although the physiology of de-astringency has been widely studied, its molecular control is poorly understood. Both CO(2) and ethylene treatments efficiently removed the astringency from 'Mopan' persimmon fruit, as indicated by a decrease in SCTs. Acetaldehyde, the putative agent for causing de-astringency, accumulated during these treatments, as did activities of the key enzymes of acetaldehyde synthesis, alcohol dehydrogenase (ADH), and pyruvate decarboxylase (PDC). Eight DkADH and DkPDC genes were isolated, and three candidates for a role in de-astringency, DkADH1, DkPDC1, and DkPDC2, were characterized by transcriptional analysis in different tissues. The significance of these specific isoforms was confirmed by principal component analysis. Transient expression in leaf tissue showed that DkPDC2 decreased SCTs. Interactions of six hypoxia-responsive ERF genes and target promoters were tested in transient assays. The results indicated that two hypoxia-responsive ERF genes, DkERF9 and DkERF10, were involved in separately regulating the DkPDC2 and DkADH1 promoters. It is proposed that a DkERF-DkADH/DkPDC cascade is involved in regulating persimmon de-astringency.

  8. Ethylene-responsive transcription factors interact with promoters of ADH and PDC involved in persimmon (Diospyros kaki) fruit de-astringency

    PubMed Central

    Min, Ting; Yin, Xue-ren; Chen, Kun-song

    2012-01-01

    The persimmon fruit is a particularly good model for studying fruit response to hypoxia, in particular, the hypoxia-response ERF (HRE) genes. An anaerobic environment reduces fruit astringency by converting soluble condensed tannins (SCTs) into an insoluble form. Although the physiology of de-astringency has been widely studied, its molecular control is poorly understood. Both CO2 and ethylene treatments efficiently removed the astringency from ‘Mopan’ persimmon fruit, as indicated by a decrease in SCTs. Acetaldehyde, the putative agent for causing de-astringency, accumulated during these treatments, as did activities of the key enzymes of acetaldehyde synthesis, alcohol dehydrogenase (ADH), and pyruvate decarboxylase (PDC). Eight DkADH and DkPDC genes were isolated, and three candidates for a role in de-astringency, DkADH1, DkPDC1, and DkPDC2, were characterized by transcriptional analysis in different tissues. The significance of these specific isoforms was confirmed by principal component analysis. Transient expression in leaf tissue showed that DkPDC2 decreased SCTs. Interactions of six hypoxia-responsive ERF genes and target promoters were tested in transient assays. The results indicated that two hypoxia-responsive ERF genes, DkERF9 and DkERF10, were involved in separately regulating the DkPDC2 and DkADH1 promoters. It is proposed that a DkERF–DkADH/DkPDC cascade is involved in regulating persimmon de-astringency. PMID:23095993

  9. Paradigm of Tunable Clustering Using Binarization of Consensus Partition Matrices (Bi-CoPaM) for Gene Discovery

    PubMed Central

    Abu-Jamous, Basel; Fa, Rui; Roberts, David J.; Nandi, Asoke K.

    2013-01-01

    Clustering analysis has a growing role in the study of co-expressed genes for gene discovery. Conventional binary and fuzzy clustering do not embrace the biological reality that some genes may be irrelevant for a problem and not be assigned to a cluster, while other genes may participate in several biological functions and should simultaneously belong to multiple clusters. Also, these algorithms cannot generate tight clusters that focus on their cores or wide clusters that overlap and contain all possibly relevant genes. In this paper, a new clustering paradigm is proposed. In this paradigm, all three eventualities of a gene being exclusively assigned to a single cluster, being assigned to multiple clusters, and being not assigned to any cluster are possible. These possibilities are realised through the primary novelty of the introduction of tunable binarization techniques. Results from multiple clustering experiments are aggregated to generate one fuzzy consensus partition matrix (CoPaM), which is then binarized to obtain the final binary partitions. This is referred to as Binarization of Consensus Partition Matrices (Bi-CoPaM). The method has been tested with a set of synthetic datasets and a set of five real yeast cell-cycle datasets. The results demonstrate its validity in generating relevant tight, wide, and complementary clusters that can meet requirements of different gene discovery studies. PMID:23409186

  10. Clusters of Antibiotic Resistance Genes Enriched Together Stay Together in Swine Agriculture

    PubMed Central

    Johnson, Timothy A.; Stedtfeld, Robert D.; Wang, Qiong; Cole, James R.; Hashsham, Syed A.; Looft, Torey; Zhu, Yong-Guan

    2016-01-01

    ABSTRACT   Antibiotic resistance is a worldwide health risk, but the influence of animal agriculture on the genetic context and enrichment of individual antibiotic resistance alleles remains unclear. Using quantitative PCR followed by amplicon sequencing, we quantified and sequenced 44 genes related to antibiotic resistance, mobile genetic elements, and bacterial phylogeny in microbiomes from U.S. laboratory swine and from swine farms from three Chinese regions. We identified highly abundant resistance clusters: groups of resistance and mobile genetic element alleles that cooccur. For example, the abundance of genes conferring resistance to six classes of antibiotics together with class 1 integrase and the abundance of IS6100-type transposons in three Chinese regions are directly correlated. These resistance cluster genes likely colocalize in microbial genomes in the farms. Resistance cluster alleles were dramatically enriched (up to 1 to 10% as abundant as 16S rRNA) and indicate that multidrug-resistant bacteria are likely the norm rather than an exception in these communities. This enrichment largely occurred independently of phylogenetic composition; thus, resistance clusters are likely present in many bacterial taxa. Furthermore, resistance clusters contain resistance genes that confer resistance to antibiotics independently of their particular use on the farms. Selection for these clusters is likely due to the use of only a subset of the broad range of chemicals to which the clusters confer resistance. The scale of animal agriculture and its wastes, the enrichment and horizontal gene transfer potential of the clusters, and the vicinity of large human populations suggest that managing this resistance reservoir is important for minimizing human risk. PMID:27073098

  11. Unbiased Functional Clustering of Gene Variants with a Phenotypic-Linkage Network

    PubMed Central

    Honti, Frantisek; Meader, Stephen; Webber, Caleb

    2014-01-01

    Groupwise functional analysis of gene variants is becoming standard in next-generation sequencing studies. As the function of many genes is unknown and their classification to pathways is scant, functional associations between genes are often inferred from large-scale omics data. Such data types—including protein–protein interactions and gene co-expression networks—are used to examine the interrelations of the implicated genes. Statistical significance is assessed by comparing the interconnectedness of the mutated genes with that of random gene sets. However, interconnectedness can be affected by confounding bias, potentially resulting in false positive findings. We show that genes implicated through de novo sequence variants are biased in their coding-sequence length and longer genes tend to cluster together, which leads to exaggerated p-values in functional studies; we present here an integrative method that addresses these bias. To discern molecular pathways relevant to complex disease, we have inferred functional associations between human genes from diverse data types and assessed them with a novel phenotype-based method. Examining the functional association between de novo gene variants, we control for the heretofore unexplored confounding bias in coding-sequence length. We test different data types and networks and find that the disease-associated genes cluster more significantly in an integrated phenotypic-linkage network than in other gene networks. We present a tool of superior power to identify functional associations among genes mutated in the same disease even after accounting for significant sequencing study bias and demonstrate the suitability of this method to functionally cluster variant genes underlying polygenic disorders. PMID:25166029

  12. A Genomics Based Discovery of Secondary Metabolite Biosynthetic Gene Clusters in Aspergillus ustus

    PubMed Central

    Pi, Borui; Yu, Dongliang; Dai, Fangwei; Song, Xiaoming; Zhu, Congyi; Li, Hongye; Yu, Yunsong

    2015-01-01

    Secondary metabolites (SMs) produced by Aspergillus have been extensively studied for their crucial roles in human health, medicine and industrial production. However, the resulting information is almost exclusively derived from a few model organisms, including A. nidulans and A. fumigatus, but little is known about rare pathogens. In this study, we performed a genomics based discovery of SM biosynthetic gene clusters in Aspergillus ustus, a rare human pathogen. A total of 52 gene clusters were identified in the draft genome of A. ustus 3.3904, such as the sterigmatocystin biosynthesis pathway that was commonly found in Aspergillus species. In addition, several SM biosynthetic gene clusters were firstly identified in Aspergillus that were possibly acquired by horizontal gene transfer, including the vrt cluster that is responsible for viridicatumtoxin production. Comparative genomics revealed that A. ustus shared the largest number of SM biosynthetic gene clusters with A. nidulans, but much fewer with other Aspergilli like A. niger and A. oryzae. These findings would help to understand the diversity and evolution of SM biosynthesis pathways in genus Aspergillus, and we hope they will also promote the development of fungal identification methodology in clinic. PMID:25706180

  13. Ethanol-Induced ADH Activity in Zebrafish: Differential Concentration-Dependent Effects on High- Versus Low-Affinity ADH Enzymes.

    PubMed

    Tran, Steven; Nowicki, Magda; Facciol, Amanda; Chatterjee, Diptendu; Gerlai, Robert

    2016-04-01

    Zebrafish express enzymes that metabolize ethanol in a manner comparable to that of mammals, including humans. We previously demonstrated that acute ethanol exposure increases alcohol dehydrogenase (ADH) activity in an inverted U-shaped dose-dependent manner. It was hypothesized that the biphasic dose-response was due to the increased activity of a high-affinity ADH isoform following exposure to low concentrations of ethanol and increased activity of a low-affinity ADH isoform following exposure to higher concentrations of ethanol. To test this hypothesis, we exposed zebrafish to different concentrations of ethanol (0%, 0.25%, 0.5%, and 1.0% v/v) for 30 min and measured the total ADH activity in the zebrafish liver. However, we also repeated this enzyme activity assay using a low concentration of the substrate (ethanol) to determine the activity of high-affinity ADH isoforms. We found that total ADH activity in response to ethanol induces an inverted U-shaped dose-response similar to our previous study. Using a lower substrate level in our enzyme assay targeting high-affinity isozymes, we found a similar dose-response. However, the difference in activity between the high and low substrate assays (high substrate activity - low substrate activity), which provide an index of activity for low-affinity ADH isoforms, revealed no significant effect of ethanol exposure. Our results suggest that the inverted U-shaped dose-response for total ADH activity in response to ethanol is driven primarily by high-affinity isoforms of ADH.

  14. Genetic analysis of chromosomal mutations in the polysialic acid gene cluster of Escherichia coli K1.

    PubMed Central

    Vimr, E R; Aaronson, W; Silver, R P

    1989-01-01

    The kps gene cluster of Escherichia coli K1 encodes functions for sialic acid synthesis, activation, polymerization, and possibly translocation of polymer to the cell surface. The size and complexity of this membrane polysaccharide biosynthetic cluster have hindered genetic mapping and functional descriptions of the kps genes. To begin a detailed investigation of the polysialic acid synthetic mechanism, acapsular mutants were characterized to determine their probable defects in polymer synthesis. The mutants were tested for complementation with kps fragments subcloned from two separately isolated, functionally intact kps gene clusters. Complementation was assayed by immunological and biochemical methods and by sensitivity to the K1-specific bacteriophage K1F. The kps cluster consisted of a central 5.8-kilobase region that contained at least two genes coding for sialic acid synthetic enzymes, a gene encoding the sialic acid-activating enzyme, and a gene encoding the sialic acid polymerase. This biosynthetic region is flanked on one side by an approximately 2.8-kilobase region that contains a potential regulatory locus and at least one structural gene for a polypeptide that appears to function in polysialic acid assembly. Flanking the biosynthetic region on the opposite side is a 6- to 8.4-kilobase region that codes for at least three proteins which may also function in polymer assembly and possibly in translocating polymer to the outer cell surface. Results of transduction crosses supported these conclusions and indicated that some of the kps genes flanking the central biosynthetic region may not function directly in transporting polymer to the cell surface. The results also demonstrate that the map position and probable function of most of the kps cluster genes have been identified. Images PMID:2644224

  15. Molecular cloning and identification of the laspartomycin biosynthetic gene cluster from Streptomyces viridochromogenes

    PubMed Central

    Wang, Yang; Chen, Ying; Shen, Qirong; Yin, Xihou

    2011-01-01

    The biosynthetic gene cluster for laspartomycins, a family of 11 amino acid peptide antibiotics, has been cloned and sequenced from Streptomyces viridochromogenes ATCC 29814. Annotation of a segment of 88912 bp of S. viridochromogenes genomic sequence revealed the putative las cluster and its flanking regions which harbor 43 open reading frames. The lpm cluster, which spans approximately 60 kb, consists of 21 open reading frames. Those include four NRPS genes (lpmA/orf18, lpmB/orf25, lpmC/orf26 and lpmD/orf27), four genes (orfs 21, 22, 24 and 29) involved in the lipid tail biosynthesis and attachment, four regulatory genes (orfs 13, 19, 32 and 33) and three putative exporters or self-resistance genes (orfs 14, 20 and 30). In addition, the gene involved in the biosynthesis of the nonproteinogenic amino acid Pip was also identified in the lpm cluster while the genes necessary for the biosynthesis of the rare residue diaminopropionic acid (Dap) were found to reside elsewhere on the chromosome. Interestingly, the dabA, dabB and dabC genes predicted to code for the biosynthesis of the unusual amino acid diaminobutyric acid (Dab) are organized into the lpm cluster even though the Dab residue was not found in the laspartomycins. Disruption of the NRPS lpmC gene completely abolished laspartomycin production in the corresponding mutant strain. These findings will allow molecular engineering and combinatorial biosynthesis approaches to expand the structural diversity of the amphomycin-group peptide antibiotics including the laspartomycins and friulimicins. PMID:21640802

  16. The Eucalyptus grandis NBS-LRR Gene Family: Physical Clustering and Expression Hotspots

    PubMed Central

    Christie, Nanette; Tobias, Peri A.; Naidoo, Sanushka; Külheim, Carsten

    2016-01-01

    Eucalyptus grandis is a commercially important hardwood species and is known to be susceptible to a number of pests and pathogens. Determining mechanisms of defense is therefore a research priority. The published genome for E. grandis has aided the identification of one important class of resistance (R) genes that incorporate nucleotide binding sites and leucine-rich repeat domains (NBS-LRR). Using an iterative search process we identified NBS-LRR gene models within the E. grandis genome. We characterized the gene models and identified their genomic arrangement. The gene expression patterns were examined in E. grandis clones, challenged with a fungal pathogen (Chrysoporthe austroafricana) and insect pest (Leptocybe invasa). One thousand two hundred and fifteen putative NBS-LRR coding sequences were located which aligned into two large classes, Toll or interleukin-1 receptor (TIR) and coiled-coil (CC) based on NB-ARC domains. NBS-LRR gene-rich regions were identified with 76% organized in clusters of three or more genes. A further 272 putative incomplete resistance genes were also identified. We determined that E. grandis has a higher ratio of TIR to CC classed genes compared to other woody plant species as well as a smaller percentage of single NBS-LRR genes. Transcriptome profiles indicated expression hotspots, within physical clusters, including expression of many incomplete genes. The clustering of putative NBS-LRR genes correlates with differential expression responses in resistant and susceptible plants indicating functional relevance for the physical arrangement of this gene family. This analysis of the repertoire and expression of E. grandis putative NBS-LRR genes provides an important resource for the identification of novel and functional R-genes; a key objective for strategies to enhance resilience. PMID:26793216

  17. Regulation of Three Nitrogenase Gene Clusters in the Cyanobacterium Anabaena variabilis ATCC 29413

    PubMed Central

    Thiel, Teresa; Pratte, Brenda S.

