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1

Essential Bacillus subtilis genes  

Microsoft Academic Search

To estimate the minimal gene set required to sustain bacterial life in nutritious conditions, we carried out a systematic inactivation of Bacillus subtilis genes. Among 4,100 genes of the organism, only 192 were shown to be indispensable by this or previous work. Another 79 genes were predicted to be essential. The vast majority of essential genes were categorized in relatively

K. Kobayashi; S. D. Ehrlichb; A. Albertini; G. Amati; K. Asaig Arnaudf; M. Arnaud; K. Asai; S. Ashikaga; S. Aymerich; P. Bessieres; F. Boland; S. C. Brignell; S. Bron; K. Bunai; J. Chapuis; L. C. Christiansen; A. Danchin; M. Débarbouillé; E. Dervyn; E. Deuerling; K. Devine; S. K. Devine; O. Dreesen; J. Errington; S. Fillinger; S. J. Foster; Y. Fujita; A. Galizzi; R. Gardan; C. Eschevins; T. Fukushima; K. Haga; C. R. Harwood; M. Hecker; D. Hosoya; M. F. Hullo; H. Kakeshita; D. Karamata; Y. Kasahara; F. Kawamura; K. Koga; P. Koski; R. Kuwana; D. Imamura; M. Ishimaru; S. Ishikawa; I. Ishio; D. Le Coq; A. Masson; C. Mauël; R. Meima; R. P. Mellado; A. Moir; S. Moriya; E. Nagakawa; H. Nanamiya; S. Nakai; P. Nygaard; M. Ogura; T. Ohanan; M. O'Reilly; M. O'Rourke; Z. Pragai; H. M. Pooley; G. Rapoport; J. P. Rawlins; L. A. Rivas; C. Rivolta; A. Sadaie; Y. Sadaie; M. Sarvas; T. Sato; H. H. Saxild; E. Scanlan; W. Schumann; J. F. Seegers; J. Sekiguchi; A. Sekowska; S. J. Seror; M. Simon; P. Stragier; R. Studer; H. Takamatsu; T. Tanaka; M. Takeuchi; H. B. Thomaides; V. Vagner; J. M. van Dijl; K. Watabe; A. Wipat; H. Yamamoto; M. Yamamoto; Y. Yamamoto; K. Yamane; K. Yata; K. Yoshida; H. Yoshikawa; U. Zuber; N. Ogasawara

2003-01-01

2

Essential Bacillus subtilis genes.  

PubMed

To estimate the minimal gene set required to sustain bacterial life in nutritious conditions, we carried out a systematic inactivation of Bacillus subtilis genes. Among approximately 4,100 genes of the organism, only 192 were shown to be indispensable by this or previous work. Another 79 genes were predicted to be essential. The vast majority of essential genes were categorized in relatively few domains of cell metabolism, with about half involved in information processing, one-fifth involved in the synthesis of cell envelope and the determination of cell shape and division, and one-tenth related to cell energetics. Only 4% of essential genes encode unknown functions. Most essential genes are present throughout a wide range of Bacteria, and almost 70% can also be found in Archaea and Eucarya. However, essential genes related to cell envelope, shape, division, and respiration tend to be lost from bacteria with small genomes. Unexpectedly, most genes involved in the Embden-Meyerhof-Parnas pathway are essential. Identification of unknown and unexpected essential genes opens research avenues to better understanding of processes that sustain bacterial life. PMID:12682299

Kobayashi, K; Ehrlich, S D; Albertini, A; Amati, G; Andersen, K K; Arnaud, M; Asai, K; Ashikaga, S; Aymerich, S; Bessieres, P; Boland, F; Brignell, S C; Bron, S; Bunai, K; Chapuis, J; Christiansen, L C; Danchin, A; Débarbouille, M; Dervyn, E; Deuerling, E; Devine, K; Devine, S K; Dreesen, O; Errington, J; Fillinger, S; Foster, S J; Fujita, Y; Galizzi, A; Gardan, R; Eschevins, C; Fukushima, T; Haga, K; Harwood, C R; Hecker, M; Hosoya, D; Hullo, M F; Kakeshita, H; Karamata, D; Kasahara, Y; Kawamura, F; Koga, K; Koski, P; Kuwana, R; Imamura, D; Ishimaru, M; Ishikawa, S; Ishio, I; Le Coq, D; Masson, A; Mauël, C; Meima, R; Mellado, R P; Moir, A; Moriya, S; Nagakawa, E; Nanamiya, H; Nakai, S; Nygaard, P; Ogura, M; Ohanan, T; O'Reilly, M; O'Rourke, M; Pragai, Z; Pooley, H M; Rapoport, G; Rawlins, J P; Rivas, L A; Rivolta, C; Sadaie, A; Sadaie, Y; Sarvas, M; Sato, T; Saxild, H H; Scanlan, E; Schumann, W; Seegers, J F M L; Sekiguchi, J; Sekowska, A; Séror, S J; Simon, M; Stragier, P; Studer, R; Takamatsu, H; Tanaka, T; Takeuchi, M; Thomaides, H B; Vagner, V; van Dijl, J M; Watabe, K; Wipat, A; Yamamoto, H; Yamamoto, M; Yamamoto, Y; Yamane, K; Yata, K; Yoshida, K; Yoshikawa, H; Zuber, U; Ogasawara, N

2003-04-07

3

Essential Bacillus subtilis genes  

PubMed Central

To estimate the minimal gene set required to sustain bacterial life in nutritious conditions, we carried out a systematic inactivation of Bacillus subtilis genes. Among ?4,100 genes of the organism, only 192 were shown to be indispensable by this or previous work. Another 79 genes were predicted to be essential. The vast majority of essential genes were categorized in relatively few domains of cell metabolism, with about half involved in information processing, one-fifth involved in the synthesis of cell envelope and the determination of cell shape and division, and one-tenth related to cell energetics. Only 4% of essential genes encode unknown functions. Most essential genes are present throughout a wide range of Bacteria, and almost 70% can also be found in Archaea and Eucarya. However, essential genes related to cell envelope, shape, division, and respiration tend to be lost from bacteria with small genomes. Unexpectedly, most genes involved in the Embden–Meyerhof–Parnas pathway are essential. Identification of unknown and unexpected essential genes opens research avenues to better understanding of processes that sustain bacterial life.

Kobayashi, K.; Ehrlich, S. D.; Albertini, A.; Amati, G.; Andersen, K. K.; Arnaud, M.; Asai, K.; Ashikaga, S.; Aymerich, S.; Bessieres, P.; Boland, F.; Brignell, S. C.; Bron, S.; Bunai, K.; Chapuis, J.; Christiansen, L. C.; Danchin, A.; Debarbouille, M.; Dervyn, E.; Deuerling, E.; Devine, K.; Devine, S. K.; Dreesen, O.; Errington, J.; Fillinger, S.; Foster, S. J.; Fujita, Y.; Galizzi, A.; Gardan, R.; Eschevins, C.; Fukushima, T.; Haga, K.; Harwood, C. R.; Hecker, M.; Hosoya, D.; Hullo, M. F.; Kakeshita, H.; Karamata, D.; Kasahara, Y.; Kawamura, F.; Koga, K.; Koski, P.; Kuwana, R.; Imamura, D.; Ishimaru, M.; Ishikawa, S.; Ishio, I.; Le Coq, D.; Masson, A.; Mauel, C.; Meima, R.; Mellado, R. P.; Moir, A.; Moriya, S.; Nagakawa, E.; Nanamiya, H.; Nakai, S.; Nygaard, P.; Ogura, M.; Ohanan, T.; O'Reilly, M.; O'Rourke, M.; Pragai, Z.; Pooley, H. M.; Rapoport, G.; Rawlins, J. P.; Rivas, L. A.; Rivolta, C.; Sadaie, A.; Sadaie, Y.; Sarvas, M.; Sato, T.; Saxild, H. H.; Scanlan, E.; Schumann, W.; Seegers, J. F. M. L.; Sekiguchi, J.; Sekowska, A.; Seror, S. J.; Simon, M.; Stragier, P.; Studer, R.; Takamatsu, H.; Tanaka, T.; Takeuchi, M.; Thomaides, H. B.; Vagner, V.; van Dijl, J. M.; Watabe, K.; Wipat, A.; Yamamoto, H.; Yamamoto, M.; Yamamoto, Y.; Yamane, K.; Yata, K.; Yoshida, K.; Yoshikawa, H.; Zuber, U.; Ogasawara, N.

2003-01-01

4

The Bysl Gene Product, Bystin, is Essential for Survival of Mouse Embryos.  

PubMed Central

Human bystin is a cytoplasmic protein directly binding to trophinin, a cell adhesion molecule potentially involved in human embryo implantation. The present study shows that bystin is expressed in luminal and glandular epithelia in the mouse uterus at peri-implantation stages. In fertilized embryos, bystin was not seen until blastocyst stage. Bystin expression started during hatching and increased in expanded blastocyst. However, bystin disappeared from the blastocyst during implantation. After implantation bystin re-appeared in the epiblast. Targeted disruption of the mouse bystin gene, Bysl, resulted in embryonic lethality shortly after implantation, indicating that bystin is essential for survival of mouse embryos.

Aoki, Rui; Suzuki, Nao; Paria, Bibhash C.; Sugihara, Kazuhiro; Akama, Tomoya O.; Raab, Gerhard; Miyoshi, Masaya; Nadano, Daita; Fukuda, Michiko N.

2006-01-01

5

Lichenicidin Biosynthesis in Escherichia coli: licFGEHI Immunity Genes Are Not Essential for Lantibiotic Production or Self-Protection ?  

PubMed Central

This study demonstrated, for the first time, that immunity genes licFGEHI are not essential for self-protection and production of the two-component lantibiotic lichenicidin in the Gram-negative heterologous host Escherichia coli BLic5. Additionally, it was experimentally demonstrated that lichenicidin lantibiotics are active against the E. coli imp4213 strain, a mutant strain possessing a permeable outer membrane.