    2014-01-01

    The filamentous cyanobacterium Anabaena variabilis ATCC 29413 fixes nitrogen under aerobic conditions in specialized cells called heterocysts that form in response to an environmental deficiency in combined nitrogen. Nitrogen fixation is mediated by the enzyme nitrogenase, which is very sensitive to oxygen. Heterocysts are microxic cells that allow nitrogenase to function in a filament comprised primarily of vegetative cells that produce oxygen by photosynthesis. A. variabilis is unique among well-characterized cyanobacteria in that it has three nitrogenase gene clusters that encode different nitrogenases, which function under different environmental conditions. The nif1 genes encode a Mo-nitrogenase that functions only in heterocysts, even in filaments grown anaerobically. The nif2 genes encode a different Mo-nitrogenase that functions in vegetative cells, but only in filaments grown under anoxic conditions. An alternative V-nitrogenase is encoded by vnf genes that are expressed only in heterocysts in an environment that is deficient in Mo. Thus, these three nitrogenases are expressed differentially in response to environmental conditions. The entire nif1 gene cluster, comprising at least 15 genes, is primarily under the control of the promoter for the first gene, nifB1. Transcriptional control of many of the downstream nif1 genes occurs by a combination of weak promoters within the coding regions of some downstream genes and by RNA processing, which is associated with increased transcript stability. The vnf genes show a similar pattern of transcriptional and post-transcriptional control of expression suggesting that the complex pattern of regulation of the nif1 cluster is conserved in other cyanobacterial nitrogenase gene clusters. PMID:25513762

  18. Sequencing and mapping hemoglobin gene clusters in the australian model dasyurid marsupial sminthopsis macroura

    SciTech Connect

    De Leo, A.A.; Wheeler, D.; Lefevre, C.; Cheng, Jan-Fang; Hope, R.; Kuliwaba, J.; Nicholas, K.R.; Westermanc, M.; Graves, J.A.M.

    2004-07-26

    Comparing globin genes and their flanking sequences across many species has allowed globin gene evolution to be reconstructed in great detail. Marsupial globin sequences have proved to be of exceptional significance. A previous finding of a beta-like omega gene in the alpha cluster in the tammar wallaby suggested that the alpha and beta cluster evolved via genome duplication and loss rather than tandem duplication. To confirm and extend this important finding we isolated and sequenced BACs containing the alpha and beta loci from the distantly related Australian marsupial Sminthopsis macroura. We report that the alpha gene lies in the same BAC as the beta-like omega gene, implying that the alpha-omega juxtaposition is likely to be conserved in all marsupials. The LUC7L gene was found 3' of the S. macroura alpha locus, a gene order shared with humans but not mouse, chicken or fugu. Sequencing a BAC contig that contained the S. macroura beta globin and epsilon globin loci showed that the globin cluster is flanked by olfactory genes, demonstrating a gene arrangement conserved for over 180 MY. Analysis of the region 5' to the S. macroura epsilon globin gene revealed a region similar to the eutherian LCR, containing sequences and potential transcription factor binding sites with homology to eutherian hypersensitive sites 1 to 5. FISH mapping of BACs containing S. macroura alpha and beta globin genes located the beta globin cluster on chromosome 3q and the alpha locus close to the centromere on 1q, resolving contradictory map locations obtained by previous radioactive in situ hybridization.

  19. An exploration of the sequence of a 2.9-Mb region of the genome of Drosophila melanogaster: the Adh region.

    PubMed Central

    Ashburner, M; Misra, S; Roote, J; Lewis, S E; Blazej, R; Davis, T; Doyle, C; Galle, R; George, R; Harris, N; Hartzell, G; Harvey, D; Hong, L; Houston, K; Hoskins, R; Johnson, G; Martin, C; Moshrefi, A; Palazzolo, M; Reese, M G; Spradling, A; Tsang, G; Wan, K; Whitelaw, K; Celniker, S

    1999-01-01

    A contiguous sequence of nearly 3 Mb from the genome of Drosophila melanogaster has been sequenced from a series of overlapping P1 and BAC clones. This region covers 69 chromosome polytene bands on chromosome arm 2L, including the genetically well-characterized "Adh region." A computational analysis of the sequence predicts 218 protein-coding genes, 11 tRNAs, and 17 transposable element sequences. At least 38 of the protein-coding genes are arranged in clusters of from 2 to 6 closely related genes, suggesting extensive tandem duplication. The gene density is one protein-coding gene every 13 kb; the transposable element density is one element every 171 kb. Of 73 genes in this region identified by genetic analysis, 49 have been located on the sequence; P-element insertions have been mapped to 43 genes. Ninety-five (44%) of the known and predicted genes match a Drosophila EST, and 144 (66%) have clear similarities to proteins in other organisms. Genes known to have mutant phenotypes are more likely to be represented in cDNA libraries, and far more likely to have products similar to proteins of other organisms, than are genes with no known mutant phenotype. Over 650 chromosome aberration breakpoints map to this chromosome region, and their nonrandom distribution on the genetic map reflects variation in gene spacing on the DNA. This is the first large-scale analysis of the genome of D. melanogaster at the sequence level. In addition to the direct results obtained, this analysis has allowed us to develop and test methods that will be needed to interpret the complete sequence of the genome of this species.Before beginning a Hunt, it is wise to ask someone what you are looking for before you begin looking for it. Milne 1926 PMID:10471707

  20. Functional Gene Networks: R/Bioc package to generate and analyse gene networks derived from functional enrichment and clustering

    PubMed Central

    Aibar, Sara; Fontanillo, Celia; Droste, Conrad; De Las Rivas, Javier

    2015-01-01

    Summary: Functional Gene Networks (FGNet) is an R/Bioconductor package that generates gene networks derived from the results of functional enrichment analysis (FEA) and annotation clustering. The sets of genes enriched with specific biological terms (obtained from a FEA platform) are transformed into a network by establishing links between genes based on common functional annotations and common clusters. The network provides a new view of FEA results revealing gene modules with similar functions and genes that are related to multiple functions. In addition to building the functional network, FGNet analyses the similarity between the groups of genes and provides a distance heatmap and a bipartite network of functionally overlapping genes. The application includes an interface to directly perform FEA queries using different external tools: DAVID, GeneTerm Linker, TopGO or GAGE; and a graphical interface to facilitate the use. Availability and implementation: FGNet is available in Bioconductor, including a tutorial. URL: http://bioconductor.org/packages/release/bioc/html/FGNet.html Contact: jrivas@usal.es Supplementary information: Supplementary data are available at Bioinformatics online. PMID:25600944

  1. Sequencing and Transcriptional Analysis of the Biosynthesis Gene Cluster of Abscisic Acid-Producing Botrytis cinerea

    PubMed Central

    Gong, Tao; Shu, Dan; Yang, Jie; Ding, Zhong-Tao; Tan, Hong

    2014-01-01

    Botrytis cinerea is a model species with great importance as a pathogen of plants and has become used for biotechnological production of ABA. The ABA cluster of B. cinerea is composed of an open reading frame without significant similarities (bcaba3), followed by the genes (bcaba1 and bcaba2) encoding P450 monooxygenases and a gene probably coding for a short-chain dehydrogenase/reductase (bcaba4). In B. cinerea ATCC58025, targeted inactivation of the genes in the cluster suggested at least three genes responsible for the hydroxylation at carbon atom C-1' and C-4' or oxidation at C-4' of ABA. Our group has identified an ABA-overproducing strain, B. cinerea TB-3-H8. To differentiate TB-3-H8 from other B. cinerea strains with the functional ABA cluster, the DNA sequence of the 12.11-kb region containing the cluster of B. cinerea TB-3-H8 was determined. Full-length cDNAs were also isolated for bcaba1, bcaba2, bcaba3 and bcaba4 from B. cinerea TB-3-H8. Sequence comparison of the four genes and their flanking regions respectively derived from B. cinerea TB-3-H8, B05.10 and T4 revealed that major variations were located in intergenic sequences. In B. cinerea TB-3-H8, the expression profiles of the four function genes under ABA high-yield conditions were also analyzed by real-time PCR. PMID:25268614

  2. Breaking the Silence: Protein Stabilization Uncovers Silenced Biosynthetic Gene Clusters in the Fungus Aspergillus nidulans

    PubMed Central

    Gerke, Jennifer; Bayram, Özgür; Feussner, Kirstin; Landesfeind, Manuel; Shelest, Ekaterina; Feussner, Ivo

    2012-01-01

    The genomes of filamentous fungi comprise numerous putative gene clusters coding for the biosynthesis of chemically and structurally diverse secondary metabolites (SMs), which are rarely expressed under laboratory conditions. Previous approaches to activate these genes were based primarily on artificially targeting the cellular protein synthesis apparatus. Here, we applied an alternative approach of genetically impairing the protein degradation apparatus of the model fungus Aspergillus nidulans by deleting the conserved eukaryotic csnE/CSN5 deneddylase subunit of the COP9 signalosome. This defect in protein degradation results in the activation of a previously silenced gene cluster comprising a polyketide synthase gene producing the antibiotic 2,4-dihydroxy-3-methyl-6-(2-oxopropyl)benzaldehyde (DHMBA). The csnE/CSN5 gene is highly conserved in fungi, and therefore, the deletion is a feasible approach for the identification of new SMs. PMID:23001671

  3. Cloning and characterization of the goadsporin biosynthetic gene cluster from Streptomyces sp. TP-A0584.

    PubMed

    Onaka, Hiroyasu; Nakaho, Mizuho; Hayashi, Keiko; Igarashi, Yasuhiro; Furumai, Tamotsu

    2005-12-01

    The biosynthetic gene cluster of goadsporin, a polypeptide antibiotic containing thiazole and oxazole rings, was cloned from Streptomyces sp. TP-A0584. The cluster contains a structural gene, godA, and nine god (goadsporin) genes involved in post-translational modification, immunity and transcriptional regulation. Although the gene organization is similar to typical bacteriocin biosynthetic gene clusters, each goadsporin biosynthetic gene shows low homology to these genes. Goadsporin biosynthesis is initiated by the translation of godA, and the subsequent cyclization, dehydration and acetylation are probably catalysed by godD, godE, godF, godG and godH gene products. godI shows high similarity to the 54 kDa subunit of the signal recognition particle and plays an important role in goadsporin immunity. Furthermore, four goadsporin analogues were produced by site-directed mutagenesis of godA, suggesting that this biosynthesis machinery is used for the heterocyclization of peptides. PMID:16339937

  4. Form gene clustering method about pan-ethnic-group products based on emotional semantic

    NASA Astrophysics Data System (ADS)

    Chen, Dengkai; Ding, Jingjing; Gao, Minzhuo; Ma, Danping; Liu, Donghui

    2016-09-01

    The use of pan-ethnic-group products form knowledge primarily depends on a designer's subjective experience without user participation. The majority of studies primarily focus on the detection of the perceptual demands of consumers from the target product category. A pan-ethnic-group products form gene clustering method based on emotional semantic is constructed. Consumers' perceptual images of the pan-ethnic-group products are obtained by means of product form gene extraction and coding and computer aided product form clustering technology. A case of form gene clustering about the typical pan-ethnic-group products is investigated which indicates that the method is feasible. This paper opens up a new direction for the future development of product form design which improves the agility of product design process in the era of Industry 4.0.

  5. Physical and genetic map of the major nif gene cluster from Azotobacter vinelandii.

    PubMed

    Jacobson, M R; Brigle, K E; Bennett, L T; Setterquist, R A; Wilson, M S; Cash, V L; Beynon, J; Newton, W E; Dean, D R

    1989-02-01

    Determination of a 28,793-base-pair DNA sequence of a region from the Azotobacter vinelandii genome that includes and flanks the nitrogenase structural gene region was completed. This information was used to revise the previously proposed organization of the major nif cluster. The major nif cluster from A. vinelandii encodes 15 nif-specific genes whose products bear significant structural identity to the corresponding nif-specific gene products from Klebsiella pneumoniae. These genes include nifH, nifD, nifK, nifT, nifY, nifE, nifN, nifX, nifU, nifS, nifV, nifW, nifZ, nifM, and nifF. Although there are significant spatial differences, the identified A. vinelandii nif-specific genes have the same sequential arrangement as the corresponding nif-specific genes from K. pneumoniae. Twelve other potential genes whose expression could be subject to nif-specific regulation were also found interspersed among the identified nif-specific genes. These potential genes do not encode products that are structurally related to the identified nif-specific gene products. Eleven potential nif-specific promoters were identified within the major nif cluster, and nine of these are preceded by an appropriate upstream activator sequence. A + T-rich regions were identified between 8 of the 11 proposed nif promoter sequences and their upstream activator sequences. Site-directed deletion-and-insertion mutagenesis was used to establish a genetic map of the major nif cluster.