Caetano, Tania; Krawczyk, Joanna M.; Mosker, Eva; Sussmuth, Roderich D.; Mendo, Sonia

2011-01-01

6

Nonribosomal Peptide Synthetase Genes pesL and pes1 Are Essential for Fumigaclavine C Production in Aspergillus fumigatus  

PubMed Central

The identity of metabolites encoded by the majority of nonribosomal peptide synthetases in the opportunistic pathogen, Aspergillus fumigatus, remains outstanding. We found that the nonribosomal peptide (NRP) synthetases PesL and Pes1 were essential for fumigaclavine C biosynthesis, the end product of the complex ergot alkaloid (EA) pathway in A. fumigatus. Deletion of either pesL (?pesL) or pes1 (?pes1) resulted in complete loss of fumigaclavine C biosynthesis, relatively increased production of fumitremorgins such as TR-2, fumitremorgin C and verruculogen, increased sensitivity to H2O2, and increased sensitivity to the antifungals, voriconazole, and amphotericin B. Deletion of pesL resulted in severely reduced virulence in an invertebrate infection model (P < 0.001). These findings indicate that NRP synthesis plays an essential role in mediating the final prenylation step of the EA pathway, despite the apparent absence of NRP synthetases in the proposed EA biosynthetic cluster for A. fumigatus. Liquid chromatography/diode array detection/mass spectrometry analysis also revealed the presence of fumiquinazolines A to F in both A. fumigatus wild-type and ?pesL strains. This observation suggests that alternative NRP synthetases can also function in fumiquinazoline biosynthesis, since PesL has been shown to mediate fumiquinazoline biosynthesis in vitro. Furthermore, we provide here the first direct link between EA biosynthesis and virulence, in agreement with the observed toxicity associated with EA exposure. Finally, we demonstrate a possible cluster cross-talk phenomenon, a theme which is beginning to emerge in the literature.

O'Hanlon, Karen A.; Gallagher, Lorna; Schrettl, Markus; Jochl, Christoph; Kavanagh, Kevin; Larsen, Thomas O.

2012-01-01

7

Regulation of essential oil production in plants  

Microsoft Academic Search

This review provides a summary of the physiological dynamics andregulation of essential oil production, from the literature and availableinformation on diverse volatile oil crops. Essential oil production is highlyintegrated with the physiology of the whole plant and so depends on themetabolic state and preset developmental differentiation programme of thesynthesising tissue. Essential oil productivity is ecophysiologically andenvironmentally friendly. These and other

N. S. Sangwan; A. H. A. Farooqi; F. Shabih; R. S. Sangwan

2001-01-01

8

The yeast NOP4 gene product is an essential nucleolar protein required for pre-rRNA processing and accumulation of 60S ribosomal subunits.  

PubMed Central

The Saccharomyces cerevisiae NOP4 gene was isolated by screening a lambda gt11 yeast genomic DNA library with a monoclonal antibody against a yeast nucleolar protein. NOP4 encodes a 78 kDa protein that contains two prototypical RNA recognition motifs (RRMs) flanking an imperfect RRM lacking characteristic RNP1 and RNP2 motifs. In addition, there is a fourth incomplete RRM. NOP4 is a single copy essential gene present on chromosome XVI, between RAD1 and PEP4. To examine the function of Nop4p, we constructed a conditional null allele of NOP4 by placing this gene under the control of the glucose-repressible GAL1 promoter. When cells are shifted from galactose-containing medium to glucose-containing medium, NOP4 transcription is terminated, Nop4 protein is depleted and cell growth is impaired. Nop4 protein depletion results in diminished accumulation of 60S ribosomal subunits, assignable to a defect in ribosome biogenesis arising from a lack of production of mature 25S rRNA from 27S precursor rRNA. Images

Sun, C; Woolford, J L

1994-01-01

9

Transcriptional control and essential roles of the Escherichia coli ccm gene products in formate-dependent nitrite reduction and cytochrome c synthesis.  

PubMed Central

The eight ccm genes located at minute 47 on the Escherichia coli chromosome, in the order ccmABCDEFGH, encode homologues of proteins which are essential for cytochrome c assembly in other bacteria. The ccm genes are immediately downstream from the napFDAGHBC genes encoding a periplasmic nitrate reductase. CcmH was previously shown to be essential for cytochrome c assembly. Deletion analysis and a two-plasmid strategy have now been used to demonstrate that CcmA, B, D, E, F and G are also essential for cytochrome c assembly, and hence for cytochrome-c-dependent nitrite reduction. The ccm genes are transcribed from a ccmA promoter located within the adjacent gene, napC, which is the structural gene for a 24 kDa membrane-bound c-type cytochrome, NapC. Transcription from this ccmA promoter is induced approximately 5-fold during anaerobic growth, independently of a functional Fnr protein: it is also not regulated by the ArcB-ArcA two-component regulatory system. The ccmA promoter is an example of the 'extended -10 sequence' group of promoters with a TGX motif immediately upstream of the -10 sequence. Mutagenesis of the TG motif to TC, CT or CC resulted in loss of about 50% of the promoter activity. A weak second promoter is suggested to permit transcription of the downstream ccmEFGH genes in the absence of transcription readthrough from the upstream napF and ccmA promoters. The results are consistent with, but do not prove, the current view that CcmA, B, C and D are part of an essential haem transport mechanism, that CcmE, F and H are required for covalent haem attachment to cysteine-histidine motifs in cytochrome c apoproteins in the periplasm, and that CcmG is required for the reduction of cysteine residues on apocytochromes c in preparation for haem ligation.

Tanapongpipat, S; Reid, E; Cole, J A; Crooke, H

1998-01-01

10

Network rewiring is an important mechanism of gene essentiality change  

PubMed Central

Gene essentiality changes are crucial for organismal evolution. However, it is unclear how essentiality of orthologs varies across species. We investigated the underlying mechanism of gene essentiality changes between yeast and mouse based on the framework of network evolution and comparative genomic analysis. We found that yeast nonessential genes become essential in mouse when their network connections rapidly increase through engagement in protein complexes. The increased interactions allowed the previously nonessential genes to become members of vital pathways. By accounting for changes in gene essentiality, we firmly reestablished the centrality-lethality rule, which proposed the relationship of essential genes and network hubs. Furthermore, we discovered that the number of connections associated with essential and non-essential genes depends on whether they were essential in ancestral species. Our study describes for the first time how network evolution occurs to change gene essentiality.

Kim, Jinho; Kim, Inhae; Han, Seong Kyu; Bowie, James U.; Kim, Sanguk

2012-01-01

11

How to identify essential genes from molecular networks?  

PubMed Central

Background The prediction of essential genes from molecular networks is a way to test the understanding of essentiality in the context of what is known about the network. However, the current knowledge on molecular network structures is incomplete yet, and consequently the strategies aimed to predict essential genes are prone to uncertain predictions. We propose that simultaneously evaluating different network structures and different algorithms representing gene essentiality (centrality measures) may identify essential genes in networks in a reliable fashion. Results By simultaneously analyzing 16 different centrality measures on 18 different reconstructed metabolic networks for Saccharomyces cerevisiae, we show that no single centrality measure identifies essential genes from these networks in a statistically significant way; however, the combination of at least 2 centrality measures achieves a reliable prediction of most but not all of the essential genes. No improvement is achieved in the prediction of essential genes when 3 or 4 centrality measures were combined. Conclusion The method reported here describes a reliable procedure to predict essential genes from molecular networks. Our results show that essential genes may be predicted only by combining centrality measures, revealing the complex nature of the function of essential genes.

del Rio, Gabriel; Koschutzki, Dirk; Coello, Gerardo

2009-01-01

12

How to identify essential genes from molecular networks?  

Microsoft Academic Search

BACKGROUND: The prediction of essential genes from molecular networks is a way to test the understanding of essentiality in the context of what is known about the network. However, the current knowledge on molecular network structures is incomplete yet, and consequently the strategies aimed to predict essential genes are prone to uncertain predictions. We propose that simultaneously evaluating different network

Gabriel del Rio; Dirk Koschützki; Gerardo Coello

2009-01-01

13

Defining the Role of Essential Genes in Human Disease  

PubMed Central

A greater understanding of the causes of human disease can come from identifying characteristics that are specific to disease genes. However, a full understanding of the contribution of essential genes to human disease is lacking, due to the premise that these genes tend to cause developmental abnormalities rather than adult disease. We tested the hypothesis that human orthologs of mouse essential genes are associated with a variety of human diseases, rather than only those related to miscarriage and birth defects. We segregated human disease genes according to whether the knockout phenotype of their mouse ortholog was lethal or viable, defining those with orthologs producing lethal knockouts as essential disease genes. We show that the human orthologs of mouse essential genes are associated with a wide spectrum of diseases affecting diverse physiological systems. Notably, human disease genes with essential mouse orthologs are over-represented among disease genes associated with cancer, suggesting links between adult cellular abnormalities and developmental functions. The proteins encoded by essential genes are highly connected in protein-protein interaction networks, which we find correlates with an over-representation of nuclear proteins amongst essential disease genes. Disease genes associated with essential orthologs also are more likely than those with non-essential orthologs to contribute to disease through an autosomal dominant inheritance pattern, suggesting that these diseases may actually result from semi-dominant mutant alleles. Overall, we have described attributes found in disease genes according to the essentiality status of their mouse orthologs. These findings demonstrate that disease genes do occupy highly connected positions in protein-protein interaction networks, and that due to the complexity of disease-associated alleles, essential genes cannot be ignored as candidates for causing diverse human diseases.

Robertson, David L.; Hentges, Kathryn E.

2011-01-01

14

An overlapping essential gene in the Potyviridae  

PubMed Central

The family Potyviridae includes >30% of known plant virus species, many of which are of great agricultural significance. These viruses have a positive sense RNA genome that is ?10 kb long and contains a single long ORF. The ORF is translated into a large polyprotein, which is cleaved into ?10 mature proteins. We report the discovery of a short ORF embedded within the P3 cistron of the polyprotein but translated in the +2 reading-frame. The ORF, termed pipo, is conserved and has a strong bioinformatic coding signature throughout the large and diverse Potyviridae family. Mutations that knock out expression of the PIPO protein in Turnip mosaic potyvirus but leave the polyprotein amino acid sequence unaltered are lethal to the virus. Immunoblotting with antisera raised against two nonoverlapping 14-aa antigens, derived from the PIPO amino acid sequence, reveals the expression of an ?25-kDa PIPO fusion product in planta. This is consistent with expression of PIPO as a P3-PIPO fusion product via ribosomal frameshifting or transcriptional slippage at a highly conserved G1-2A6-7 motif at the 5? end of pipo. This discovery suggests that other short overlapping genes may remain hidden even in well studied virus genomes (as well as cellular organisms) and demonstrates the utility of the software package MLOGD as a tool for identifying such genes.

Chung, Betty Y.-W.; Miller, W. Allen; Atkins, John F.; Firth, Andrew E.

2008-01-01

15

Genetics of germination-arrest factor (GAF) production by Pseudomonas fluorescens WH6: identification of a gene cluster essential for GAF biosynthesis.  

PubMed

The genetic basis of the biosynthesis of the germination-arrest factor (GAF) produced by Pseudomonas fluorescens WH6, and previously identified as 4-formylaminooxyvinylglycine, has been investigated here. In addition to inhibiting the germination of a wide range of grassy weeds, GAF exhibits a selective antimicrobial activity against the bacterial plant pathogen Erwinia amylovora. We utilized the in vitro response of E. amylovora to GAF as a rapid screen for loss-of-function GAF phenotypes generated by transposon mutagenesis. A Tn5 mutant library consisting of 6364 WH6 transformants was screened in this Erwinia assay, resulting in the identification of 18 non-redundant transposon insertion sites that led to loss of GAF production in WH6, as confirmed by TLC analysis. These insertions mapped to five different genes and four intergenic regions. Three of these genes, including two putative regulatory genes (gntR and iopB homologues), were clustered in a 13 kb chromosomal region containing 13 putative ORFs. A GAF mutation identified previously as affecting an aminotransferase also maps to this region. We suggest that three of the genes in this region (a carbamoyltransferase, an aminotransferase and a formyltransferase) encode the enzymes necessary to synthesize dihydroGAF, the putative immediate precursor of GAF in a proposed GAF biosynthetic pathway. RT-qPCR analyses demonstrated that mutations in the gntR and iopB regulatory genes, as well as in a prtR homologue identified earlier as controlling GAF formation, suppressed transcription of at least two of the putative GAF biosynthetic genes (encoding the aminotransferase and formyltransferase) located in this 13 kb region. PMID:23125119

Halgren, Anne; Maselko, Maciej; Azevedo, Mark; Mills, Dallice; Armstrong, Donald; Banowetz, Gary

2012-11-01

16

A Penicillium expansum glucose oxidase-encoding gene, GOX2, is essential for gluconic acid production and acidification during colonization of deciduous fruit.  

PubMed

Penicillium expansum, the causal agent of blue mold rot, causes severe postharvest maceration of fruit through secretion of total, d-gluconic acid (GLA). Two P. expansum glucose oxidase (GOX)-encoding genes, GOX1 and GOX2, were analyzed. GOX activity and GLA accumulation were strongly related to GOX2 expression, which increased with pH to a maximum at pH 7.0, whereas GOX1 was expressed at pH 4.0, where no GOX activity or extracellular GLA were detected. This differential expression was also observed at the leading edge of the decaying tissue, where GOX2 expression was dominant. The roles of the GOX genes in pathogenicity were further studied through i) development of P. expansum goxRNAi mutants exhibiting differential downregulation of GOX2, ii) heterologous expression of the P. expansum GOX2 gene in the nondeciduous fruit-pathogen P. chrysogenum, and iii) modulation of GLA production by FeSO(4) chelation. Interestingly, in P. expansum, pH and GLA production elicited opposite effects on germination and biomass accumulation: 26% of spores germinated at pH 7.0 when GOX activity and GLA were highest whereas, in P. chrysogenum at the same pH, when GLA did not accumulate, 72% of spores germinated. Moreover, heterologous expression of P. expansum GOX2 in P. chrysogenum resulted in enhanced GLA production and reduced germination, suggesting negative regulation of spore germination and GLA production. These results demonstrate that pH modulation, mediated by GLA accumulation, is an important factor in generating the initial signal or signals for fungal development leading to host-tissue colonization by P. expansum. PMID:22352719

Barad, Shiri; Horowitz, Sigal Brown; Moscovitz, Oren; Lichter, Amnon; Sherman, Amir; Prusky, Dov

2012-06-01

17

High Confidence Prediction of Essential Genes in Burkholderia Cenocepacia  

PubMed Central

Background Essential genes are absolutely required for the survival of an organism. The identification of essential genes, besides being one of the most fundamental questions in biology, is also of interest for the emerging science of synthetic biology and for the development of novel antimicrobials. New antimicrobial therapies are desperately needed to treat multidrug-resistant pathogens, such as members of the Burkholderia cepacia complex. Methodology/Principal Findings We hypothesize that essential genes may be highly conserved within a group of evolutionary closely related organisms. Using a bioinformatics approach we determined that the core genome of the order Burkholderiales consists of 649 genes. All but two of these identified genes were located on chromosome 1 of Burkholderia cenocepacia. Although many of the 649 core genes of Burkholderiales have been shown to be essential in other bacteria, we were also able to identify a number of novel essential genes present mainly, or exclusively, within this order. The essentiality of some of the core genes, including the known essential genes infB, gyrB, ubiB, and valS, as well as the so far uncharacterized genes BCAL1882, BCAL2769, BCAL3142 and BCAL3369 has been confirmed experimentally in B. cenocepacia. Conclusions/Significance We report on the identification of essential genes using a novel bioinformatics strategy and provide bioinformatics and experimental evidence that the large majority of the identified genes are indeed essential. The essential genes identified here may represent valuable targets for the development of novel antimicrobials and their detailed study may shed new light on the functions required to support life.

Juhas, Mario; Stark, Manuel; von Mering, Christian; Lumjiaktase, Puthapoom; Crook, Derrick W.; Valvano, Miguel A.; Eberl, Leo

2012-01-01

18

Mutualistic polydnaviruses share essential replication gene functions with pathogenic ancestors.  

PubMed

Viruses are usually thought to form parasitic associations with hosts, but all members of the family Polydnaviridae are obligate mutualists of insects called parasitoid wasps. Phylogenetic data founded on sequence comparisons of viral genes indicate that polydnaviruses in the genus Bracovirus (BV) are closely related to pathogenic nudiviruses and baculoviruses. However, pronounced differences in the biology of BVs and baculoviruses together with high divergence of many shared genes make it unclear whether BV homologs still retain baculovirus-like functions. Here we report that virions from Microplitis demolitor bracovirus (MdBV) contain multiple baculovirus-like and nudivirus-like conserved gene products. We further show that RNA interference effectively and specifically knocks down MdBV gene expression. Coupling RNAi knockdown methods with functional assays, we examined the activity of six genes in the MdBV conserved gene set that are known to have essential roles in transcription (lef-4, lef-9), capsid assembly (vp39, vlf-1), and envelope formation (p74, pif-1) during baculovirus replication. Our results indicated that MdBV produces a baculovirus-like RNA polymerase that transcribes virus structural genes. Our results also supported a conserved role for vp39, vlf-1, p74, and pif-1 as structural components of MdBV virions. Additional experiments suggested that vlf-1 together with the nudivirus-like gene int-1 also have novel functions in regulating excision of MdBV proviral DNAs for packaging into virions. Overall, these data provide the first experimental insights into the function of BV genes in virion formation. PMID:23671417

Burke, Gaelen R; Thomas, Sarah A; Eum, Jai H; Strand, Michael R

2013-05-09

19

Conditional Lethal Amber Mutations in Essential Escherichia coli Genes  

Microsoft Academic Search

The essential genes of microorganisms encode biological functions important for survival and thus tend to be of high scientific interest. Drugs that interfere with essential functions are likely to be interesting candidates for antimicrobials. However, these genes are hard to study genetically because knockout mutations in them are by definition inviable. We recently described a conditional mutation system in Escherichia

Christopher D. Herring; Frederick R. Blattner

2004-01-01

20

Essential genes in proximal 3L heterochromatin of Drosophila melanogaster  

Microsoft Academic Search

We have further characterized essential loci within the centric heterochromatin of the left arm of chromosome 3 (3L) of Drosophila melanogaster, using EMS, radiation and P element mutagenesis. We failed to find any new essential genes, a result that suggests a lower-than-average gene density in this region. Mutations affecting expression of the most proximal gene [lethal 1, l1 or l(3)80Fj

S. Schulze; D. A. R. Sinclair; E. Silva; K. A. Fitzpatrick; M. Singh; V. K. Lloyd; K. A. Morin; J. Kim; D. G. Holm; J. A. Kennison; B. M. Honda

2001-01-01

21

In Vivo-Validated Essential Genes Identified in Acinetobacter baumannii by Using Human Ascites Overlap Poorly with Essential Genes Detected on Laboratory Media  

PubMed Central

ABSTRACT A critical feature of a potential antimicrobial target is the characteristic of being essential for growth and survival during host infection. For bacteria, genome-wide essentiality screens are usually performed on rich laboratory media. This study addressed whether genes detected in that manner were optimal for the identification of antimicrobial targets since the in vivo milieu is fundamentally different. Mutant derivatives of a clinical isolate of Acinetobacter baumannii were screened for growth on human ascites, an ex vivo medium that reflects the infection environment. A subset of 34 mutants with unique gene disruptions that demonstrated little to no growth on ascites underwent evaluation in a rat subcutaneous abscess model, establishing 18 (53%) of these genes as in vivo essential. The putative gene products all had annotated biological functions, represented unrecognized or underexploited antimicrobial targets, and could be grouped into five functional categories: metabolic, two-component signaling systems, DNA/RNA synthesis and regulation, protein transport, and structural. These A. baumannii in vivo essential genes overlapped poorly with the sets of essential genes from other Gram-negative bacteria catalogued in the Database of Essential Genes (DEG), including those of Acinetobacter baylyi, a closely related species. However, this finding was not due to the absence of orthologs. None of the 18 in vivo essential genes identified in this study, or their putative gene products, were targets of FDA-approved drugs or drugs in the developmental pipeline, indicating that a significant portion of the available target space within pathogenic Gram-negative bacteria is currently neglected.

Umland, Timothy C.; Schultz, L. Wayne; MacDonald, Ulrike; Beanan, Janet M.; Olson, Ruth; Russo, Thomas A.