  6. Physical and genetic map of the major nif gene cluster from Azotobacter vinelandii.

    PubMed Central

    Jacobson, M R; Brigle, K E; Bennett, L T; Setterquist, R A; Wilson, M S; Cash, V L; Beynon, J; Newton, W E; Dean, D R

    1989-01-01

    Determination of a 28,793-base-pair DNA sequence of a region from the Azotobacter vinelandii genome that includes and flanks the nitrogenase structural gene region was completed. This information was used to revise the previously proposed organization of the major nif cluster. The major nif cluster from A. vinelandii encodes 15 nif-specific genes whose products bear significant structural identity to the corresponding nif-specific gene products from Klebsiella pneumoniae. These genes include nifH, nifD, nifK, nifT, nifY, nifE, nifN, nifX, nifU, nifS, nifV, nifW, nifZ, nifM, and nifF. Although there are significant spatial differences, the identified A. vinelandii nif-specific genes have the same sequential arrangement as the corresponding nif-specific genes from K. pneumoniae. Twelve other potential genes whose expression could be subject to nif-specific regulation were also found interspersed among the identified nif-specific genes. These potential genes do not encode products that are structurally related to the identified nif-specific gene products. Eleven potential nif-specific promoters were identified within the major nif cluster, and nine of these are preceded by an appropriate upstream activator sequence. A + T-rich regions were identified between 8 of the 11 proposed nif promoter sequences and their upstream activator sequences. Site-directed deletion-and-insertion mutagenesis was used to establish a genetic map of the major nif cluster. PMID:2644218

  7. Contributions of vertical descent, horizontal transfer and gene loss to the distribution of mycotoxin biosynthetic gene clusters in Fusarium

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The genus Fusarium produces a diverse array of mycotoxins and other secondary metabolites, but individual species contribute to only a small fraction of this diversity. Here, we employed comparative genomic and phylogenetic analyses to investigate the distribution and evolution of gene clusters resp...

  8. Delineation of metabolic gene clusters in plant genomes by chromatin signatures

    PubMed Central

    Yu, Nan; Nützmann, Hans-Wilhelm; MacDonald, James T.; Moore, Ben; Field, Ben; Berriri, Souha; Trick, Martin; Rosser, Susan J.; Kumar, S. Vinod; Freemont, Paul S.; Osbourn, Anne

    2016-01-01

    Plants are a tremendous source of diverse chemicals, including many natural product-derived drugs. It has recently become apparent that the genes for the biosynthesis of numerous different types of plant natural products are organized as metabolic gene clusters, thereby unveiling a highly unusual form of plant genome architecture and offering novel avenues for discovery and exploitation of plant specialized metabolism. Here we show that these clustered pathways are characterized by distinct chromatin signatures of histone 3 lysine trimethylation (H3K27me3) and histone 2 variant H2A.Z, associated with cluster repression and activation, respectively, and represent discrete windows of co-regulation in the genome. We further demonstrate that knowledge of these chromatin signatures along with chromatin mutants can be used to mine genomes for cluster discovery. The roles of H3K27me3 and H2A.Z in repression and activation of single genes in plants are well known. However, our discovery of highly localized operon-like co-regulated regions of chromatin modification is unprecedented in plants. Our findings raise intriguing parallels with groups of physically linked multi-gene complexes in animals and with clustered pathways for specialized metabolism in filamentous fungi. PMID:26895889

  9. Delineation of metabolic gene clusters in plant genomes by chromatin signatures.

    PubMed

    Yu, Nan; Nützmann, Hans-Wilhelm; MacDonald, James T; Moore, Ben; Field, Ben; Berriri, Souha; Trick, Martin; Rosser, Susan J; Kumar, S Vinod; Freemont, Paul S; Osbourn, Anne

    2016-03-18

    Plants are a tremendous source of diverse chemicals, including many natural product-derived drugs. It has recently become apparent that the genes for the biosynthesis of numerous different types of plant natural products are organized as metabolic gene clusters, thereby unveiling a highly unusual form of plant genome architecture and offering novel avenues for discovery and exploitation of plant specialized metabolism. Here we show that these clustered pathways are characterized by distinct chromatin signatures of histone 3 lysine trimethylation (H3K27me3) and histone 2 variant H2A.Z, associated with cluster repression and activation, respectively, and represent discrete windows of co-regulation in the genome. We further demonstrate that knowledge of these chromatin signatures along with chromatin mutants can be used to mine genomes for cluster discovery. The roles of H3K27me3 and H2A.Z in repression and activation of single genes in plants are well known. However, our discovery of highly localized operon-like co-regulated regions of chromatin modification is unprecedented in plants. Our findings raise intriguing parallels with groups of physically linked multi-gene complexes in animals and with clustered pathways for specialized metabolism in filamentous fungi. PMID:26895889

  10. ToppCluster: a multiple gene list feature analyzer for comparative enrichment clustering and network-based dissection of biological systems.

    PubMed

    Kaimal, Vivek; Bardes, Eric E; Tabar, Scott C; Jegga, Anil G; Aronow, Bruce J

    2010-07-01

    ToppCluster is a web server application that leverages a powerful enrichment analysis and underlying data environment for comparative analyses of multiple gene lists. It generates heatmaps or connectivity networks that reveal functional features shared or specific to multiple gene lists. ToppCluster uses hypergeometric tests to obtain list-specific feature enrichment P-values for currently 17 categories of annotations of human-ortholog genes, and provides user-selectable cutoffs and multiple testing correction methods to control false discovery. Each nameable gene list represents a column input to a resulting matrix whose rows are overrepresented features, and individual cells per-list P-values and corresponding genes per feature. ToppCluster provides users with choices of tabular outputs, hierarchical clustering and heatmap generation, or the ability to interactively select features from the functional enrichment matrix to be transformed into XGMML or GEXF network format documents for use in Cytoscape or Gephi applications, respectively. Here, as example, we demonstrate the ability of ToppCluster to enable identification of list-specific phenotypic and regulatory element features (both cis-elements and 3'UTR microRNA binding sites) among tissue-specific gene lists. ToppCluster's functionalities enable the identification of specialized biological functions and regulatory networks and systems biology-based dissection of biological states. ToppCluster can be accessed freely at http://toppcluster.cchmc.org.

  11. Birth, death and horizontal transfer of the fumonisin biosynthetic gene cluster during the evolutionary diversification of Fusarium.

    PubMed

    Proctor, Robert H; Van Hove, François; Susca, Antonia; Stea, Gaetano; Busman, Mark; van der Lee, Theo; Waalwijk, Cees; Moretti, Antonio; Ward, Todd J

    2013-10-01

    Fumonisins are a family of carcinogenic secondary metabolites produced by members of the Fusarium fujikuroi species complex (FFSC) and rare strains of Fusarium oxysporum. In Fusarium, fumonisin biosynthetic genes (FUM) are clustered, and the cluster is uniform in gene organization. Here, sequence analyses indicated that the cluster exists in five different genomic contexts, defining five cluster types. In FUM gene genealogies, evolutionary relationships between fusaria with different cluster types were largely incongruent with species relationships inferred from primary-metabolism (PM) gene genealogies, and FUM cluster types are not trans-specific. In addition, synonymous site divergence analyses indicated that three FUM cluster types predate diversification of FFSC. The data are not consistent with balancing selection or interspecific hybridization, but they are consistent with two competing hypotheses: (i) multiple horizontal transfers of the cluster from unknown donors to FFSC recipients and (ii) cluster duplication and loss (birth and death). Furthermore, low levels of FUM gene divergence in F. bulbicola, an FFSC species, and F. oxysporum provide evidence for horizontal transfer of the cluster from the former, or a closely related species, to the latter. Thus, uniform gene organization within the FUM cluster belies a complex evolutionary history that has not always paralleled the evolution of Fusarium.

  12. Comprehensive annotation of secondary metabolite biosynthetic genes and gene clusters of Aspergillus nidulans, A. fumigatus, A. niger and A. oryzae

    PubMed Central

    2013-01-01

    Background Secondary metabolite production, a hallmark of filamentous fungi, is an expanding area of research for the Aspergilli. These compounds are potent chemicals, ranging from deadly toxins to therapeutic antibiotics to potential anti-cancer drugs. The genome sequences for multiple Aspergilli have been determined, and provide a wealth of predictive information about secondary metabolite production. Sequence analysis and gene overexpression strategies have enabled the discovery of novel secondary metabolites and the genes involved in their biosynthesis. The Aspergillus Genome Database (AspGD) provides a central repository for gene annotation and protein information for Aspergillus species. These annotations include Gene Ontology (GO) terms, phenotype data, gene names and descriptions and they are crucial for interpreting both small- and large-scale data and for aiding in the design of new experiments that further Aspergillus research. Results We have manually curated Biological Process GO annotations for all genes in AspGD with recorded functions in secondary metabolite production, adding new GO terms that specifically describe each secondary metabolite. We then leveraged these new annotations to predict roles in secondary metabolism for genes lacking experimental characterization. As a starting point for manually annotating Aspergillus secondary metabolite gene clusters, we used antiSMASH (antibiotics and Secondary Metabolite Analysis SHell) and SMURF (Secondary Metabolite Unknown Regions Finder) algorithms to identify potential clusters in A. nidulans, A. fumigatus, A. niger and A. oryzae, which we subsequently refined through manual curation. Conclusions This set of 266 manually curated secondary metabolite gene clusters will facilitate the investigation of novel Aspergillus secondary metabolites. PMID:23617571

  13. Arrangements of alpha-globin gene cluster in Taiwan.

    PubMed

    Peng, H W; Choo, K B; Ho, C H; Yen, M S; Liung, W Y; Lin, C K; Yang, Z L; Ng, H T; Ching, K N; Han, S H

    1989-01-01

    In a gene mapping study on 217 newborn babies in Taiwan with alpha- and zeta-globin probes, we have observed 4 cases (1.84%) of alpha-thalassemia-2 heterozygotes (zeta zeta-alpha/zeta zeta alpha alpha) without increased levels of hemoglobin (Hb) Bart's in the cord blood. Eleven subjects (5.07%) were found to have the South East Asian alpha-thalassemia-1 haplotype (zeta zeta--SEA/zeta zeta alpha alpha) with increased Hb Bart's levels ranging from 2.2 to 9%. One case, with Hb Bart's level of 14% in the cord blood, was found to have the genotype of zeta zeta--SEA/zeta zeta alpha alpha T (0.46%). Four heterozygotes (1.84%) were found with the triple alpha gene anti-rightward arrangement (zeta zeta alpha alpha alpha 3.7/zeta zeta alpha alpha). Twenty-one heterozygotes (9.68%) were found to have the triple zeta-globin gene arrangement (zeta zeta zeta alpha alpha/zeta zeta alpha alpha). A new triple zeta-globin gene variant with a BamHI polymorphism was also observed in this study.

  14. microRNAs in the Same Clusters Evolve to Coordinately Regulate Functionally Related Genes.

    PubMed

    Wang, Yirong; Luo, Junjie; Zhang, Hong; Lu, Jian

    2016-09-01

    MicroRNAs (miRNAs) are endogenously expressed small noncoding RNAs. The genomic locations of animal miRNAs are significantly clustered in discrete loci. We found duplication and de novo formation were important mechanisms to create miRNA clusters and the clustered miRNAs tend to be evolutionarily conserved. We proposed a "functional co-adaptation" model to explain how clustering helps newly emerged miRNAs survive and develop functions. We presented evidence that abundance of miRNAs in the same clusters were highly correlated and those miRNAs exerted cooperative repressive effects on target genes in human tissues. By transfecting miRNAs into human and fly cells and extensively profiling the transcriptome alteration with deep-sequencing, we further demonstrated the functional co-adaptation between new and old miRNAs in the miR-17-92 cluster. Our population genomic analysis suggest that positive Darwinian selection might be the driving force underlying the formation and evolution of miRNA clustering. Our model provided novel insights into mechanisms and evolutionary significance of miRNA clustering. PMID:27189568

  15. microRNAs in the Same Clusters Evolve to Coordinately Regulate Functionally Related Genes

    PubMed Central

    Wang, Yirong; Luo, Junjie; Zhang, Hong; Lu, Jian

    2016-01-01

    MicroRNAs (miRNAs) are endogenously expressed small noncoding RNAs. The genomic locations of animal miRNAs are significantly clustered in discrete loci. We found duplication and de novo formation were important mechanisms to create miRNA clusters and the clustered miRNAs tend to be evolutionarily conserved. We proposed a “functional co-adaptation” model to explain how clustering helps newly emerged miRNAs survive and develop functions. We presented evidence that abundance of miRNAs in the same clusters were highly correlated and those miRNAs exerted cooperative repressive effects on target genes in human tissues. By transfecting miRNAs into human and fly cells and extensively profiling the transcriptome alteration with deep-sequencing, we further demonstrated the functional co-adaptation between new and old miRNAs in the miR-17–92 cluster. Our population genomic analysis suggest that positive Darwinian selection might be the driving force underlying the formation and evolution of miRNA clustering. Our model provided novel insights into mechanisms and evolutionary significance of miRNA clustering. PMID:27189568

  16. Recombination and spontaneous mutation at the major cluster of resistance genes in lettuce (Lactuca sativa).

    PubMed Central

    Chin, D B; Arroyo-Garcia, R; Ochoa, O E; Kesseli, R V; Lavelle, D O; Michelmore, R W

    2001-01-01

    Two sets of overlapping experiments were conducted to examine recombination and spontaneous mutation events within clusters of resistance genes in lettuce. Multiple generations were screened for recombinants using PCR-based markers flanking Dm3. The Dm3 region is not highly recombinagenic, exhibiting a recombination frequency 18-fold lower than the genome average. Recombinants were identified only rarely within the cluster of Dm3 homologs and no crossovers within genes were detected. Three populations were screened for spontaneous mutations in downy mildew resistance. Sixteen Dm mutants were identified corresponding to spontaneous mutation rates of 10(-3) to 10(-4) per generation for Dm1, Dm3, and Dm7. All mutants carried single locus, recessive mutations at the corresponding Dm locus. Eleven of the 12 Dm3 mutations were associated with large chromosome deletions. When recombination could be analyzed, deletion events were associated with exchange of flanking markers, consistent with unequal crossing over; however, although the number of Dm3 paralogs was changed, no novel chimeric genes were detected. One mutant was the result of a gene conversion event between Dm3 and a closely related homolog, generating a novel chimeric gene. In two families, spontaneous deletions were correlated with elevated levels of recombination. Therefore, the short-term evolution of the major cluster of resistance genes in lettuce involves several genetic mechanisms including unequal crossing over and gene conversion. PMID:11157000