2012-01-01

22

The AcMNPV pp31 gene is not essential for productive AcMNPV replication or late gene transcription but appears to increase levels of most viral transcripts  

PubMed Central

The pp31 gene of Autographa californica Multicapsid Nucleopolyhedrovirus (AcMNPV) encodes a phosphorylated DNA binding protein that associates with virogenic stroma in the nuclei of infected cells. Prior studies of pp31 by transient late expression assays suggested that pp31 may play an important role in transcription of AcMNPV late genes (Todd et al., 1995. J. Virol. 69, 968–974) although genetic studies of the closely related BmNPV pp31 gene suggested that pp31 may be dispensable (Gomi et al., 1997. Virology 230, 35–47). In the current study, we examined the role of the pp31 gene in the context of the AcMNPV genome during infection. We used a BACmid-based system to generate a pp31 knockout in the AcMNPV genome. The pp31 knockout was subsequently rescued by reinserting the pp31 gene into the polyhedrin locus of the same virus genome. We found that pp31 was not essential for viral replication although the absence of pp31 resulted in a lower viral titer. Analysis of viral DNA replication in the absence of pp31 showed that the kinetics of viral DNA replication were unaffected. An AcMNPV oligonucleotide microarray was used to compare gene expression from all AcMNPV genes in the presence or absence of pp31. In the absence of pp31 a modest reduction in transcripts was detected for many viral genes (99 genes) while no substantial increase or decrease was observed for 43 genes. Transcripts from 6 genes (p6.9, ORF 97, ORF 60, ORF 98, ORF 102 and chitinase) were reduced by 66% or more compared to the levels detected from the control virus. Microarray results were further examined by qPCR analysis of selected genes. In combination, these data show that deletion of the pp31 gene was not lethal and did not appear to affect viral DNA replication, but resulted in an apparent modest down-regulation of a subset of AcMNPV genes that included both early and late genes.

Yamagishi, Junya; Burnett, Erik D.; Harwood, Steven H.; Blissard, Gary W.

2009-01-01

23

Genome scanning in Haemophilus influenzae for identification of essential genes.  

PubMed

We have developed a method for identifying essential genes by using an in vitro transposition system, with a small (975 bp) insertional element containing an antibiotic resistance cassette, and mapping these inserts relative to the deduced open reading frames of Haemophilus influenzae by PCR and Southern analysis. Putative essential genes are identified by two methods: mutation exclusion or zero time analysis. Mutation exclusion consists of growing an insertional library and identifying open reading frames that do not contain insertional elements: in a growing population of bacteria, insertions in essential genes are excluded. Zero time analysis consists of monitoring the fate of individual insertions after transformation in a growing culture: the loss of inserts in essential genes is observed over time. Both methods of analysis permit the identification of genes required for bacterial survival. Details of the mutant library construction and the mapping strategy, examples of mutant exclusion, and zero time analysis are presented. PMID:10438768

Reich, K A; Chovan, L; Hessler, P

1999-08-01

24

Essential oil production: relationship with abundance of glandular trichomes in aerial surface of plants  

Microsoft Academic Search

The terpenoids, or isoprenoids, are a large family of natural products that are best known as constituents of the essential\\u000a oils in plants. Because of their pleasant flavor and aromatic properties, essential oils have an economic importance in perfumery,\\u000a cosmetic, pharmaceutical and various other industries. However, expression profiles of regulatory genes in essential oil production\\u000a have not been dissected entirely,

Kamal K. Biswas; Adam J. Foster; Theingi Aung; Soheil S. Mahmoud

2009-01-01

25

Experimental Determination and System Level Analysis of Essential Genes in Escherichia coli MG1655  

PubMed Central

Defining the gene products that play an essential role in an organism's functional repertoire is vital to understanding the system level organization of living cells. We used a genetic footprinting technique for a genome-wide assessment of genes required for robust aerobic growth of Escherichia coli in rich media. We identified 620 genes as essential and 3,126 genes as dispensable for growth under these conditions. Functional context analysis of these data allows individual functional assignments to be refined. Evolutionary context analysis demonstrates a significant tendency of essential E. coli genes to be preserved throughout the bacterial kingdom. Projection of these data over metabolic subsystems reveals topologic modules with essential and evolutionarily preserved enzymes with reduced capacity for error tolerance.

Gerdes, S. Y.; Scholle, M. D.; Campbell, J. W.; Balazsi, G.; Ravasz, E.; Daugherty, M. D.; Somera, A. L.; Kyrpides, N. C.; Anderson, I.; Gelfand, M. S.; Bhattacharya, A.; Kapatral, V.; D'Souza, M.; Baev, M. V.; Grechkin, Y.; Mseeh, F.; Fonstein, M. Y.; Overbeek, R.; Barabasi, A.-L.; Oltvai, Z. N.; Osterman, A. L.

2003-01-01

26

Processing of Vietnamese Essential Oils and Related Natural Products.  

National Technical Information Service (NTIS)

A project document on processing of Vietnamese Essential Oils and related natural products was drawn up between the United Nations Industrial Development Organization (UNIDO) and the Socialist Republic of Vietnam to develop an essential oils industry by u...

R. Gupta

1990-01-01

27

The endopolyphosphatase gene: Essential in Saccharomyces cerevisiae  

PubMed Central

Endopolyphosphatases (Ppn1) from yeast and animal cells hydrolyze inorganic polyphosphate (poly P) chains of many hundreds of phosphate residues into shorter lengths. The limit digest consists predominantly of chains of 60 (P60) and 3 (P3) Pi residues. Ppn1 of Saccharomyces cerevisiae, a homodimer of 35-kDa subunits (about 352-aa) is of vacuolar origin and requires the protease activation of a 75-kDa (674-aa) precursor polypeptide. The Ppn1 gene (PPN1) now has been cloned, sequenced, overexpressed, and deleted. That PPN1 encodes Ppn1 was verified by a 25-fold increase in Ppn1 when overexpressed under a GAL promoter and also by several peptide sequences that match exactly with sequences in a yeast genome ORF, the mutation of which abolishes Ppn1 activity. Null mutants in Ppn1 accumulate long-chain poly P and are defective in growth in minimal media. A double mutant of PPN1 and PPX1 (the gene encoding a potent exopolyphosphatase) loses viability rapidly in stationary phase. Whether this loss is a result of the excess of long-chain poly P or to the lack of shorter chains (i.e., poly P60 and P3) is unknown. Overexpression of the processed form of Ppn1 should provide a unique and powerful reagent to analyze poly P when the chain termini are unavailable to the actions of polyPase and poly P kinase.

Sethuraman, Anand; Rao, Narayana N.; Kornberg, Arthur

2001-01-01

28

Chromatin Regulation and Gene Centrality Are Essential for Controlling Fitness Pleiotropy in Yeast  

PubMed Central

Background There are a wide range of phenotypes that are due to loss-of-function or null mutations. Previously, the functions of gene products that distinguish essential from nonessential genes were characterized. However, the functions of products of non-essential genes that contribute to fitness remain minimally understood. Principal Findings Using data from Saccharomyces cerevisiae, we investigated several gene characteristics, which we are able to measure, that are significantly associated with a gene's fitness pleiotropy. Fitness pleiotropy is a measurement of the gene's importance to fitness. These characteristics include: 1) whether the gene's product functions in chromatin regulation, 2) whether the regulation of the gene is influenced by chromatin state, measured by chromatin regulation effect (CRE), 3) whether the gene's product functions as a transcription factor (TF) and the number of genes a TF regulates, 4) whether the gene contains TATA-box, and 5) whether the gene's product is central in a protein interaction network. Partial correlation analysis was used to study how these characteristics interact to influence fitness pleiotropy. We show that all five characteristics that were measured are statistically significantly associated with fitness pleiotropy. However, fitness pleiotropy is not associated with the presence of TATA-box when CRE is controlled. In particular, two characteristics: 1) whether the regulation of a gene is more likely to be influenced by chromatin state, and 2) whether the gene product is central in a protein interaction network measured by the number of protein interactions were found to play the most important roles affecting a gene's fitness pleiotropy. Conclusions These findings highlight the significance of both epigenetic gene regulation and protein interaction networks in influencing the fitness pleiotropy.

Zhou, Linqi; Ma, Xiaotu; Arbeitman, Michelle N.; Sun, Fengzhu

2009-01-01

29

Essential technical parameters for effective biogas production  

Microsoft Academic Search

Rising agrarian raw material prices in 2006 had a negative impact on the biogas sector in Germany, leading to the search for potentials for optimisation of production. A problem is the workload of the block-type thermal power station (BTPS). This is caused by biogas process disturbances, construction errors, technical problems and management mistakes as well as by oversizing of the

M. Schlegel; N. Kanswohl; D. Rössel; A. Sakalauskas

30

Aptamer-regulated expression of essential genes in yeast.  

PubMed

Conditional gene expression systems are important tools for the functional analysis of essential genes. Tetracycline (tc)-binding aptamers can be exploited as artificial riboswitches for the efficient control of gene expression by inserting them into the 5' untranslated region of an mRNA. The ligand-bound form of those mRNAs inhibits gene expression by interfering with translation initiation. In contrast to previous tc-dependent regulatory systems, where tc inhibits or activates transcription upon binding to the repressor protein TetR, the tc-binding aptamer system inhibits translation of the respective mRNA. We describe here a simple and powerful PCR-based strategy which allows easy tagging of any target gene in yeast using a tc aptamer-containing insertion cassette. The expression window can be adjusted with different promoters and protein synthesis is rapidly switched off. PMID:22160910

Suess, Beatrix; Entian, Karl-Dieter; Kötter, Peter; Weigand, Julia E

2012-01-01

31

Highly parallel identification of essential genes in cancer cells.  