  17. DNase I hypersensitive sites within the inducible qa gene cluster of Neurospora crassa.

    PubMed Central

    Baum, J A; Giles, N H

    1986-01-01

    DNase I hypersensitive regions were mapped within the 17.3-kilobase qa (quinic acid) gene cluster of Neurospora crassa. The 5'-flanking regions of the five qa structural genes and the two qa regulatory genes each contain DNase I hypersensitive sites under noninducing conditions and generally exhibit increases in DNase I cleavage upon induction of transcription with quinic acid. The two large intergenic regions of the qa gene cluster appear to be similarly organized with respect to the positions of constitutive and inducible DNase I hypersensitive sites. Inducible hypersensitive sites on the 5' side of one qa gene, qa-x, appear to be differentially regulated. Employing these and previously published data, we have identified a conserved sequence element that may mediate the activator function of the qa-1F regulatory gene. Variants of the 16-base-pair consensus sequence are consistently found within DNase I-protected regions adjacent to inducible DNase I hypersensitive sites within the gene cluster. Images PMID:2944110

  18. Copy Number Variants in the Kallikrein Gene Cluster

    PubMed Central

    Lindahl, Pernilla; Säll, Torbjörn; Bjartell, Anders; Johansson, Anna M.; Lilja, Hans; Halldén, Christer

    2013-01-01

    The kallikrein gene family (KLK1-KLK15) is the largest contiguous group of protease genes within the human genome and is associated with both risk and outcome of cancer and other diseases. We searched for copy number variants in all KLK genes using quantitative PCR analysis and analysis of inheritance patterns of single nucleotide polymorphisms. Two deletions were identified: one 2235-bp deletion in KLK9 present in 1.2% of alleles, and one 3394-bp deletion in KLK15 present in 4.0% of alleles. Each deletion eliminated one complete exon and created out-of-frame coding that eliminated the catalytic triad of the resulting truncated gene product, which therefore likely is a non-functional protein. Deletion breakpoints identified by DNA sequencing located the KLK9 deletion breakpoint to a long interspersed element (LINE) repeated sequence, while the deletion in KLK15 is located in a single copy sequence. To search for an association between each deletion and risk of prostate cancer (PC), we analyzed a cohort of 667 biopsied men (266 PC cases and 401 men with no evidence of PC at biopsy) using short deletion-specific PCR assays. There was no association between evidence of PC in this cohort and the presence of either gene deletion. Haplotyping revealed a single origin of each deletion, with most recent common ancestor estimates of 3000-8000 and 6000-14 000 years for the deletions in KLK9 and KLK15, respectively. The presence of the deletions on the same haplotypes in 1000 Genomes data of both European and African populations indicate an early origin of both deletions. The old age in combination with homozygous presence of loss-of-function variants suggests that some kallikrein-related peptidases have non-essential functions. PMID:23894413

  19. Intact cluster and chordate-like expression of ParaHox genes in a sea star

    PubMed Central

    2013-01-01

    Background The ParaHox genes are thought to be major players in patterning the gut of several bilaterian taxa. Though this is a fundamental role that these transcription factors play, their activities are not limited to the endoderm and extend to both ectodermal and mesodermal tissues. Three genes compose the ParaHox group: Gsx, Xlox and Cdx. In some taxa (mostly chordates but to some degree also in protostomes) the three genes are arranged into a genomic cluster, in a similar fashion to what has been shown for the better-known Hox genes. Sea urchins possess the full complement of ParaHox genes but they are all dispersed throughout the genome, an arrangement that, perhaps, represented the primitive condition for all echinoderms. In order to understand the evolutionary history of this group of genes we cloned and characterized all ParaHox genes, studied their expression patterns and identified their genomic loci in a member of an earlier branching group of echinoderms, the asteroid Patiria miniata. Results We identified the three ParaHox orthologs in the genome of P. miniata. While one of them, PmGsx is provided as maternal message, with no zygotic activation afterwards, the other two, PmLox and PmCdx are expressed during embryogenesis, within restricted domains of both endoderm and ectoderm. Screening of a Patiria bacterial artificial chromosome (BAC) library led to the identification of a clone containing the three genes. The transcriptional directions of PmGsx and PmLox are opposed to that of the PmCdx gene within the cluster. Conclusions The identification of P. miniata ParaHox genes has revealed the fact that these genes are clustered in the genome, in contrast to what has been reported for echinoids. Since the presence of an intact cluster, or at least a partial cluster, has been reported in chordates and polychaetes respectively, it becomes clear that within echinoderms, sea urchins have modified the original bilaterian arrangement. Moreover, the sea star

  20. Hybrid coexpression link similarity graph clustering for mining biological modules from multiple gene expression datasets

    PubMed Central

    2014-01-01

    Background Advances in genomic technologies have enabled the accumulation of vast amount of genomic data, including gene expression data for multiple species under various biological and environmental conditions. Integration of these gene expression datasets is a promising strategy to alleviate the challenges of protein functional annotation and biological module discovery based on a single gene expression data, which suffers from spurious coexpression. Results We propose a joint mining algorithm that constructs a weighted hybrid similarity graph whose nodes are the coexpression links. The weight of an edge between two coexpression links in this hybrid graph is a linear combination of the topological similarities and co-appearance similarities of the corresponding two coexpression links. Clustering the weighted hybrid similarity graph yields recurrent coexpression link clusters (modules). Experimental results on Human gene expression datasets show that the reported modules are functionally homogeneous as evident by their enrichment with biological process GO terms and KEGG pathways. PMID:25221624

  1. Co-expression of TAL1 and ADH1 in recombinant xylose-fermenting Saccharomyces cerevisiae improves ethanol production from lignocellulosic hydrolysates in the presence of furfural.

    PubMed

    Hasunuma, Tomohisa; Ismail, Ku Syahidah Ku; Nambu, Yumiko; Kondo, Akihiko

    2014-02-01

    Lignocellulosic biomass dedicated to bioethanol production usually contains pentoses and inhibitory compounds such as furfural that are not well tolerated by Saccharomyces cerevisiae. Thus, S. cerevisiae strains with the capability of utilizing both glucose and xylose in the presence of inhibitors such as furfural are very important in industrial ethanol production. Under the synergistic conditions of transaldolase (TAL) and alcohol dehydrogenase (ADH) overexpression, S. cerevisiae MT8-1X/TAL-ADH was able to produce 1.3-fold and 2.3-fold more ethanol in the presence of 70 mM furfural than a TAL-expressing strain and a control strain, respectively. We also tested the strains' ability by mimicking industrial ethanol production from hemicellulosic hydrolysate containing fermentation inhibitors, and ethanol production was further improved by 16% when using MT8-1X/TAL-ADH compared to the control strain. Transcript analysis further revealed that besides the pentose phosphate pathway genes TKL1 and TAL1, ADH7 was also upregulated in response to furfural stress, which resulted in higher ethanol production compared to the TAL-expressing strain. The improved capability of our modified strain was based on its capacity to more quickly reduce furfural in situ resulting in higher ethanol production. The co-expression of TAL/ADH genes is one crucial strategy to fully utilize undetoxified lignocellulosic hydrolysate, leading to cost-competitive ethanol production.

  2. Identification of a gene cluster associated with triclosan catabolism.

    PubMed

    Kagle, Jeanne M; Paxson, Clayton; Johnstone, Precious; Hay, Anthony G

    2015-06-01

    Aerobic degradation of bis-aryl ethers like the antimicrobial triclosan typically proceeds through oxygenase-dependent catabolic pathways. Although several studies have reported on bacteria capable of degrading triclosan aerobically, there are no reports describing the genes responsible for this process. In this study, a gene encoding the large subunit of a putative triclosan oxygenase, designated tcsA was identified in a triclosan-degrading fosmid clone from a DNA library of Sphingomonas sp. RD1. Consistent with tcsA's similarity to two-part dioxygenases, a putative FMN-dependent ferredoxin reductase, designated tcsB was found immediately downstream of tcsA. Both tcsAB were found in the midst of a putative chlorocatechol degradation operon. We show that RD1 produces hydroxytriclosan and chlorocatechols during triclosan degradation and that tcsA is induced by triclosan. This is the first study to report on the genetics of triclosan degradation.

  3. Teaching Gene Technology in an Outreach Lab: Students' Assigned Cognitive Load Clusters and the Clusters' Relationships to Learner Characteristics, Laboratory Variables, and Cognitive Achievement

    ERIC Educational Resources Information Center

    Scharfenberg, Franz-Josef; Bogner, Franz X.

    2013-01-01

    This study classified students into different cognitive load (CL) groups by means of cluster analysis based on their experienced CL in a gene technology outreach lab which has instructionally been designed with regard to CL theory. The relationships of the identified student CL clusters to learner characteristics, laboratory variables, and…

  4. The Response of Maize Seedlings of Different Ages to Hypoxic and Anoxic Stress (Changes in Induction of Adh1 mRNA, ADH Activity, and Survival of Anoxia).

    PubMed Central

    Andrews, D. L.; Drew, M. C.; Johnson, J. R.; Cobb, B. G.

    1994-01-01

    Previously we showed that there is only a transient induction of alcohol dehydrogenase 1 (Adh1) transcripts and only a small induction of alcohol dehydrogenase (ADH) enzyme activity in root tips of maize (Zea mays L.) seedlings subjected to strict anaerobiosis without prior acclimation by exposure to low O2 (D.L. Andrews, B.G. Cobb, J.R. Johnson, M.C. Drew [1993] Plant Physiol 101: 403-414). Acclimation of root tips of seedlings by low O2 before anoxia appeared to be necessary for full induction of ADH. Here we have examined the effect of seedling age on changes in the protein content, induction of Adh1 transcripts, and ADH enzyme activity in 5-mm root tips, root axes, and shoots of maize (cv TX5855). Their ability to survive anoxia was also recorded. Some seedlings were sparged with 4% O2 for 6 or 18 h (a hypoxic pretreatment) followed by anoxia (sparged with N2) for up to 48 h. Other seedlings were not acclimated before anoxia. In general, younger seedlings had higher initial (aerobic) levels of total protein, Adh1 transcripts, and ADH activity than did seedlings that were 2 d older. For younger seedlings, anoxia alone induced Adh1 transcripts, which reached a peak within 6 to 12 h, whereas ADH activity increased throughout the 48-h treatment. For older seedlings, anoxia caused only a small, transient induction of Adh1 transcripts or ADH activity. For seedlings of either age, hypoxia induced Adh1 transcripts and ADH activity, both of which were increased further by subsequent anoxia in the younger seedlings but to a lesser extent in the older seedlings. Despite differences in ADH activity, roots of seedlings of either age showed a similar resistance to anoxia. Thus, acclimation of maize seedlings to survive anoxia does not appear to be related to induction of high levels of ADH activity. PMID:12232185

  5. Regulated Expression of Three Alcohol Dehydrogenase Genes in Barley Aleurone Layers 1

    PubMed Central

    Hanson, Andrew D.; Jacobsen, John V.; Zwar, John A.

    1984-01-01

    Three genes specify alcohol dehydrogenase (EC 1.1.1.1.; ADH) enzymes in barley (Hordeum vulgare L.) (Adh 1, Adh 2, and Adh 3). Their polypeptide products (ADH 1, ADH 2, ADH 3) dimerize to give a total of six ADH isozymes which can be resolved by native gel electrophoresis and stained for enzyme activity. Under fully aerobic conditions, aleurone layers of cv Himalaya had a high titer of a single isozyme, the homodimer containing ADH 1 monomers. This isozyme was accumulated by the aleurone tissue during the later part of seed development, and survived seed drying and rehydration. The five other possible ADH isozymes were induced by O2 deficit. The staining of these five isozymes on electrophoretic gels increased progressively in intensity as O2 levels were reduced below 5%, and were most intense at 0% O2. In vivo35S labeling and specific immunoprecipitation of ADH peptides, followed by isoelectric focusing of the ADH peptides in the presence of 8 molar urea (urea-IEF) demonstrated the following. (a) Aleurone layers incubated in air synthesized ADH 1 and a trace of ADH 2; immature layers from developing seeds behaved similarly. (b) At 5% O2, synthesis of ADH 2 increased and ADH 3 appeared. (c) At 2% and 0% O2, the synthesis of all three ADH peptides increased markedly. Cell-free translation of RNA isolated from aleurone layers, followed by immunoprecipitation and urea-IEF of in vitro synthesized ADH peptides, showed that levels of mRNA for all three ADH peptides rose sharply during 1 day of O2 deprivation. Northern hybridizations with a maize Adh 2 cDNA clone established that the clone hybridized with barley mRNA comparable in size to maize Adh 2 mRNA, and that the level of this barley mRNA increased 15- to 20-fold after 1 day at 5% or 2% O2, and about 100-fold after 1 day at 0% O2. We conclude that in aleurone layers, expression of the three barley Adh genes is maximal in the absence of O2, that regulation of mRNA level is likely to be a major controlling factor, and

  6. In vitro expression of Candida albicans alcohol dehydrogenase genes involved in acetaldehyde metabolism.

    PubMed

    Bakri, M M; Rich, A M; Cannon, R D; Holmes, A R

    2015-02-01

    Alcohol consumption is a risk factor for oral cancer, possibly via its conversion to acetaldehyde, a known carcinogen. The oral commensal yeast Candida albicans may be one of the agents responsible for this conversion intra-orally. The alcohol dehydrogenase (Adh) family of enzymes are involved in acetaldehyde metabolism in yeast but, for C. albicans it is not known which family member is responsible for the conversion of ethanol to acetaldehyde. In this study we determined the expression of mRNAs from three C. albicans Adh genes (CaADH1, CaADH2 and CaCDH3) for cells grown in different culture media at different growth phases by Northern blot analysis and quantitative reverse transcription polymerase chain reaction. CaADH1 was constitutively expressed under all growth conditions but there was differential expression of CaADH2. CaADH3 expression was not detected. To investigate whether CaAdh1p or CaAdh2p can contribute to alcohol catabolism in C. albicans, each gene from the reference strain C. albicans SC5314 was expressed in Saccharomyces cerevisiae. Cell extracts from an CaAdh1p-expressing S. cerevisiae recombinant, but not an CaAdh2p-expressing recombinant, or an empty vector control strain, possessed ethanol-utilizing Adh activity above endogenous S. cerevisiae activity. Furthermore, expression of C. albicans Adh1p in a recombinant S. cerevisiae strain in which the endogenous ScADH2 gene (known to convert ethanol to acetaldehyde in this yeast) had been deleted, conferred an NAD-dependent ethanol-utilizing, and so acetaldehyde-producing, Adh activity. We conclude that CaAdh1p is the enzyme responsible for ethanol use under in vitro growth conditions, and may contribute to the intra-oral production of acetaldehyde.