PubMed

More complete knowledge of the molecular mechanisms underlying cancer will improve prevention, diagnosis and treatment. Efforts such as The Cancer Genome Atlas are systematically characterizing the structural basis of cancer, by identifying the genomic mutations associated with each cancer type. A powerful complementary approach is to systematically characterize the functional basis of cancer, by identifying the genes essential for growth and related phenotypes in different cancer cells. Such information would be particularly valuable for identifying potential drug targets. Here, we report the development of an efficient, robust approach to perform genome-scale pooled shRNA screens for both positive and negative selection and its application to systematically identify cell essential genes in 12 cancer cell lines. By integrating these functional data with comprehensive genetic analyses of primary human tumors, we identified known and putative oncogenes such as EGFR, KRAS, MYC, BCR-ABL, MYB, CRKL, and CDK4 that are essential for cancer cell proliferation and also altered in human cancers. We further used this approach to identify genes involved in the response of cancer cells to tumoricidal agents and found 4 genes required for the response of CML cells to imatinib treatment: PTPN1, NF1, SMARCB1, and SMARCE1, and 5 regulators of the response to FAS activation, FAS, FADD, CASP8, ARID1A and CBX1. Broad application of this highly parallel genetic screening strategy will not only facilitate the rapid identification of genes that drive the malignant state and its response to therapeutics but will also enable the discovery of genes that participate in any biological process. PMID:19091943

Luo, Biao; Cheung, Hiu Wing; Subramanian, Aravind; Sharifnia, Tanaz; Okamoto, Michael; Yang, Xiaoping; Hinkle, Greg; Boehm, Jesse S; Beroukhim, Rameen; Weir, Barbara A; Mermel, Craig; Barbie, David A; Awad, Tarif; Zhou, Xiaochuan; Nguyen, Tuyen; Piqani, Bruno; Li, Cheng; Golub, Todd R; Meyerson, Matthew; Hacohen, Nir; Hahn, William C; Lander, Eric S; Sabatini, David M; Root, David E

2008-12-17

32

Shielding Production: An Essential Step in Production Control  

Microsoft Academic Search

Effective production control systems are structured around the assignment as the unit of analysis. The quality of work assignments to production units such as construction crews and engineering squads is the key to production control and a key factor determining production unit productivity. Research has revealed that the quality of assignments can be substantially improved by forming and selecting them

Glenn Ballard; Gregory Howell

1997-01-01

33

Probing the limits of genetic recoding in essential genes.  

PubMed

Engineering radically altered genetic codes will allow for genomically recoded organisms that have expanded chemical capabilities and are isolated from nature. We have previously reassigned the translation function of the UAG stop codon; however, reassigning sense codons poses a greater challenge because such codons are more prevalent, and their usage regulates gene expression in ways that are difficult to predict. To assess the feasibility of radically altering the genetic code, we selected a panel of 42 highly expressed essential genes for modification. Across 80 Escherichia coli strains, we removed all instances of 13 rare codons from these genes and attempted to shuffle all remaining codons. Our results suggest that the genome-wide removal of 13 codons is feasible; however, several genome design constraints were apparent, underscoring the importance of a strategy that rapidly prototypes and tests many designs in small pieces. PMID:24136967

Lajoie, M J; Kosuri, S; Mosberg, J A; Gregg, C J; Zhang, D; Church, G M

2013-10-18

34

A crtB homolog essential for photochromogenicity in Mycobacterium marinum: isolation, characterization, and gene disruption via homologous recombination.  

PubMed Central

A gene essential for light-induced pigment production was isolated from the photochromogen Mycobacterium marinum by heterologous complementation of an M. marinum cosmid library in the nonchromogen Mycobacterium smegmatis. This gene is part of an operon and homologous to the Streptomyces griseus and Myxococcus xanthus crtB genes encoding phytoene synthase. Gene replacement at this locus was achieved via homologous recombination, demonstrating that its expression is essential for photochromogenicity. The ease of targeted gene disruption in this pathogenic Mycobacterium allows for the dissection of the molecular basis of mycobacterial pathogenesis.

Ramakrishnan, L; Tran, H T; Federspiel, N A; Falkow, S

1997-01-01

35

Essential genes in thyroid cancers: focus on fascin  

PubMed Central

Although thyroid cancers are not among common malignancies, they rank as the first prevalent endocrine cancers in human. According to the results of published studies it has been shown the gradual progress from normal to the neoplastic cell in the process of tumor formation is the result of sequential genetic events. Among them we may point the mutations and rearrangements occurred in a group of proto-oncogenes, transcription factors and metastasis elements such as P53, RAS,RET,BRAF, PPAR? and Fascin. In the present article,we reviewed the most important essential genes in thyroid cancers, the role of epithelial mesenchymal transition and Fascin has been highlighted in this paper.

2013-01-01

36

Identifying essential genes in bacterial metabolic networks with machine learning methods  

Microsoft Academic Search

BACKGROUND: Identifying essential genes in bacteria supports to identify potential drug targets and an understanding of minimal requirements for a synthetic cell. However, experimentally assaying the essentiality of their coding genes is resource intensive and not feasible for all bacterial organisms, in particular if they are infective. RESULTS: We developed a machine learning technique to identify essential genes using the

Kitiporn Plaimas; Roland Eils; Rainer König

2010-01-01

37

Molecular genetic analysis of a vaccinia virus gene with an essential role in DNA replication.  

PubMed Central

We have identified a gene encoded by vaccinia virus which is essential for DNA replication. The gene, located in the HindIII D fragment of the viral genome, is transcribed early after infection into two transcripts of 3.0 and 3.7 kilobases which share a 3' terminus. The lesions of three temperature-sensitive DNA replication mutants with defects in this gene have been localized by marker rescue with progressively smaller DNA fragments. We have determined by hybrid selection that the gene encodes an 82-kilodalton protein. An antibody has been prepared against this polypeptide and used to quantitate expression of the protein after infection with wild-type virus or with a viral mutant whose lesion maps within this gene. The temporal pattern of expression in the mutant is unaffected, but the product encoded by the mutant is significantly more thermolabile than the wild-type protein. Images

Evans, E; Traktman, P

1987-01-01

38

Partial AUC maximization for essential gene prediction using genetic algorithms.  

PubMed

Identifying genes indispensable for an organism's life and their characteristics is one of the central questions in current biological research, and hence it would be helpful to develop computational approaches towards the prediction of essential genes. The performance of a predictor is usually measured by the area under the receiver operating characteristic curve (AUC). We propose a novel method by implementing genetic algorithms to maximize the partial AUC that is restricted to a specific interval of lower false positive rate (FPR), the region relevant to follow-up experimental validation. Our predictor uses various features based on sequence information, protein-protein interaction network topology, and gene expression profiles. A feature selection wrapper was developed to alleviate the over-fitting problem and to weigh each feature's relevance to prediction. We evaluated our method using the proteome of budding yeast. Our implementation of genetic algorithms maximizing the partial AUC below 0.05 or 0.10 of FPR outperformed other popular classification methods. PMID:23351383

Hwang, Kyu-Baek; Ha, Beom-Yong; Ju, Sanghun; Kim, Sangsoo

2013-01-01

39

The tobacco plastid accD gene is essential and is required for leaf development.  

PubMed

Angiosperm plastid genomes typically encode approximately 80 polypeptides, mainly specifying plastid-localized functions such as photosynthesis and gene expression. Plastid protein synthesis and expression of the plastid clpP1 gene are essential for development in tobacco, indicating the presence of one or more plastid genes whose influence extends beyond the plastid compartment. The plastid accD gene encodes the beta-carboxyl transferase subunit of acetyl-CoA carboxylase and is present in the plastids of most flowering plants, including non-photosynthetic parasitic plants. We replaced the wild-type accD gene with an aadA-disrupted mutant allele using homologous recombination. Persistent heteroplasmy in the presence of antibiotics indicated that the wild-type accD allele was essential. The phenotype of the accD knockout was revealed in plastid transformants grown in the absence of antibiotics. Leaves contained pale green sectors and lacked part or all of the leaf lamina due to arrested division or loss of cells. Abnormal structures were present in plastids found in mutant plants, indicating that accD might be required to maintain the plastid compartment. Loss of the plastid compartment would be expected to be lethal. These results provide genetic evidence showing the essential role of plastid ACCase in the pathway leading to the synthesis of products required for the extra-plastidic processes needed for leaf development. PMID:16212603

Kode, Vasumathi; Mudd, Elisabeth A; Iamtham, Siriluck; Day, Anil

2005-10-01

40

Characterization of the angR Gene of Vibrio anguillarum: Essential Role in Virulence  

PubMed Central

The ability to utilize the iron bound by high-affinity iron-binding proteins in the vertebrate host is an important virulence factor for the marine fish pathogen Vibrio anguillarum. Virulence in septicemic infections is due to the presence of a highly efficient plasmid-encoded iron transport system. AngR, a 110-kDa protein component of this system, appears to play a role in both regulation of the expression of the iron transport genes fatDCBA and the production of the siderophore anguibactin. Therefore, study of the expression of the angR gene and the properties of its product, the AngR protein, may contribute to the understanding of the mechanisms of virulence of this pathogen. In this work, we present genetic and molecular evidence from transposition mutagenesis experiments and RNA analysis that angR, which maps immediately downstream of the fatA gene, is part of a polycistronic transcript that also includes the iron transport genes fatDCBA and angT, a gene located downstream of angR which showed domain homology to certain thioesterases involved in nonribosomal peptide synthesis of siderophores and antibiotics. In order to dissect the specific domains of AngR associated with regulation of iron transport gene expression, anguibactin production, and virulence, we also generated a panel of site-directed angR mutants, as well as deletion derivatives. Both virulence and anguibactin production were dramatically affected by each one of the angR modifications. In contrast to the need for an intact AngR molecule for anguibactin production and virulence, the regulation of iron transport gene expression does not require the entire AngR molecule, since truncation of the carboxy terminus carrying the nonribosomal peptide synthetase cores, as well as the site-directed mutations, resulted in derivatives that retained their ability to regulate gene expression which was only abolished after truncation of amino-terminal sequences containing helix-turn-helix and leucine zipper motifs and a specialized heterocyclization and condensation domain found in certain nonribosomal peptide synthetases. The evidence, while not rigorously eliminating the possibility that a separate regulatory polypeptide exists and is encoded somewhere within the 5?-end region of the angR gene, strongly supports the idea that AngR is a bifunctional protein and that it plays an essential role in the virulence mechanisms of V. anguillarum. We also show in this study that the angT gene, found downstream of angR, intervenes in the mechanism of anguibactin production but is not essential for virulence or iron transport gene expression.

Wertheimer, A. M.; Verweij, W.; Chen, Q.; Crosa, L. M.; Nagasawa, M.; Tolmasky, M. E.; Actis, L. A.; Crosa, J. H.