  7. Diversity and depth-specific distribution of SAR11 cluster rRNA genes from marine planktonic bacteria.

    PubMed Central

    Field, K G; Gordon, D; Wright, T; Rappé, M; Urback, E; Vergin, K; Giovannoni, S J

    1997-01-01

    Small-subunit (SSU) ribosomal DNA (rDNA) gene clusters are phylogenetically related sets of SSU rRNA genes, commonly encountered in genes amplified from natural populations. Genetic variability in gene clusters could result from artifacts (polymerase error or PCR chimera formation), microevolution (variation among rrn copies within strains), or macroevolution (genetic divergence correlated with long-term evolutionary divergence). To better understand gene clusters this study assessed genetic diversity and distribution of a single environmental SSU rDNA gene cluster, the SAR11 cluster. SAR11 cluster genes, from an uncultured group of the alpha subclass of the class Proteobacteria, have been recovered from coastal and midoceanic waters of the North Atlantic and Pacific. We cloned and bidirectionally sequenced 23 new SAR11 cluster 16S rRNA genes, from 80 and 250 m in the Sargasso Sea and from surface coastal waters of the Atlantic and Pacific, and analyzed them with previously published sequences. Two SAR11 genes were obviously PCR chimeras, but the biological (nonchimeric) origins of most subgroups within the cluster were confirmed by independent recovery from separate gene libraries. Using group-specific oligonucleotide probes, we analyzed depth profiles of nucleic acids, targeting both amplified rDNAs and bulk RNAs. Two subgroups within the SAR11 cluster showed different highly depth-specific distributions. We conclude that some of the genetic diversity within the SAR11 gene cluster represents macroevolutionary divergence correlated with niche specialization. Furthermore, we demonstrate the utility for marine microbial ecology of oligonucleotide probes based on gene sequences amplified from natural populations and show that a detailed knowledge of sequence variability may be needed to effectively design these probes. PMID:8979340

  8. Diversity and depth-specific distribution of SAR11 cluster rRNA genes from marine planktonic bacteria

    SciTech Connect

    Field, K.G.; Gordon, D.; Wright, T.

    1997-01-01

    Small-subunit (SSU) ribosomal DNA (rDNA) gene clusters are phylogenetically related sets of SSU rRNA genes, commonly encountered in genes amplified from natural populations. Genetic variability in gene clusters could result form artifacts (polymerase error or PCR chimera formation), microevolution (variation among rrn copies within strains), or macroevolution (genetic divergence correlated with long-term evolutionary divergence). To better understand gene clusters, this study assessed genetic diversity and distribution of a single environmental SSU rDNA gene cluster, the SAR11 cluster. SAR11 cluster genes, from an uncultured group of the {alpha} subclass of the class Proteobacteria, have been recovered from coastal and midoceanic waters of the North Atlantic and Pacific. We cloned and bidirectionally sequenced 23 new SAR11 cluster 16S rRNA genes, from 80 and 250 m im the Sargasso Sea and from surface coastal waters of the Atlantic and Pacific, and analyzed them with previously published sequences. Two SAR11 genes were obviously PCR chimeras, but the biological (nonchimeric) origins of most subgroups within the cluster were confirmed by independent recovery from separate gene libraries. Using group-specific oligonucleotide probes, we analyzed depth profiles of nucleic acids, targeting both amplified rDNAs and bulk RNAs. Two subgroups within the SAR11 cluster showed different highly depth-specific distributions. We conclude that some of the genetic diversity within the SAR11 gene cluster represents macroevolutionary divergence correlated with niche specialization. Furthermore, we demonstrate the utility for marine microbial ecology of oligonucleotide probes based on gene sequences amplified from natural populations and show that a detailed knowledge of sequence variability may be needed to effectively design these probes. 48 refs., 7 figs., 3 tabs.

  9. The Magea gene cluster regulates male germ cell apoptosis without affecting the fertility in mice.

    PubMed

    Hou, Siyuan; Xian, Li; Shi, Peiliang; Li, Chaojun; Lin, Zhaoyu; Gao, Xiang

    2016-01-01

    While apoptosis is essential for male germ cell development, improper activation of apoptosis in the testis can affect spermatogenesis and cause reproduction defects. Members of the MAGE-A (melanoma antigen family A) gene family are frequently clustered in mammalian genomes and are exclusively expressed in the testes of normal animals but abnormally activated in a wide variety of cancers. We investigated the potential roles of these genes in spermatogenesis by generating a mouse model with a 210-kb genomic deletion encompassing six members of the Magea gene cluster (Magea1, Magea2, Magea3, Magea5, Magea6 and Magea8). Male mice carrying the deletion displayed smaller testes from 2 months old with a marked increase in apoptotic germ cells in the first wave of spermatogenesis. Furthermore, we found that Magea genes prevented stress-induced spermatogenic apoptosis after N-ethyl-N-nitrosourea (ENU) treatment during the adult stage. Mechanistically, deletion of the Magea gene cluster resulted in a dramatic increase in apoptotic germ cells, predominantly spermatocytes, with activation of p53 and induction of Bax in the testes. These observations demonstrate that the Magea genes are crucial in maintaining normal testicular size and protecting germ cells from excessive apoptosis under genotoxic stress. PMID:27226137

  10. The Magea gene cluster regulates male germ cell apoptosis without affecting the fertility in mice

    PubMed Central

    Hou, Siyuan; Xian, Li; Shi, Peiliang; Li, Chaojun; Lin, Zhaoyu; Gao, Xiang

    2016-01-01

    While apoptosis is essential for male germ cell development, improper activation of apoptosis in the testis can affect spermatogenesis and cause reproduction defects. Members of the MAGE-A (melanoma antigen family A) gene family are frequently clustered in mammalian genomes and are exclusively expressed in the testes of normal animals but abnormally activated in a wide variety of cancers. We investigated the potential roles of these genes in spermatogenesis by generating a mouse model with a 210-kb genomic deletion encompassing six members of the Magea gene cluster (Magea1, Magea2, Magea3, Magea5, Magea6 and Magea8). Male mice carrying the deletion displayed smaller testes from 2 months old with a marked increase in apoptotic germ cells in the first wave of spermatogenesis. Furthermore, we found that Magea genes prevented stress-induced spermatogenic apoptosis after N-ethyl-N-nitrosourea (ENU) treatment during the adult stage. Mechanistically, deletion of the Magea gene cluster resulted in a dramatic increase in apoptotic germ cells, predominantly spermatocytes, with activation of p53 and induction of Bax in the testes. These observations demonstrate that the Magea genes are crucial in maintaining normal testicular size and protecting germ cells from excessive apoptosis under genotoxic stress. PMID:27226137

  11. Rotation of photoreceptor clusters in the developing Drosophila eye requires the nemo gene.

    PubMed

    Choi, K W; Benzer, S

    1994-07-15

    The Drosophila eye consists of a reiterative hexagonal array of photoreceptor cell clusters, the ommatidia. During normal morphogenesis, the clusters in the dorsal or ventral halves of the disc rotate 90 degrees in opposite directions, forming mirror images across a dorsoventral equator. In the mutant nemo (nmo), there is an initial turning of approximately 45 degrees, but further rotation is blocked. Genetic mosaic analysis indicates that the nmo gene acts upon each cluster as a whole; normal nmo function in one or more photoreceptor cells appears to be sufficient to induce full rotation. The nmo gene sequence encodes a serine/threonine protein kinase homolog, suggesting that the kinase is required to initiate the second step of rotation. In another mutant, roulette, excessive rotation through varying angles occurs in many ommatidia. This defect is suppressed by nmo, indicating that nmo acts upstream in a rotation-regulating pathway.

  12. TreeParser-Aided Klee Diagrams Display Taxonomic Clusters in DNA Barcode and Nuclear Gene Datasets

    PubMed Central

    Stoeckle, Mark Y.; Coffran, Cameron

    2013-01-01

    Indicator vector analysis of a nucleotide sequence alignment generates a compact heat map, called a Klee diagram, with potential insight into clustering patterns in evolution. However, so far this approach has examined only mitochondrial cytochrome c oxidase I (COI) DNA barcode sequences. To further explore, we developed TreeParser, a freely-available web-based program that sorts a sequence alignment according to a phylogenetic tree generated from the dataset. We applied TreeParser to nuclear gene and COI barcode alignments from birds and butterflies. Distinct blocks in the resulting Klee diagrams corresponded to species and higher-level taxonomic divisions in both groups, and this enabled graphic comparison of phylogenetic information in nuclear and mitochondrial genes. Our results demonstrate TreeParser-aided Klee diagrams objectively display taxonomic clusters in nucleotide sequence alignments. This approach may help establish taxonomy in poorly studied groups and investigate higher-level clustering which appears widespread but not well understood. PMID:24022383

  13. TreeParser-aided Klee diagrams display taxonomic clusters in DNA barcode and nuclear gene datasets.

    PubMed

    Stoeckle, Mark Y; Coffran, Cameron

    2013-01-01

    Indicator vector analysis of a nucleotide sequence alignment generates a compact heat map, called a Klee diagram, with potential insight into clustering patterns in evolution. However, so far this approach has examined only mitochondrial cytochrome c oxidase I (COI) DNA barcode sequences. To further explore, we developed TreeParser, a freely-available web-based program that sorts a sequence alignment according to a phylogenetic tree generated from the dataset. We applied TreeParser to nuclear gene and COI barcode alignments from birds and butterflies. Distinct blocks in the resulting Klee diagrams corresponded to species and higher-level taxonomic divisions in both groups, and this enabled graphic comparison of phylogenetic information in nuclear and mitochondrial genes. Our results demonstrate TreeParser-aided Klee diagrams objectively display taxonomic clusters in nucleotide sequence alignments. This approach may help establish taxonomy in poorly studied groups and investigate higher-level clustering which appears widespread but not well understood.

  14. Identification of the Viridicatumtoxin and Griseofulvin Gene Clusters from Penicillium aethiopicum

    PubMed Central

    Chooi, Yit-Heng; Cacho, Ralph; Tang, Yi

    2010-01-01

    SUMMARY Penicillium aethiopicum produces two structurally interesting and biologically active polyketides: the tetracycline-like viridicatumtoxin 1 and the classic antifungal agent griseofulvin 2. Here, we report the concurrent discovery of the two corresponding biosynthetic gene clusters (vrt and gsf) by 454 shotgun sequencing. Gene deletions confirmed two nonreducing PKSs (NRPKS), vrtA and gsfA, are required for the biosynthesis of 1 and 2, respectively. Both PKSs share similar domain architectures and lack a C-terminal thioesterase domain. We identified gsfI as the chlorinase involved in the biosynthesis of 2, as deletion of gsfI resulted in the accumulation of decholorogriseofulvin 3. Comparative analysis with the P. chrysogenum genome revealed that both clusters are embedded within conserved syntenic regions of P. aethiopicum chromosomes. Discovery of the vrt and gsf clusters provided the basis for genetic and biochemical studies of the pathways. PMID:20534346

  15. Regulation of alkyl-dihydrothiazole-carboxylates (ATCs) by iron and the pyochelin gene cluster in Pseudomonas aeruginosa.

    PubMed

    Vinayavekhin, Nawaporn; Saghatelian, Alan

    2009-08-21

    Using the pyochelin (pch) gene cluster as an example, we demonstrate the utility of untargeted metabolomics in the discovery and characterization of secondary metabolites regulated by biosynthetic gene clusters. Comparison of the extracellular metabolomes of pch gene cluster mutants to the wild-type Pseudomonas aeruginosa (strain PA 14) identified 198 ions regulated by the pch genes. In addition to known metabolites, we characterized the structure of a pair of novel metabolites regulated by the pch gene cluster as 2-alkyl-4,5-dihydrothiazole-4-carboxylates (ATCs), using a combination of mass spectrometry, chemical synthesis, and stable isotope labeling. Subsequent assays revealed that ATCs bind iron and are regulated by iron levels in the media in a similar fashion as other metabolites associated with the pch gene cluster. Further genetic complementation and overexpression analyses of the pch genes revealed ATC production to be dependent on the pchE gene in the pch gene cluster. Overall, these studies highlight the ability of untargeted metabolomics to reveal regulatory connections between gene clusters and secondary metabolites, including novel metabolites. PMID:19621937

  16. Resolving misassembled cattle immune gene clusters with hierarchical, long read sequencing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Animal health is a critical component of productivity; however, current genomic selection genotyping tools have a paucity of genetic markers within key immune gene clusters (IGC) involved in the cattle innate and adaptive immune systems. With diseases such as Bovine Tuberculosis and Johne’s disease ...

  17. Expanded Natural Product Diversity Revealed by Analysis of Lanthipeptide-Like Gene Clusters in Actinobacteria

    PubMed Central

    Zhang, Qi; Doroghazi, James R.; Zhao, Xiling; Walker, Mark C.

    2015-01-01

    Lanthionine-containing peptides (lanthipeptides) are a rapidly growing family of polycyclic peptide natural products belonging to the large class of ribosomally synthesized and posttranslationally modified peptides (RiPPs). Lanthipeptides are widely distributed in taxonomically distant species, and their currently known biosynthetic systems and biological activities are diverse. Building on the recent natural product gene cluster family (GCF) project, we report here large-scale analysis of lanthipeptide-like biosynthetic gene clusters from Actinobacteria. Our analysis suggests that lanthipeptide biosynthetic pathways, and by extrapolation the natural products themselves, are much more diverse than currently appreciated and contain many different posttranslational modifications. Furthermore, lanthionine synthetases are much more diverse in sequence and domain topology than currently characterized systems, and they are used by the biosynthetic machineries for natural products other than lanthipeptides. The gene cluster families described here significantly expand the chemical diversity and biosynthetic repertoire of lanthionine-related natural products. Biosynthesis of these novel natural products likely involves unusual and unprecedented biochemistries, as illustrated by several examples discussed in this study. In addition, class IV lanthipeptide gene clusters are shown not to be silent, setting the stage to investigate their biological activities. PMID:25888176

  18. Characterization of the complete zwittermicin A biosynthesis gene cluster from Bacillus cereus.

    PubMed

    Kevany, Brian M; Rasko, David A; Thomas, Michael G

    2009-02-01

    Bacillus cereus UW85 produces the linear aminopolyol antibiotic zwittermicin A (ZmA). This antibiotic has diverse biological activities, such as suppression of disease in plants caused by protists, inhibition of fungal and bacterial growth, and amplification of the insecticidal activity of the toxin protein from Bacillus thuringiensis. ZmA has an unusual chemical structure that includes a d amino acid and ethanolamine and glycolyl moieties, as well as having an unusual terminal amide that is generated from the modification of the nonproteinogenic amino acid beta-ureidoalanine. The diverse biological activities and unusual structure of ZmA have stimulated our efforts to understand how this antibiotic is biosynthesized. Here, we present the identification of the complete ZmA biosynthesis gene cluster from B. cereus UW85. A nearly identical gene cluster is identified on a plasmid from B. cereus AH1134, and we show that this strain is also capable of producing ZmA. Bioinformatics and biochemical analyses of the ZmA biosynthesis enzymes strongly suggest that ZmA is initially biosynthesized as part of a larger metabolite that is processed twice, resulting in the formation of ZmA and two additional metabolites. Additionally, we propose that the biosynthesis gene cluster for the production of the amino sugar kanosamine is contained within the ZmA biosynthesis gene cluster in B. cereus UW85.