1999-01-01

41

Concurrent Growth Rate and Transcript Analyses Reveal Essential Gene Stringency in Escherichia coli  

Microsoft Academic Search

BackgroundGenes essential for bacterial growth are of particular scientific interest. Many putative essential genes have been identified or predicted in several species, however, little is known about gene expression requirement stringency, which may be an important aspect of bacterial physiology and likely a determining factor in drug target development.Methodology\\/Principal FindingsWorking from the premise that essential genes differ in absolute requirement

Shan Goh; Jaroslaw M. Boberek; Nobutaka Nakashima; Jem Stach; Liam Good; Laurent Rénia

2009-01-01

42

A postgenomic method for predicting essential genes at subsaturation levels of mutagenesis: Application to Mycobacterium tuberculosis  

Microsoft Academic Search

We describe a postgenomic in silico approach for identifying genes that are likely to be essential and estimate their proportion in haploid genomes. With the knowledge of all sites eligible for mutagenesis and an experimentally determined partial list of nonessential genes from genome mutagenesis, a Bayesian statistical method provides reasonable predictions of essential genes with a subsaturation level of random

Gyanu Lamichhane; Matteo Zignol; Natalie J. Blades; Deborah E. Geiman; Annette Dougherty; Jacques Grosset; Karl W. Broman; William R. Bishai

2003-01-01

43

The Essential Gene EMB1611 Maintains Shoot Apical Meristem Function During Arabidopsis Development  

Technology Transfer Automated Retrieval System (TEKTRAN)

The Arabidopsis thaliana genome contains hundreds of genes essential for seed development. Because null mutations in these genes cause embryo lethality, their specific molecular and developmental functions are largely unknown. Here, we identify a role for EMB1611/MEE22, an essential gene in Arabidop...

44

Interdomain lateral gene transfer of an essential ferrochelatase gene in human parasitic nematodes  

PubMed Central

Lateral gene transfer events between bacteria and animals highlight an avenue for evolutionary genomic loss/gain of function. Herein, we report functional lateral gene transfer in animal parasitic nematodes. Members of the Nematoda are heme auxotrophs, lacking the ability to synthesize heme; however, the human filarial parasite Brugia malayi has acquired a bacterial gene encoding ferrochelatase (BmFeCH), the terminal step in heme biosynthesis. BmFeCH, encoded by a 9-exon gene, is a mitochondrial-targeted, functional ferrochelatase based on enzyme assays, complementation, and inhibitor studies. Homologs have been identified in several filariae and a nonfilarial nematode. RNAi and ex vivo inhibitor experiments indicate that BmFeCH is essential for viability, validating it as a potential target for filariasis control.

Wu, Bo; Novelli, Jacopo; Jiang, Daojun; Dailey, Harry A.; Landmann, Frederic; Ford, Louise; Taylor, Mark J.; Carlow, Clotilde K. S.; Kumar, Sanjay; Foster, Jeremy M.; Slatko, Barton E.

2013-01-01

45

A Statistical Framework for Improving Genomic Annotations of Prokaryotic Essential Genes  

PubMed Central

Large-scale systematic analysis of gene essentiality is an important step closer toward unraveling the complex relationship between genotypes and phenotypes. Such analysis cannot be accomplished without unbiased and accurate annotations of essential genes. In current genomic databases, most of the essential gene annotations are derived from whole-genome transposon mutagenesis (TM), the most frequently used experimental approach for determining essential genes in microorganisms under defined conditions. However, there are substantial systematic biases associated with TM experiments. In this study, we developed a novel Poisson model–based statistical framework to simulate the TM insertion process and subsequently correct the experimental biases. We first quantitatively assessed the effects of major factors that potentially influence the accuracy of TM and subsequently incorporated relevant factors into the framework. Through iteratively optimizing parameters, we inferred the actual insertion events occurred and described each gene’s essentiality on probability measure. Evaluated by the definite mapping of essential gene profile in Escherichia coli, our model significantly improved the accuracy of original TM datasets, resulting in more accurate annotations of essential genes. Our method also showed encouraging results in improving subsaturation level TM datasets. To test our model’s broad applicability to other bacteria, we applied it to Pseudomonas aeruginosa PAO1 and Francisella tularensis novicida TM datasets. We validated our predictions by literature as well as allelic exchange experiments in PAO1. Our model was correct on six of the seven tested genes. Remarkably, among all three cases that our predictions contradicted the TM assignments, experimental validations supported our predictions. In summary, our method will be a promising tool in improving genomic annotations of essential genes and enabling large-scale explorations of gene essentiality. Our contribution is timely considering the rapidly increasing essential gene sets. A Webserver has been set up to provide convenient access to this tool. All results and source codes are available for download upon publication at http://research.cchmc.org/essentialgene/.

Deng, Jingyuan; Su, Shengchang; Lin, Xiaodong; Hassett, Daniel J.; Lu, Long Jason

2013-01-01

46

The ROX3 gene encodes an essential nuclear protein involved in CYC7 gene expression in Saccharomyces cerevisiae.  

PubMed Central

The ROX3 gene was identified during a hunt for mutants with increased expression of the heme-regulated CYC7 gene, which encodes the minor species of cytochrome c in the yeast Saccharomyces cerevisiae. The rox3 mutants caused a 10-fold increase in CYC7 expression both in the presence and absence of heme, had slightly increased anaerobic expression of the heme-activated CYC1 gene, and caused decreases in the anaerobic expression of the heme-repressed ANB1 gene and the aerobic expression of its heme-induced homolog. The wild-type ROX3 gene was cloned, and the sequence indicated that it encodes a 220-amino-acid protein. This protein is essential; deletion of the coding sequence was lethal. The coding sequence for beta-galactosidase was fused to the 3' end of the ROX3 coding sequence, and the fusion product was found to be localized in the nucleus, strongly suggesting that the wild-type protein carries out a nuclear function. Mutations in the rox3 gene showed an interesting pattern of intragenic complementation. A deletion of the 5' coding region complemented a nonsense mutation at codon 128 but could not prevent the lethality of the null mutation. These results suggest that the amino-terminal domain is required for an essential function, while the carboxy-terminal domain can be supplied in trans to achieve the wild-type expression of CYC7. Finally, RNA blots demonstrated that the ROX3 mRNA was expressed at higher levels anaerobically but was not subject to heme repression. The nuclear localization and the lack of viability of null mutants suggest that the ROX3 protein is a general regulatory factor. Images

Rosenblum-Vos, L S; Rhodes, L; Evangelista, C C; Boayke, K A; Zitomer, R S

1991-01-01

47

Evolutionary conservation of essential and highly expressed genes in Pseudomonas aeruginosa  

Microsoft Academic Search

BACKGROUND: The constant increase in development and spread of bacterial resistance to antibiotics poses a serious threat to human health. New sequencing technologies are now on the horizon that will yield massive increases in our capacity for DNA sequencing and will revolutionize the drug discovery process. Since essential genes are promising novel antibiotic targets, the prediction of gene essentiality based

Andreas Dötsch; Frank Klawonn; Michael Jarek; Maren Scharfe; Helmut Blöcker; Susanne Häussler

2010-01-01

48

Investigating the predictability of essential genes across distantly related organisms using an integrative approach  

PubMed Central

Rapid and accurate identification of new essential genes in under-studied microorganisms will significantly improve our understanding of how a cell works and the ability to re-engineer microorganisms. However, predicting essential genes across distantly related organisms remains a challenge. Here, we present a machine learning-based integrative approach that reliably transfers essential gene annotations between distantly related bacteria. We focused on four bacterial species that have well-characterized essential genes, and tested the transferability between three pairs among them. For each pair, we trained our classifier to learn traits associated with essential genes in one organism, and applied it to make predictions in the other. The predictions were then evaluated by examining the agreements with the known essential genes in the target organism. Ten-fold cross-validation in the same organism yielded AUC scores between 0.86 and 0.93. Cross-organism predictions yielded AUC scores between 0.69 and 0.89. The transferability is likely affected by growth conditions, quality of the training data set and the evolutionary distance. We are thus the first to report that gene essentiality can be reliably predicted using features trained and tested in a distantly related organism. Our approach proves more robust and portable than existing approaches, significantly extending our ability to predict essential genes beyond orthologs.

Deng, Jingyuan; Deng, Lei; Su, Shengchang; Zhang, Minlu; Lin, Xiaodong; Wei, Lan; Minai, Ali A.; Hassett, Daniel J.; Lu, Long J.

2011-01-01

49

Investigating the predictability of essential genes across distantly related organisms using an integrative approach.  

PubMed

Rapid and accurate identification of new essential genes in under-studied microorganisms will significantly improve our understanding of how a cell works and the ability to re-engineer microorganisms. However, predicting essential genes across distantly related organisms remains a challenge. Here, we present a machine learning-based integrative approach that reliably transfers essential gene annotations between distantly related bacteria. We focused on four bacterial species that have well-characterized essential genes, and tested the transferability between three pairs among them. For each pair, we trained our classifier to learn traits associated with essential genes in one organism, and applied it to make predictions in the other. The predictions were then evaluated by examining the agreements with the known essential genes in the target organism. Ten-fold cross-validation in the same organism yielded AUC scores between 0.86 and 0.93. Cross-organism predictions yielded AUC scores between 0.69 and 0.89. The transferability is likely affected by growth conditions, quality of the training data set and the evolutionary distance. We are thus the first to report that gene essentiality can be reliably predicted using features trained and tested in a distantly related organism. Our approach proves more robust and portable than existing approaches, significantly extending our ability to predict essential genes beyond orthologs. PMID:20870748

Deng, Jingyuan; Deng, Lei; Su, Shengchang; Zhang, Minlu; Lin, Xiaodong; Wei, Lan; Minai, Ali A; Hassett, Daniel J; Lu, Long J

2010-09-24

50

A statistical framework for improving genomic annotations of prokaryotic essential genes.  