  19. Functional Analysis of the Yeast Glc7-Binding Protein Reg1 Identifies a Protein Phosphatase Type 1-Binding Motif as Essential for Repression of ADH2 Expression

    PubMed Central

    Dombek, Kenneth M.; Voronkova, Valentina; Raney, Alexa; Young, Elton T.

    1999-01-01

    In Saccharomyces cerevisiae, the protein phosphatase type 1 (PP1)-binding protein Reg1 is required to maintain complete repression of ADH2 expression during growth on glucose. Surprisingly, however, mutant forms of the yeast PP1 homologue Glc7, which are unable to repress expression of another glucose-regulated gene, SUC2, fully repressed ADH2. Constitutive ADH2 expression in reg1 mutant cells did require Snf1 protein kinase activity like constitutive SUC2 expression and was inhibited by unregulated cyclic AMP-dependent protein kinase activity like ADH2 expression in derepressed cells. To further elucidate the functional role of Reg1 in repressing ADH2 expression, deletions scanning the entire length of the protein were analyzed. Only the central region of the protein containing the putative PP1-binding sequence RHIHF was found to be indispensable for repression. Introduction of the I466M F468A substitutions into this sequence rendered Reg1 almost nonfunctional. Deletion of the central region or the double substitution prevented Reg1 from significantly interacting with Glc7 in two-hybrid analyses. Previous experimental evidence had indicated that Reg1 might target Glc7 to nuclear substrates such as the Snf1 kinase complex. Subcellular localization of a fully functional Reg1-green fluorescent protein fusion, however, indicated that Reg1 is cytoplasmic and excluded from the nucleus independently of the carbon source. When the level of Adr1 was modestly elevated, ADH2 expression was no longer fully repressed in glc7 mutant cells, providing the first direct evidence that Glc7 can repress ADH2 expression. These results suggest that the Reg1-Glc7 phosphatase is a cytoplasmic component of the machinery responsible for returning Snf1 kinase activity to its basal level and reestablishing glucose repression. This implies that the activated form of the Snf1 kinase complex must cycle between the nucleus and the cytoplasm. PMID:10454550

  20. Following tetraploidy in an Arabidopsis ancestor, genes were removed preferentially from one homeolog leaving clusters enriched in dose-sensitive genes.

    PubMed

    Thomas, Brian C; Pedersen, Brent; Freeling, Michael

    2006-07-01

    Approximately 90% of Arabidopsis' unique gene content is found in syntenic blocks that were formed during the most recent whole-genome duplication. Within these blocks, 28.6% of the genes have a retained pair; the remaining genes have been lost from one of the homeologs. We create a minimized genome by condensing local duplications to one gene, removing transposons, and including only genes within blocks defined by retained pairs. We use a moving average of retained and non-retained genes to find clusters of retention and then identify the types of genes that appear in clusters at frequencies above expectations. Significant clusters of retention exist for almost all chromosomal segments. Detailed alignments show that, for 85% of the genome, one homeolog was preferentially (1.6x) targeted for fractionation. This homeolog fractionation bias suggests an epigenetic mechanism. We find that islands of retention contain "connected genes," those genes predicted-by the gene balance hypothesis-to be resistant to removal because the products they encode interact with other products in a dose-sensitive manner, creating a web of dependency. Gene families that are overrepresented in clusters include those encoding components of the proteasome/protein modification complexes, signal transduction machinery, ribosomes, and transcription factor complexes. Gene pair fractionation following polyploidy or segmental duplication leaves a genome enriched for "connected" genes. These clusters of duplicate genes may help explain the evolutionary origin of coregulated chromosomal regions and new functional modules. PMID:16760422

  1. Following tetraploidy in an Arabidopsis ancestor, genes were removed preferentially from one homeolog leaving clusters enriched in dose-sensitive genes.

    PubMed

    Thomas, Brian C; Pedersen, Brent; Freeling, Michael

    2006-07-01

    Approximately 90% of Arabidopsis' unique gene content is found in syntenic blocks that were formed during the most recent whole-genome duplication. Within these blocks, 28.6% of the genes have a retained pair; the remaining genes have been lost from one of the homeologs. We create a minimized genome by condensing local duplications to one gene, removing transposons, and including only genes within blocks defined by retained pairs. We use a moving average of retained and non-retained genes to find clusters of retention and then identify the types of genes that appear in clusters at frequencies above expectations. Significant clusters of retention exist for almost all chromosomal segments. Detailed alignments show that, for 85% of the genome, one homeolog was preferentially (1.6x) targeted for fractionation. This homeolog fractionation bias suggests an epigenetic mechanism. We find that islands of retention contain "connected genes," those genes predicted-by the gene balance hypothesis-to be resistant to removal because the products they encode interact with other products in a dose-sensitive manner, creating a web of dependency. Gene families that are overrepresented in clusters include those encoding components of the proteasome/protein modification complexes, signal transduction machinery, ribosomes, and transcription factor complexes. Gene pair fractionation following polyploidy or segmental duplication leaves a genome enriched for "connected" genes. These clusters of duplicate genes may help explain the evolutionary origin of coregulated chromosomal regions and new functional modules.

  2. Organization of the qa Gene Cluster in NEUROSPORA CRASSA: Direction of Transcription of the qa-3 Gene

    PubMed Central

    Strøman, Per; Reinert, William; Case, Mary E.; Giles, Norman H.

    1979-01-01

    In Neurospora crassa, the enzyme quinate (shikimate) dehydrogenase catalyzes the first reaction in the inducible quinic acid catabolic pathway and is encoded in the qa-3 gene of the qa cluster. In this cluster, the order of genes has been established as qa-1 qa-3 qa-4 qa-2. Amino-terminal sequences have been determined for purified quinate dehydrogenase from wild type and from UV-induced revertants in two different qa-3 mutants. These two mutants (M16 and M45) map at opposite ends of the qa-3 locus. In addition, mapping data (Case et al. 1978) indicate that the end of the qa-3 gene specified by M45 is closer to the adjacent qa-1 gene than is the end specified by the M16 mutant site. In one of the revertants (R45 from qa-3 mutant M45), the aminoterminal sequence for the first ten amino acids is identical to that of wild type. The other revertant (R1 from qa-3 mutant M16) differs from wild type at the amino-terminal end by a single altered residue at position three in the sequence. The observed change involves the substitution of an isoleucine in M16-R1 for a proline in wild type. This substitution requires a two-nucleotide change in the corresponding wild-type codon.——The combined genetic and biochemical data indicate that the qa-3 mutants M16 and M45 carry amino acid substitutions near the amino-terminal and carboxyl-terminal ends of the quinate dehydrogenase enzyme, respectively. On this basis we conclude that transcription of the qa-3 gene proceeds from the end specified by the M16 mutant site in the direction of the qa-1 gene. It appears probable that transcription is initiated from a promoter site within the qa cluster, possibly immediately adjacent to the qa-3 gene. PMID:159203

  3. Clusters of genes encoding fructan biosynthesizing enzymes in wheat and barley.

    PubMed

    Huynh, Bao-Lam; Mather, Diane E; Schreiber, Andreas W; Toubia, John; Baumann, Ute; Shoaei, Zahra; Stein, Nils; Ariyadasa, Ruvini; Stangoulis, James C R; Edwards, James; Shirley, Neil; Langridge, Peter; Fleury, Delphine

    2012-10-01

    Fructans are soluble carbohydrates with health benefits and possible roles in plant adaptation. Fructan biosynthetic genes were isolated using comparative genomics and physical mapping followed by BAC sequencing in barley. Genes encoding sucrose:sucrose 1-fructosyltransferase (1-SST), fructan:fructan 1-fructosyltransferase (1-FFT) and sucrose:fructan 6-fructosyltransferase (6-SFT) were clustered together with multiple copies of vacuolar invertase genes and a transposable element on two barley BAC. Intron-exon structures of the genes were similar. Phylogenetic analysis of the fructosyltransferases and invertases in the Poaceae showed that the fructan biosynthetic genes may have evolved from vacuolar invertases. Quantitative real-time PCR was performed using leaf RNA extracted from three wheat cultivars grown under different conditions. The 1-SST, 1-FFT and 6-SFT genes had correlated expression patterns in our wheat experiment and in existing barley transcriptome database. Single nucleotide polymorphism (SNP) markers were developed and successfully mapped to a major QTL region affecting wheat grain fructan accumulation in two independent wheat populations. The alleles controlling high- and low- fructan in parental lines were also found to be associated in fructan production in a diverse set of 128 wheat lines. To the authors' knowledge, this is the first report on the mapping and sequencing of a fructan biosynthetic gene cluster and in particular, the isolation of a novel 1-FFT gene from barley.

  4. Clusters of genes encoding fructan biosynthesizing enzymes in wheat and barley.

    PubMed

    Huynh, Bao-Lam; Mather, Diane E; Schreiber, Andreas W; Toubia, John; Baumann, Ute; Shoaei, Zahra; Stein, Nils; Ariyadasa, Ruvini; Stangoulis, James C R; Edwards, James; Shirley, Neil; Langridge, Peter; Fleury, Delphine

    2012-10-01

    Fructans are soluble carbohydrates with health benefits and possible roles in plant adaptation. Fructan biosynthetic genes were isolated using comparative genomics and physical mapping followed by BAC sequencing in barley. Genes encoding sucrose:sucrose 1-fructosyltransferase (1-SST), fructan:fructan 1-fructosyltransferase (1-FFT) and sucrose:fructan 6-fructosyltransferase (6-SFT) were clustered together with multiple copies of vacuolar invertase genes and a transposable element on two barley BAC. Intron-exon structures of the genes were similar. Phylogenetic analysis of the fructosyltransferases and invertases in the Poaceae showed that the fructan biosynthetic genes may have evolved from vacuolar invertases. Quantitative real-time PCR was performed using leaf RNA extracted from three wheat cultivars grown under different conditions. The 1-SST, 1-FFT and 6-SFT genes had correlated expression patterns in our wheat experiment and in existing barley transcriptome database. Single nucleotide polymorphism (SNP) markers were developed and successfully mapped to a major QTL region affecting wheat grain fructan accumulation in two independent wheat populations. The alleles controlling high- and low- fructan in parental lines were also found to be associated in fructan production in a diverse set of 128 wheat lines. To the authors' knowledge, this is the first report on the mapping and sequencing of a fructan biosynthetic gene cluster and in particular, the isolation of a novel 1-FFT gene from barley. PMID:22864927

  5. Clustered Transcription Factor Genes Regulate Nicotine Biosynthesis in Tobacco[W][OA

    PubMed Central

    Shoji, Tsubasa; Kajikawa, Masataka; Hashimoto, Takashi

    2010-01-01

    Tobacco (Nicotiana tabacum) synthesizes nicotine and related pyridine alkaloids in the root, and their synthesis increases upon herbivory on the leaf via a jasmonate-mediated signaling cascade. Regulatory NIC loci that positively regulate nicotine biosynthesis have been genetically identified, and their mutant alleles have been used to breed low-nicotine tobacco varieties. Here, we report that the NIC2 locus, originally called locus B, comprises clustered transcription factor genes of an ethylene response factor (ERF) subfamily; in the nic2 mutant, at least seven ERF genes are deleted altogether. Overexpression, suppression, and dominant repression experiments using transgenic tobacco roots showed both functional redundancy and divergence among the NIC2-locus ERF genes. These transcription factors recognized a GCC-box element in the promoter of a nicotine pathway gene and specifically activated all known structural genes in the pathway. The NIC2-locus ERF genes are expressed in the root and upregulated by jasmonate with kinetics that are distinct among the members. Thus, gene duplication events generated a cluster of highly homologous transcription factor genes with transcriptional and functional diversity. The NIC2-locus ERFs are close homologs of ORCA3, a jasmonate-responsive transcriptional activator of indole alkaloid biosynthesis in Catharanthus roseus, indicating that the NIC2/ORCA3 ERF subfamily was recruited independently to regulate jasmonate-inducible secondary metabolism in distinct plant lineages. PMID:20959558

  6. Biosynthetic Gene Cluster for Surugamide A Encompasses an Unrelated Decapeptide, Surugamide F.

    PubMed

    Ninomiya, Akihiro; Katsuyama, Yohei; Kuranaga, Takefumi; Miyazaki, Masayuki; Nogi, Yuichi; Okada, Shigeru; Wakimoto, Toshiyuki; Ohnishi, Yasuo; Matsunaga, Shigeki; Takada, Kentaro

    2016-09-15

    Genome mining is a powerful method for finding novel secondary metabolites. In our study on the biosynthetic gene cluster for the cyclic octapeptides surugamides A-E (inhibitors of cathepsin B), we found a putative gene cluster consisting of four successive non-ribosomal peptide synthetase (NRPS) genes, surA, surB, surC, and surD. Prediction of amino acid sequence based on the NRPSs and gene inactivation revealed that surugamides A-E are produced by two NRPS genes, surA and surD, which were separated by two NRPS genes, surB and surC. The latter genes are responsible for the biosynthesis of an unrelated peptide, surugamide F. The pattern of intercalation observed in the sur genes is unprecedented. The structure of surugamide F, a linear decapeptide containing one 3-amino-2-methylpropionic acid (AMPA) residue, was determined by spectroscopic methods and was confirmed by solid-phase peptide synthesis. PMID:27443244

  7. Complete Genome Sequence of the Filamentous Fungus Aspergillus westerdijkiae Reveals the Putative Biosynthetic Gene Cluster of Ochratoxin A

    PubMed Central

    Chakrabortti, Alolika; Li, Jinming

    2016-01-01

    Ochratoxin A (OTA) is a common mycotoxin that contaminates food and agricultural products. Sequencing of the complete genome of Aspergillus westerdijkiae, a major producer of OTA, reveals more than 50 biosynthetic gene clusters, including a putative OTA biosynthetic gene cluster that encodes a dozen of enzymes, transporters, and regulatory proteins. PMID:27635003

  8. Complete Genome Sequence of the Filamentous Fungus Aspergillus westerdijkiae Reveals the Putative Biosynthetic Gene Cluster of Ochratoxin A.