PubMed

Large-scale systematic analysis of gene essentiality is an important step closer toward unraveling the complex relationship between genotypes and phenotypes. Such analysis cannot be accomplished without unbiased and accurate annotations of essential genes. In current genomic databases, most of the essential gene annotations are derived from whole-genome transposon mutagenesis (TM), the most frequently used experimental approach for determining essential genes in microorganisms under defined conditions. However, there are substantial systematic biases associated with TM experiments. In this study, we developed a novel Poisson model-based statistical framework to simulate the TM insertion process and subsequently correct the experimental biases. We first quantitatively assessed the effects of major factors that potentially influence the accuracy of TM and subsequently incorporated relevant factors into the framework. Through iteratively optimizing parameters, we inferred the actual insertion events occurred and described each gene's essentiality on probability measure. Evaluated by the definite mapping of essential gene profile in Escherichia coli, our model significantly improved the accuracy of original TM datasets, resulting in more accurate annotations of essential genes. Our method also showed encouraging results in improving subsaturation level TM datasets. To test our model's broad applicability to other bacteria, we applied it to Pseudomonas aeruginosa PAO1 and Francisella tularensis novicida TM datasets. We validated our predictions by literature as well as allelic exchange experiments in PAO1. Our model was correct on six of the seven tested genes. Remarkably, among all three cases that our predictions contradicted the TM assignments, experimental validations supported our predictions. In summary, our method will be a promising tool in improving genomic annotations of essential genes and enabling large-scale explorations of gene essentiality. Our contribution is timely considering the rapidly increasing essential gene sets. A Webserver has been set up to provide convenient access to this tool. All results and source codes are available for download upon publication at http://research.cchmc.org/essentialgene/. PMID:23520492

Deng, Jingyuan; Su, Shengchang; Lin, Xiaodong; Hassett, Daniel J; Lu, Long Jason

2013-03-08

51

Systematic Identification of Novel, Essential Host Genes Affecting Bromovirus RNA Replication  

PubMed Central

Positive-strand RNA virus replication involves viral proteins and cellular proteins at nearly every replication step. Brome mosaic virus (BMV) is a well-established model for dissecting virus-host interactions and is one of very few viruses whose RNA replication, gene expression and encapsidation have been reproduced in the yeast Saccharomyces cerevisiae. Previously, our laboratory identified ?100 non-essential host genes whose loss inhibited or enhanced BMV replication at least 3-fold. However, our isolation of additional BMV-modulating host genes by classical genetics and other results underscore that genes essential for cell growth also contribute to BMV RNA replication at a frequency that may be greater than that of non-essential genes. To systematically identify novel, essential host genes affecting BMV RNA replication, we tested a collection of ?900 yeast strains, each with a single essential gene promoter replaced by a doxycycline-repressible promoter, allowing repression of gene expression by adding doxycycline to the growth medium. Using this strain array of ?81% of essential yeast genes, we identified 24 essential host genes whose depleted expression reproducibly inhibited or enhanced BMV RNA replication. Relevant host genes are involved in ribosome biosynthesis, cell cycle regulation and protein homeostasis, among other cellular processes. BMV 2aPol levels were significantly increased in strains depleted for a heat shock protein (HSF1) or proteasome components (PRE1 and RPT6), suggesting these genes may affect BMV RNA replication by directly or indirectly modulating 2aPol localization, post-translational modification or interacting partners. Investigating the diverse functions of these newly identified essential host genes should advance our understanding of BMV-host interactions and normal cellular pathways, and suggest new modes of virus control.

Gancarz, Brandi L.; Hao, Linhui; He, Qiuling; Newton, Michael A.; Ahlquist, Paul

2011-01-01

52

An ABC Transporter from Bacillus thuringiensis Is Essential for ?-Exotoxin I Production  

PubMed Central

?-Exotoxin I is a nonspecific insecticidal metabolite secreted by some Bacillus thuringiensis strains. Several studies of B. thuringiensis strains that have lost the capacity to produce ?-exotoxin I have suggested that there is a strong correlation between high levels of ?-exotoxin I production and the ability to synthesize crystal proteins. In this study, we showed that a mutant strain, B. thuringiensis 407-1(Cry?)(Pig+), with no crystal gene, produced considerable amounts of ?-exotoxin I together with a soluble brown melanin pigment. Therefore, ?-exotoxin I production can take place after a strain has lost the plasmids bearing the cry genes, which suggests that these curable plasmids probably contain determinants involved in the regulation of ?-exotoxin I production. Using a mini-Tn10 transposon, we constructed a library of strain 407-1(Cry?)(Pig+) mutants. We screened for nonpigmented mutants with impaired ?-exotoxin I production and identified a genetic locus harboring two genes (berA and berB) essential for ?-exotoxin I production. The deduced amino acid sequence of the berA gene displayed significant similarity to the ATP-binding domains of the DRI (drug resistance and immunity) family of ATP-binding cassette (ABC) proteins involved in drug resistance and immunity to bacteriocins and lantibiotics. The berB gene encodes a protein with six putative transmembrane helices, which probably constitutes the integral membrane component of the transporter. The demonstration that berAB is required for ?-exotoxin I production and/or resistance in B. thuringiensis adds an adenine nucleotide analog to the wide range of substrates of the superfamily of ABC proteins. We suggest that berAB confers ?-exotoxin I immunity in B. thuringiensis, through active efflux of the molecule.

Espinasse, Sylvain; Gohar, Michel; Lereclus, Didier; Sanchis, Vincent

2002-01-01

53

An ABC transporter from Bacillus thuringiensis is essential for beta-exotoxin I production.  

PubMed

beta-Exotoxin I is a nonspecific insecticidal metabolite secreted by some Bacillus thuringiensis strains. Several studies of B. thuringiensis strains that have lost the capacity to produce beta-exotoxin I have suggested that there is a strong correlation between high levels of beta-exotoxin I production and the ability to synthesize crystal proteins. In this study, we showed that a mutant strain, B. thuringiensis 407-1(Cry(-))(Pig(+)), with no crystal gene, produced considerable amounts of beta-exotoxin I together with a soluble brown melanin pigment. Therefore, beta-exotoxin I production can take place after a strain has lost the plasmids bearing the cry genes, which suggests that these curable plasmids probably contain determinants involved in the regulation of beta-exotoxin I production. Using a mini-Tn10 transposon, we constructed a library of strain 407-1(Cry(-))(Pig(+)) mutants. We screened for nonpigmented mutants with impaired beta-exotoxin I production and identified a genetic locus harboring two genes (berA and berB) essential for beta-exotoxin I production. The deduced amino acid sequence of the berA gene displayed significant similarity to the ATP-binding domains of the DRI (drug resistance and immunity) family of ATP-binding cassette (ABC) proteins involved in drug resistance and immunity to bacteriocins and lantibiotics. The berB gene encodes a protein with six putative transmembrane helices, which probably constitutes the integral membrane component of the transporter. The demonstration that berAB is required for beta-exotoxin I production and/or resistance in B. thuringiensis adds an adenine nucleotide analog to the wide range of substrates of the superfamily of ABC proteins. We suggest that berAB confers beta-exotoxin I immunity in B. thuringiensis, through active efflux of the molecule. PMID:12374817

Espinasse, Sylvain; Gohar, Michel; Lereclus, Didier; Sanchis, Vincent

2002-11-01

54

From Mouse to Human: Evolutionary Genomics Analysis of Human Orthologs of Essential Genes  

PubMed Central

Understanding the core set of genes that are necessary for basic developmental functions is one of the central goals in biology. Studies in model organisms identified a significant fraction of essential genes through the analysis of null-mutations that lead to lethality. Recent large-scale next-generation sequencing efforts have provided unprecedented data on genetic variation in human. However, evolutionary and genomic characteristics of human essential genes have never been directly studied on a genome-wide scale. Here we use detailed phenotypic resources available for the mouse and deep genomics sequencing data from human populations to characterize patterns of genetic variation and mutational burden in a set of 2,472 human orthologs of known essential genes in the mouse. Consistent with the action of strong, purifying selection, these genes exhibit comparatively reduced levels of sequence variation, skew in allele frequency towards more rare, and exhibit increased conservation across the primate and rodent lineages relative to the remainder of genes in the genome. In individual genomes we observed ?12 rare mutations within essential genes predicted to be damaging. Consistent with the hypothesis that mutations in essential genes are risk factors for neurodevelopmental disease, we show that de novo variants in patients with Autism Spectrum Disorder are more likely to occur in this collection of genes. While incomplete, our set of human orthologs shows characteristics fully consistent with essential function in human and thus provides a resource to inform and facilitate interpretation of sequence data in studies of human disease.

Georgi, Benjamin; Voight, Benjamin F.; Bucan, Maja

2013-01-01

55

From mouse to human: evolutionary genomics analysis of human orthologs of essential genes.  

PubMed

Understanding the core set of genes that are necessary for basic developmental functions is one of the central goals in biology. Studies in model organisms identified a significant fraction of essential genes through the analysis of null-mutations that lead to lethality. Recent large-scale next-generation sequencing efforts have provided unprecedented data on genetic variation in human. However, evolutionary and genomic characteristics of human essential genes have never been directly studied on a genome-wide scale. Here we use detailed phenotypic resources available for the mouse and deep genomics sequencing data from human populations to characterize patterns of genetic variation and mutational burden in a set of 2,472 human orthologs of known essential genes in the mouse. Consistent with the action of strong, purifying selection, these genes exhibit comparatively reduced levels of sequence variation, skew in allele frequency towards more rare, and exhibit increased conservation across the primate and rodent lineages relative to the remainder of genes in the genome. In individual genomes we observed ~12 rare mutations within essential genes predicted to be damaging. Consistent with the hypothesis that mutations in essential genes are risk factors for neurodevelopmental disease, we show that de novo variants in patients with Autism Spectrum Disorder are more likely to occur in this collection of genes. While incomplete, our set of human orthologs shows characteristics fully consistent with essential function in human and thus provides a resource to inform and facilitate interpretation of sequence data in studies of human disease. PMID:23675308

Georgi, Benjamin; Voight, Benjamin F; Bu?an, Maja

2013-05-09

56

The mouse polyubiquitin gene Ubb is essential for meiotic progression  

Microsoft Academic Search

Ubiquitin is encoded in mice by two polyubiquitin genes, Ubb and Ubc, that are considered to be stress inducible and two constitutively expressed monoubiquitin (Uba) genes. Here we report that targeted disruption of Ubb results in male and female infertility due to failure of germ cells to progress through meiosis I and hypogonadism. In the absence of Ubb, spermatocytes and

Kwon-Yul Ryu; Shamim A. Sinnar; Laura G. Reinholdt; Sergio Vaccari; S. Hall; M. A. Garcia; T. S. Zaitseva; D. M. Bouley; K. Boekelheide; M. A. Handel; M. Conti; R. R. Kopito

2008-01-01

57

Gene Targeting Approaches to Complex Genetic Diseases: Atherosclerosis and Essential Hypertension  

Microsoft Academic Search

Gene targeting allows precise, predetermined changes to be made in a chosen gene in the mouse genome. To date, targeting has been used most often for generation of animals completely lacking the product of a gene of interest. The resulting \\

Oliver Smithies; Nobuyo Maeda

1995-01-01

58

A pestivirus DNA vaccine based on a non-antibiotic resistance Escherichia coli essential gene marker.  