    PubMed

    Chakrabortti, Alolika; Li, Jinming; Liang, Zhao-Xun

    2016-01-01

    Ochratoxin A (OTA) is a common mycotoxin that contaminates food and agricultural products. Sequencing of the complete genome of Aspergillus westerdijkiae, a major producer of OTA, reveals more than 50 biosynthetic gene clusters, including a putative OTA biosynthetic gene cluster that encodes a dozen of enzymes, transporters, and regulatory proteins. PMID:27635003

  9. Clustering of two genes putatively involved in cyanate detoxification evolved recently and independently in multiple fungal lineages

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fungi that have the enzymes cyanase and carbonic anhydrase show a limited capacity to detoxify cyanate, a fungicide employed by both plants and humans. Here, we describe a novel two-gene cluster that comprises duplicated cyanase and carbonic anhydrase copies, which we name the CCA gene cluster, trac...

  10. Cloning and Analysis of the Planosporicin Lantibiotic Biosynthetic Gene Cluster of Planomonospora alba

    PubMed Central

    Sherwood, Emma J.; Hesketh, Andrew R.

    2013-01-01

    The increasing prevalence of antibiotic resistance in bacterial pathogens has renewed focus on natural products with antimicrobial properties. Lantibiotics are ribosomally synthesized peptide antibiotics that are posttranslationally modified to introduce (methyl)lanthionine bridges. Actinomycetes are renowned for their ability to produce a large variety of antibiotics, many with clinical applications, but are known to make only a few lantibiotics. One such compound is planosporicin produced by Planomonospora alba, which inhibits cell wall biosynthesis in Gram-positive pathogens. Planosporicin is a type AI lantibiotic structurally similar to those which bind lipid II, the immediate precursor for cell wall biosynthesis. The gene cluster responsible for planosporicin biosynthesis was identified by genome mining and subsequently isolated from a P. alba cosmid library. A minimal cluster of 15 genes sufficient for planosporicin production was defined by heterologous expression in Nonomuraea sp. strain ATCC 39727, while deletion of the gene encoding the precursor peptide from P. alba, which abolished planosporicin production, was also used to confirm the identity of the gene cluster. Deletion of genes encoding likely biosynthetic enzymes identified through bioinformatic analysis revealed that they, too, are essential for planosporicin production in the native host. Reverse transcription-PCR (RT-PCR) analysis indicated that the planosporicin gene cluster is transcribed in three operons. Expression of one of these, pspEF, which encodes an ABC transporter, in Streptomyces coelicolor A3(2) conferred some degree of planosporicin resistance on the heterologous host. The inability to delete these genes from P. alba suggests that they play an essential role in immunity in the natural producer. PMID:23475977

  11. Cloning and analysis of the planosporicin lantibiotic biosynthetic gene cluster of Planomonospora alba.

    PubMed

    Sherwood, Emma J; Hesketh, Andrew R; Bibb, Mervyn J

    2013-05-01

    The increasing prevalence of antibiotic resistance in bacterial pathogens has renewed focus on natural products with antimicrobial properties. Lantibiotics are ribosomally synthesized peptide antibiotics that are posttranslationally modified to introduce (methyl)lanthionine bridges. Actinomycetes are renowned for their ability to produce a large variety of antibiotics, many with clinical applications, but are known to make only a few lantibiotics. One such compound is planosporicin produced by Planomonospora alba, which inhibits cell wall biosynthesis in Gram-positive pathogens. Planosporicin is a type AI lantibiotic structurally similar to those which bind lipid II, the immediate precursor for cell wall biosynthesis. The gene cluster responsible for planosporicin biosynthesis was identified by genome mining and subsequently isolated from a P. alba cosmid library. A minimal cluster of 15 genes sufficient for planosporicin production was defined by heterologous expression in Nonomuraea sp. strain ATCC 39727, while deletion of the gene encoding the precursor peptide from P. alba, which abolished planosporicin production, was also used to confirm the identity of the gene cluster. Deletion of genes encoding likely biosynthetic enzymes identified through bioinformatic analysis revealed that they, too, are essential for planosporicin production in the native host. Reverse transcription-PCR (RT-PCR) analysis indicated that the planosporicin gene cluster is transcribed in three operons. Expression of one of these, pspEF, which encodes an ABC transporter, in Streptomyces coelicolor A3(2) conferred some degree of planosporicin resistance on the heterologous host. The inability to delete these genes from P. alba suggests that they play an essential role in immunity in the natural producer.

  12. Identification and analysis of the paulomycin biosynthetic gene cluster and titer improvement of the paulomycins in Streptomyces paulus NRRL 8115.

    PubMed

    Li, Jine; Xie, Zhoujie; Wang, Min; Ai, Guomin; Chen, Yihua

    2015-01-01

    The paulomycins are a group of glycosylated compounds featuring a unique paulic acid moiety. To locate their biosynthetic gene clusters, the genomes of two paulomycin producers, Streptomyces paulus NRRL 8115 and Streptomyces sp. YN86, were sequenced. The paulomycin biosynthetic gene clusters were defined by comparative analyses of the two genomes together with the genome of the third paulomycin producer Streptomyces albus J1074. Subsequently, the identity of the paulomycin biosynthetic gene cluster was confirmed by inactivation of two genes involved in biosynthesis of the paulomycose branched chain (pau11) and the ring A moiety (pau18) in Streptomyces paulus NRRL 8115. After determining the gene cluster boundaries, a convergent biosynthetic model was proposed for paulomycin based on the deduced functions of the pau genes. Finally, a paulomycin high-producing strain was constructed by expressing an activator-encoding gene (pau13) in S. paulus, setting the stage for future investigations. PMID:25822496

  13. Web-type evolution of rhodococcus gene clusters associated with utilization of naphthalene.

    PubMed

    Kulakov, Leonid A; Chen, Shenchang; Allen, Christopher C R; Larkin, Michael J

    2005-04-01

    Clusters of genes which include determinants for the catalytic subunits of naphthalene dioxygenase (narAa and narAb) were analyzed in naphthalene-degrading Rhodococcus strains. We demonstrated (i) that in the region analyzed homologous gene clusters are separated from each other by nonhomologous DNA, (ii) that there are various degrees of homology between related genes, and (iii) that nar genes are located on plasmids in strains NCIMB12038 and P400 and on a chromosome in P200. These observations suggest that genetic exchange and reshuffling of genetic modules, as well as vertical descent of the genetic information, were the main routes in the evolution of naphthalene degradation in Rhodococcus. These conclusions were supported by studies of transcription patterns in the region analyzed. It was found that the nar region is not organized into a single operon but there are several transcription units which differ in the strains investigated. The narA and narB genes were found to be transcribed as a single unit in all strains analyzed, and their transcription was induced by naphthalene. The putative aldolase gene (narC) was found on the same transcript only in strains P200 and P400. In NCIMB12038 transcription of two more gene clusters was induced by growth on naphthalene. Transcription start sites for narA and narB were found to be different in all of the strains studied. Putative regulatory genes (narR1 and narR2) were transcribed as a single mRNA in naphthalene-induced cells. At the same time, a number of the genes known to be essential for naphthalene catabolism in gram-negative bacteria were not found in the region analyzed.

  14. Clustering of multiallele DNA markers near the Huntington's disease gene.

    PubMed

    MacDonald, M E; Cheng, S V; Zimmer, M; Haines, J L; Poustka, A; Allitto, B; Smith, B; Whaley, W L; Romano, D M; Jagadeesh, J

    1989-09-01

    Five highly informative multiallele restriction fragment length polymorphisms (RFLPs) of value for preclinical diagnosis of Huntington's disease (HD) have been genetically characterized. One RFLP was uncovered by expansion of the D4S43 locus while three others are at D4S111 and D4S115, loci defined by NotI-linking clones. The final marker, D4S125, represents a recently discovered VNTR locus. All four loci map closer to the HD gene and to the telomere than D4S10, the original linked marker for HD. In combination with two multiallele RFLPs previously identified for D4S43 and another linked locus, D4S95, these five new multiallele markers will dramatically improve the speed and accuracy of predictive testing in HD, and increase its applicability by maximizing the chances of an informative test for anyone with appropriate family structure.

  15. The Yersinia kristensenii O11 O-antigen gene cluster was acquired by lateral gene transfer and incorporated at a novel chromosomal locus.

    PubMed

    Cunneen, Monica M; Reeves, Peter R

    2007-06-01

    We have sequenced the O-antigen gene clusters for the Escherichia coli O98 and Yersinia kristensenii O11 O antigens. The basic structures of these O antigens are identical, and the sequence data indicate that Y. kristensenii O11 gained its O-antigen gene cluster by lateral gene transfer (LGT). Escherichia coli O98 has a typical O-antigen gene cluster between galF and gnd as is usual in E. coli. However, the O-antigen gene cluster of Y. kristensenii O11 is not located at the traditional Yersinia O-antigen gene cluster locus, between hemH and gsk, but at a novel chromosomal locus between aroA and cmk where it is flanked by remnant galF and gnd genes that indicate the probable source of the gene cluster. Phylogenetic analysis indicated that the source was not E. coli itself but a species in the Escherichia, Salmonella, and Klebsiella group of genera. Although other O-antigen studies imply LGT on the basis of the hypervariability of the loci and GC content, this report also identifies a potential donor and provides evidence for the mechanism involved. Remnant insertion sequence (IS) sequences flank the galF and gnd remnants and suggest that LGT of the gene cluster was IS mediated.

  16. Coexpression of Lactobacillus brevis ADH with GDH or G6PDH in Arxula adeninivorans for the synthesis of 1-(R)-phenylethanol.

    PubMed

    Rauter, Marion; Prokoph, Alexandra; Kasprzak, Jakub; Becker, Karin; Baronian, Keith; Bode, Rüdiger; Kunze, Gotthard; Vorbrodt, H- Matthias

    2015-06-01

    The yeast Arxula adeninivorans was used for the overexpression of an ADH gene of Lactobacillus brevis coding for (R)-specific alcohol dehydrogenase (LbADH) to synthesise enantiomerically pure 1-(R)-phenylethanol. Glucose dehydrogenase gene from Bacillus megaterium (BmGDH) or glucose 6-phosphate dehydrogenase of Bacillus pumilus (BpG6PDH) were coexpressed in Arxula to regenerate the cofactor NADPH by oxidising glucose or glucose 6-phosphate. The yeast strain expressing LbADH and BpG6PDH produced 5200 U l(-1) ADH and 370 U l(-1) G6PDH activity, whereas the strain expressing LbADH and BmGDH produced 2700 U l(-1) ADH and 170 U l(-1) GDH activity. However, the crude extract of both strains reduced 40 mM acetophenone to pure 1-(R)-phenylethanol with an enantiomeric excess (ee) of >99 % in 60 min without detectable by-products. An increase in yield was achieved using immobilised crude extracts (IEs), Triton X-100 permeabilised cells (PCs) and permeabilised immobilised cells (PICs) with PICs being most stable with GDH regeneration over 52 cycles. Even though the activity and synthesis rate of 1-(R)-phenylethanol with the BpG6PDH and LbADH coexpressing strain was higher, the BmGDH-LbADH strain was more stable over successive reaction cycles. This, combined with its higher total turnover number (TTN) of 391 mol product per mole NADP(+), makes it the preferred strain for continuous reaction systems. The initial non-optimised semi-continuous reaction produced 9.74 g l(-1) day(-1) or 406 g kg(-1) dry cell weight (dcw) day(-1) isolated 1-(R)-phenylethanol with an ee of 100 % and a TTN of 206 mol product per mole NADP(+). In conclusion, A. adeninivorans is a promising host for LbADH and BpG6PDH or BmGDH production and offers a simple method for the production of enantiomerically pure alcohols.

  17. Classification and Clustering on Microarray Data for Gene Functional Prediction Using R.

    PubMed

    López-Kleine, Liliana; Kleine, Liliana López; Montaño, Rosa; Torres-Avilés, Francisco

    2016-01-01

    Gene expression data (microarrays and RNA-sequencing data) as well as other kinds of genomic data can be extracted from publicly available genomic data. Here, we explain how to apply multivariate cluster and classification methods on gene expression data. These methods have become very popular and are implemented in freely available software in order to predict the participation of gene products in a specific functional category of interest. Taking into account the availability of data and of these methods, every biological study should apply them in order to obtain knowledge on the organism studied and functional category of interest. A special emphasis is made on the nonlinear kernel classification methods. PMID:25762300

  18. GIP2, a Putative Transcription Factor That Regulates the Aurofusarin Biosynthetic Gene Cluster in Gibberella zeae

    PubMed Central

    Kim, Jung-Eun; Jin, Jianming; Kim, Hun; Kim, Jin-Cheol; Yun, Sung-Hwan; Lee, Yin-Won

    2006-01-01

    Gibberella zeae (anamorph: Fusarium graminearum) is an important pathogen of maize, wheat, and rice. Colonies of G. zeae produce yellow-to-tan mycelia with the white-to-carmine red margins. In this study, we focused on nine putative open reading frames (ORFs) closely linked to PKS12 and GIP1, which are required for aurofusarin biosynthesis in G. zeae. Among them is an ORF designated GIP2 (for Gibberella zeae pigment gene 2), which encodes a putative protein of 398 amino acids that carries a Zn(II)2Cys6 binuclear cluster DNA-binding domain commonly found in transcription factors of yeasts and filamentous fungi. Targeted gene deletion and complementation analyses confirmed that GIP2 is required for aurofusarin biosynthesis. Expression of GIP2 in carrot medium correlated with aurofusarin production by G. zeae and was restricted to vegetative mycelia. Inactivation of the 10 contiguous genes in the ΔGIP2 strain delineates an aurofusarin biosynthetic gene cluster. Overexpression of GIP2 in both the ΔGIP2 and the wild-type strains increases aurofusarin production and reduces mycelial growth. Thus, GIP2 is a putative positive regulator of the aurofusarin biosynthetic gene cluster, and aurofusarin production is negatively correlated with vegetative growth by G. zeae. PMID:16461721

  19. Strategies to regulate transcription factor–mediated gene positioning and interchromosomal clustering at the nuclear periphery

    PubMed Central

    Randise-Hinchliff, Carlo; Coukos, Robert; Sood, Varun; Sumner, Michael Chas; Zdraljevic, Stefan; Meldi Sholl, Lauren; Garvey Brickner, Donna; Ahmed, Sara; Watchmaker, Lauren

    2016-01-01

    In budding yeast, targeting of active genes to the nuclear pore complex (NPC) and interchromosomal clustering is mediated by transcription factor (TF) binding sites in the gene promoters. For example, the binding sites for the TFs Put3, Ste12, and Gcn4 are necessary and sufficient to promote positioning at the nuclear periphery and interchromosomal clustering. However, in all three cases, gene positioning and interchromosomal clustering are regulated. Under uninducing conditions, local recruitment of the Rpd3(L) histone deacetylase by transcriptional repressors blocks Put3 DNA binding. This is a general function of yeast repressors: 16 of 21 repressors blocked Put3-mediated subnuclear positioning; 11 of these required Rpd3. In contrast, Ste12-mediated gene positioning is regulated independently of DNA binding by mitogen-activated protein kinase phosphorylation of the Dig2 inhibitor, and Gcn4-dependent targeting is up-regulated by increasing Gcn4 protein levels. These different regulatory strategies provide either qualitative switch-like control or quantitative control of gene positioning over different time scales. PMID:26953353

  20. Next-generation sequencing approach for connecting secondary metabolites to biosynthetic gene clusters in fungi

    PubMed Central

    Cacho, Ralph A.; Tang, Yi; Chooi, Yit-Heng

    2015-01-01

    Genomics has revolutionized the research on fungal secondary metabolite (SM) biosynthesis. To elucidate the molecular and enzymatic mechanisms underlying the biosynthesis of a specific SM compound, the important first step is often to find the genes that responsible for its synthesis. The accessibility to fungal genome sequences allows the bypass of the cumbersome traditional library construction and screening approach. The advance in next-generation sequencing (NGS) technologies have further improved the speed and reduced the cost of microbial genome sequencing in the past few years, which has accelerated the research in this field. Here, we will present an example work flow for identifying the gene cluster encoding the biosynthesis of SMs of interest using an NGS approach. We will also review the different strategies that can be employed to pinpoint the targeted gene clusters rapidly by giving several examples stemming from our work. PMID:25642215

  1. Comprehensive curation and analysis of fungal biosynthetic gene clusters of published natural products.

    PubMed

    Li, Yong Fuga; Tsai, Kathleen J S; Harvey, Colin J B; Li, James Jian; Ary, Beatrice E; Berlew, Erin E; Boehman, Brenna L; Findley, David M; Friant, Alexandra G; Gardner, Christopher A; Gould, Michael P; Ha, Jae H; Lilley, Brenna K; McKinstry, Emily L; Nawal, Saadia; Parry, Robert C; Rothchild, Kristina W; Silbert, Samantha D; Tentilucci, Michael D; Thurston, Alana M; Wai, Rebecca B; Yoon, Yongjin; Aiyar, Raeka S; Medema, Marnix H; Hillenmeyer, Maureen E; Charkoudian, Louise K

    2016-04-01

    Microorganisms produce a wide range of natural products (NPs) with clinically and agriculturally relevant biological activities. In bacteria and fungi, genes encoding successive steps in a biosynthetic pathway tend to be clustered on the chromosome as biosynthetic gene clusters (BGCs). Historically, "activity-guided" approaches to NP discovery have focused on bioactivity screening of NPs produced by culturable microbes. In contrast, recent "genome mining" approaches first identify candidate BGCs, express these biosynthetic genes using synthetic biology methods, and finally test for the production of NPs. Fungal genome mining efforts and the exploration of novel sequence and NP space are limited, however, by the lack of a comprehensive catalog of BGCs encoding experimentally-validated products. In this study, we generated a comprehensive reference set of fungal NPs whose biosynthetic gene clusters are described in the published literature. To generate this dataset, we first identified NCBI records that included both a peer-reviewed article and an associated nucleotide record. We filtered these records by text and homology criteria to identify putative NP-related articles and BGCs. Next, we manually curated the resulting articles, chemical structures, and protein sequences. The resulting catalog contains 197 unique NP compounds covering several major classes of fungal NPs, including polyketides, non-ribosomal peptides, terpenoids, and alkaloids. The distribution of articles published per compound shows a bias toward the study of certain popular compounds, such as the aflatoxins. Phylogenetic analysis of biosynthetic genes suggests that much chemical and enzymatic diversity remains to be discovered in fungi. Our catalog was incorporated into the recently launched Minimum Information about Biosynthetic Gene cluster (MIBiG) repository to create the largest known set of fungal BGCs and associated NPs, a resource that we anticipate will guide future genome mining and

  2. Identification, isolation, and analysis of a gene cluster involved in iron acquisition by Pseudomonas mendocina ymp

    PubMed Central

    Awaya, Jonathan D.

    2013-01-01

    Microbial acquisition of iron from natural sources in aerobic environments is a little-studied process that may lead to mineral instability and trace metal mobilization. Pseudomonas mendocina ymp was isolated from the Yucca Mountain Site for long-term nuclear waste storage. Its ability to solubilize a variety of Fe-containing minerals under aerobic conditions has been previously investigated but its molecular and genetic potential remained uncharacterized. Here, we have shown that the organism produces a hydroxamate and not a catecholate-based siderophore that is synthesized via non-ribosomal peptide synthetases. Gene clustering patterns observed in other Pseudomonads suggested that hybridizing multiple probes to the same library could allow for the identification of one or more clusters of syntenic siderophore-associated genes. Using this approach, two independent clusters were identified. An unfinished draft genome sequence of P. mendocina ymp indicated that these mapped to two independent contigs. The sequenced clusters were investigated informatically and shown to contain respectively a potentially complete set of genes responsible for siderophore biosynthesis, uptake, and regulation, and an incomplete set of genes with low individual homology to siderophore-associated genes. A mutation in the cluster’s pvdA homolog (pmhA) resulted in a siderophore-null phenotype, which could be reversed by complementation. The organism likely produces one siderophore with possibly different isoforms and a peptide backbone structure containing seven residues (predicted sequence: Acyl-Asp-Dab-Ser-fOHOrn-Ser-fOHorn). A similar approach could be applied for discovery of Fe− and siderophore-associated genes in unsequenced or poorly annotated organisms. PMID:18058194

  3. A Telomeric Cluster of Antimony Resistance Genes on Chromosome 34 of Leishmania infantum.

    PubMed

    Tejera Nevado, Paloma; Bifeld, Eugenia; Höhn, Katharina; Clos, Joachim

    2016-09-01

    The mechanisms underlying the drug resistance of Leishmania spp. are manifold and not completely identified. Apart from the highly conserved multidrug resistance gene family known from higher eukaryotes, Leishmania spp. also possess genus-specific resistance marker genes. One of them, ARM58, was first identified in Leishmania braziliensis using a functional cloning approach, and its domain structure was characterized in L. infantum Here we report that L. infantum ARM58 is part of a gene cluster at the telomeric end of chromosome 34 also comprising the neighboring genes ARM56 and HSP23. We show that overexpression of all three genes can confer antimony resistance to intracellular amastigotes. Upon overexpression in L. donovani, ARM58 and ARM56 are secreted via exosomes, suggesting a scavenger/secretion mechanism of action. Using a combination of functional cloning and next-generation sequencing, we found that the gene cluster was selected only under antimonyl tartrate challenge and weakly under Cu(2+) challenge but not under sodium arsenite, Cd(2+), or miltefosine challenge. The selective advantage is less pronounced in intracellular amastigotes treated with the sodium stibogluconate, possibly due to the known macrophage-stimulatory activity of this drug, against which these resistance markers may not be active. Our data point to the specificity of these three genes for antimony resistance. PMID:27324767

  4. Discovery of five conserved beta -defensin gene clusters using a computational search strategy.

    PubMed

    Schutte, Brian C; Mitros, Joseph P; Bartlett, Jennifer A; Walters, Jesse D; Jia, Hong Peng; Welsh, Michael J; Casavant, Thomas L; McCray, Paul B

    2002-02-19

    The innate immune system includes antimicrobial peptides that protect multicellular organisms from a diverse spectrum of microorganisms. beta-Defensins comprise one important family of mammalian antimicrobial peptides. The annotation of the human genome fails to reveal the expected diversity, and a recent query of the draft sequence with the blast search engine found only one new beta-defensin gene (DEFB3). To define better the beta-defensin gene family, we adopted a genomics approach that uses hmmer, a computational search tool based on hidden Markov models, in combination with blast. This strategy identified 28 new human and 43 new mouse beta-defensin genes in five syntenic chromosomal regions. Within each syntenic cluster, the gene sequences and organization were similar, suggesting each cluster pair arose from a common ancestor and was retained because of conserved functions. Preliminary analysis indicates that at least 26 of the predicted genes are transcribed. These results demonstrate the value of a genomewide search strategy to identify genes with conserved structural motifs. Discovery of these genes represents a new starting point for exploring the role of beta-defensins in innate immunity.

  5. Dynamic and physical clustering of gene expression during epidermal barrier formation in differentiating keratinocytes.

    PubMed

    Taylor, Jennifer M; Street, Teresa L; Hao, Lizhong; Copley, Richard; Taylor, Martin S; Hayden, Patrick J; Stolper, Gina; Mott, Richard; Hein, Jotun; Moffatt, Miriam F; Cookson, William O C M

    2009-01-01

    The mammalian epidermis is a continually renewing structure that provides the interface between the organism and an innately hostile environment. The keratinocyte is its principal cell. Keratinocyte proteins form a physical epithelial barrier, protect against microbial damage, and prepare immune responses to danger. Epithelial immunity is disordered in many common diseases and disordered epithelial differentiation underlies many cancers. In order to identify the genes that mediate epithelial development we used a tissue model of the skin derived from primary human keratinocytes. We measured global gene expression in triplicate at five times over the ten days that the keratinocytes took to fully differentiate. We identified 1282 gene transcripts that significantly changed during differentiation (false discovery rate <0.01%). We robustly grouped these transcripts by K-means clustering into modules with distinct temporal expression patterns, shared regulatory motifs, and biological functions. We found a striking cluster of late expressed genes that form the structural and innate immune defences of the epithelial barrier. Gene Ontology analyses showed that undifferentiated keratinocytes were characterised by genes for motility and the adaptive immune response. We systematically identified calcium-binding genes, which may operate with the epidermal calcium gradient to control keratinocyte division during skin repair. The results provide multiple novel insights into keratinocyte biology, in particular providing a comprehensive list of known and previously unrecognised major components of the epidermal barrier. The findings provide a reference for subsequent understanding of how the barrier functions in health and disease. PMID:19888454

  6. Genetic Characterization of the Klebsiella pneumoniae waa Gene Cluster, Involved in Core Lipopolysaccharide Biosynthesis

    PubMed Central

    Regué, Miguel; Climent, Núria; Abitiu, Nihal; Coderch, Núria; Merino, Susana; Izquierdo, Luis; Altarriba, Maria; Tomás, Juan M.

    2001-01-01

    A recombinant cosmid containing genes involved in Klebsiella pneumoniae C3 core lipopolysaccharide biosynthesis was identified by its ability to confer bacteriocin 28b resistance to Escherichia coli K-12. The recombinant cosmid contains 12 genes, the whole waa gene cluster, flanked by kbl and coaD genes, as was found in E. coli K-12. PCR amplification analysis showed that this cluster is conserved in representative K. pneumoniae strains. Partial nucleotide sequence determination showed that the same genes and gene order are found in K. pneumoniae subsp. ozaenae, for which the core chemical structure is known. Complementation analysis of known waa mutants from E. coli K-12 and/or Salmonella enterica led to the identification of genes involved in biosynthesis of the inner core backbone that are shared by these three members of the Enterobacteriaceae. K. pneumoniae orf10 mutants showed a two-log-fold reduction in a mice virulence assay and a strong decrease in capsule amount. Analysis of a constructed K. pneumoniae waaE deletion mutant suggests that the WaaE protein is involved in the transfer of the branch β-d-Glc to the O-4 position of l-glycero-d-manno-heptose I, a feature shared by K. pneumoniae, Proteus mirabilis, and Yersinia enterocolitica. PMID:11371519

  7. Sequencing and Analysis of the Biosynthetic Gene Cluster of the Lipopeptide Antibiotic Friulimicin in Actinoplanes friuliensis▿

    PubMed Central

    Müller, C.; Nolden, S.; Gebhardt, P.; Heinzelmann, E.; Lange, C.; Puk, O.; Welzel, K.; Wohlleben, W.; Schwartz, D.

    2007-01-01

    Actinoplanes friuliensis produces the lipopeptide antibiotic friulimicin, which is a cyclic peptide with one exocyclic amino acid linked to a branched-chain fatty acid acyl residue. The structural relationship to daptomycin and the excellent antibacterial performance of friulimicin make the antibiotic an attractive drug candidate. The complete friulimicin biosynthetic gene cluster of 24 open reading frames from A. friuliensis was sequenced and analyzed. In addition to genes for regulation, self-resistance, and transport, the cluster contains genes encoding peptide synthetases, proteins involved in the synthesis and linkage of the fatty acid component of the antibiotic, and proteins involved in the synthesis of the nonproteinogenic amino acids pipecolinic acid, methylaspartic acid, and 2,3-diaminobutyric acid. By using heterologous gene expression in Escherichia coli, we provide biochemical evidence for the stereoselective synthesis of l-pipecolinic acid by the deduced protein of the lysine cyclodeaminase gene pip. Furthermore, we show the involvement of the dabA and dabB genes in the biosynthesis of 2,3-diaminobutyric acid by gene inactivation and subsequent feeding experiments. PMID:17220414

  8. Visualizing the HoxD Gene Cluster at the Nanoscale Level.

    PubMed

    Fabre, Pierre J; Benke, Alexander; Manley, Suliana; Duboule, Denis

    2015-01-01

    Transcription of HoxD cluster genes in limbs is coordinated by two topologically associating domains (TADs), neighboring the cluster and containing various enhancers. Here, we use a combination of microscopy approaches and chromosome conformation capture to assess the structural changes occurring in this global architecture in various functional states. We observed that despite their spatial juxtaposition, the TADs are consistently kept as distinct three-dimensional units. Hox