PubMed

Antibiotic resistance genes are widely used to produce plasmid DNA vaccines, but risk unwanted exposure to antibiotic residues and the spread of resistance genes. To overcome the limitations of existing selection technologies, we developed an alternative system applying the widely used household biocide triclosan as the selective agent and an endogenous growth essential target gene, fabI, as the plasmid-borne marker in Escherichia coli. The fabI/triclosan system enables efficient, non-antibiotic selection of transformed bacteria, with improved safety and plasmid production features. Here we aimed to evaluate the performance of this non-antibiotic selection system using a plasmid DNA vaccine against bovine viral diarrhoea virus as an example. The new system displayed high-yield plasmid DNA production in a standard E. coli host strain and growth media. Notably, the purified pDNA provided efficient in vitro protein expression and a strong in vivo neutralising antibody response in a mouse model, with measures comparable to that of the parental plasmid DNA based on ampicillin resistance. The fabI/triclosan system requires only low levels of triclosan for selection (1 ?M) and residual triclosan in isolated DNA was below the limit of detection (< 20 parts per trillion). The fabI/triclosan selection system provides a simple, non-antibiotic resistance marker for plasmid selection, applicable to DNA vaccines and possibly other recombinant vaccine applications. PMID:22212129

El-Attar, Laila M R; Scott, Sally; Goh, Shan; Good, Liam

2011-12-31

59

Burkholderia cenocepacia conditional growth mutant library created by random promoter replacement of essential genes.  

PubMed

Identification of essential genes by construction of conditional knockouts with inducible promoters allows the identification of essential genes and creation of conditional growth (CG) mutants that are then available as genetic tools for further studies. We used large-scale transposon delivery of the rhamnose-inducible promoter, PrhaB followed by robotic screening of rhamnose-dependent growth to construct a genomic library of 106 Burkholderia cenocepacia CG mutants. Transposon insertions were found where PrhaB was in the same orientation of widely conserved, well-characterized essential genes as well as genes with no previous records of essentiality in other microorganisms. Using previously reported global gene-expression analyses, we demonstrate that PrhaB can achieve the wide dynamic range of expression levels required for essential genes when the promoter is delivered randomly and mutants with rhamnose-dependent growth are selected. We also show specific detection of the target of an antibiotic, novobiocin, by enhanced sensitivity of the corresponding CG mutant (PrhaB controlling gyrB expression) within the library. Modulation of gene expression to achieve 30-60% of wild-type growth created conditions for specific hypersensitivity demonstrating the value of the CG mutant library for chemogenomic experiments. In summary, CG mutants can be obtained on a large scale by random delivery of a tightly regulated inducible promoter into the bacterial chromosome followed by a simple screening for the CG phenotype, without previous information on gene essentiality. PMID:23389959

Bloodworth, Ruhi A M; Gislason, April S; Cardona, Silvia T

2013-02-07

60

Quorum sensing modulation of a putative glycosyltransferase gene cluster essential for Xanthomonas campestris biofilm formation.  

PubMed

Findings from previous studies suggest that the quorum sensing signal DSF (diffusible signal factor) negatively regulates biofilm formation in Xanthomonas campestris pv. campestris (Xcc) by affecting the expression of manA encoding biofilm dispersion and an unknown factor(s). In this study, by analysing the double deletion mutant ?rpfF?manA, in which DSF biosynthesis gene rpfF and biofilm dispersal gene manA were deleted, we found that DSF modulated biofilm development by suppression of a mechanism essential for biofilm formation. Transposon mutagenesis of ?rpfF?manA and subsequent analyses led to the identification of a novel gene locus xagABC encoding a putative glycosyl transferase system. Genetic analysis revealed that the transcriptional expression of xagABC was negatively regulated by DSF through the RpfC/RpfG two-component regulatory system. Deletion of the xag genes resulted in decreased extracellular polysaccharide production, abolished Xcc biofilm formation and attenuated the bacterial resistance to oxidative stress. Furthermore, we provide evidence that xagABC and manA were differentially expressed in Xcc and the biofilm formed by overexpression of xagABC in wild-type Xcc could be dispersed by ManA. These results provide new insight into the molecular mechanisms by which Xcc switches between planktonic growth and biofilm lifestyle. PMID:20636376

Tao, Fei; Swarup, Sanjay; Zhang, Lian-Hui

2010-12-01

61

Chromogranin A: a novel susceptibility gene for essential hypertension  

Microsoft Academic Search

Chromogranin A (CHGA) is ubiquitously expressed in secretory cells of the endocrine, neuroendocrine, and neuronal tissues.\\u000a Although this protein has long been known as a marker for neuroendocrine tumors, its role in cardiovascular disease states\\u000a including essential hypertension (EH) has only recently been recognized. It acts as a prohormone giving rise to bioactive\\u000a peptides such as vasostatin-I (human CHGA1–76) and

Bhavani S. Sahu; Parshuram J. Sonawane; Nitish R. Mahapatra

2010-01-01

62

The fission yeast dis3+ gene encodes a 110-kDa essential protein implicated in mitotic control.  

PubMed

The fission yeast mutant dis3-54 is defective in mitosis and fails in chromosome disjunction. Its phenotype is similar to that of dis2-11, a mutant with a mutation in the type 1 protein phosphatase gene. We cloned the dis3+ gene by transformation. Nucleotide sequencing predicts a coding region of 970 amino acids interrupted by a 164-bp intron at the 65th codon. The predicted dis3+ protein shares a weak but significant similarity with the budding yeast SSD1 or SRK1 gene product, the gene for which is a suppressor for the absence of a protein phosphatase SIT4 gene or the BCY1 regulatory subunit of cyclic AMP-dependent protein kinase. Anti-dis3 antibodies recognized the 110-kDa dis3+ gene product, which is part of a 250- to 350-kDa oligomer and is enriched in the nucleus. The cellular localization of the dis3+ protein is reminiscent of that of the dis2+ protein, but these two proteins do not form a complex. A type 1 protein phosphatase activity in the dis3-54 mutant extracts is apparently not affected. The dis3+ gene is essential for growth; gene disruptant cells do not germinate and fail in cell division. Increased dis3+ gene dosage reverses the Ts+ phenotype of a cdc25 wee1 strain, as does increased type 1 protein phosphatase gene dosage. Double mutant dis3 dis2 is lethal even at the permissive temperature, suggesting that the dis2+ and dis3+ genes may be functionally overlapped. The role of the dis3+ gene product in mitosis is unknown, but this gene product may be directly or indirectly involved in the regulation of mitosis. PMID:1944266

Kinoshita, N; Goebl, M; Yanagida, M

1991-12-01

63

An Epimerase Gene Essential for Capsule Synthesis in Vibrio vulnificus  

Microsoft Academic Search

The extracellular capsule polysaccharide (CPS) of Vibrio vulnificus is a primary virulence factor which allows survival of the bacteria in the human host. To study the genes involved in expression of the capsule, we generated mutants that lost the ability to produce CPS following the insertion of a minitransposon into the genome of an encapsulated, clinical strain of V. vulnificus.

AMY B. ZUPPARDO; RONALD J. SIEBELING

1998-01-01

64

The effect of feeding a commercial essential oil product on Clostridium perfringens numbers in the intestine of broiler chickens measured by real-time PCR targeting the ?-toxin-encoding gene ( plc)  

Microsoft Academic Search

Proliferation of Clostridium perfringens type A in the broiler intestinal tract is related to poor growth and litter quality, and can under certain conditions lead to the development of necrotic enteritis (NE), a severe gastrointestinal disease in broilers. The aim of the present study was to investigate the influence of a commercial essential oil blend, CRINA® Poultry, on the intestinal

L. Abildgaard; O. Hojberg; A. Schramm; K. M. Balle; R. M. Engberg

2010-01-01

65

A postgenomic method for predicting essential genes at subsaturation levels of mutagenesis: Application to Mycobacterium tuberculosis  

PubMed Central

We describe a postgenomic in silico approach for identifying genes that are likely to be essential and estimate their proportion in haploid genomes. With the knowledge of all sites eligible for mutagenesis and an experimentally determined partial list of nonessential genes from genome mutagenesis, a Bayesian statistical method provides reasonable predictions of essential genes with a subsaturation level of random mutagenesis. For mutagenesis, a transposon such as Himar1 is suitable as it inserts randomly into TA sites. All of the possible insertion sites may be determined a priori from the genome sequence and with this information, data on experimentally hit TA sites may be used to predict the proportion of genes that cannot be mutated. As a model, we used the Mycobacterium tuberculosis genome. Using the Himar1 transposon, we created a genetically defined collection of 1,425 insertion mutants. Based on our Bayesian statistical analysis using Markov chain Monte Carlo and the observed frequencies of transposon insertions in all of the genes, we estimated that the M. tuberculosis genome contains 35% (95% confidence interval, 28%–41%) essential genes. This analysis further revealed seven functional groups with high probabilities of being enriched in essential genes. The PE-PGRS (Pro-Glu polymorphic GC-rich repetitive sequence) family of genes, which are unique to mycobacteria, the polyketide/nonribosomal peptide synthase family, and mycolic and fatty acid biosynthesis gene families were disproportionately enriched in essential genes. At subsaturation levels of mutagenesis with a random transposon such as Himar1, this approach permits a statistical prediction of both the proportion and identities of essential genes of sequenced genomes.

Lamichhane, Gyanu; Zignol, Matteo; Blades, Natalie J.; Geiman, Deborah E.; Dougherty, Annette; Grosset, Jacques; Broman, Karl W.; Bishai, William R.

2003-01-01

66

The effects of protein interactions, gene essentiality and regulatory regions on expression variation  

PubMed Central

Background Identifying factors affecting gene expression variation is a challenging problem in genetics. Previous studies have shown that the presence of TATA box, the number of cis-regulatory elements, gene essentiality, and protein interactions significantly affect gene expression variation. Nonetheless, the need to obtain a more complete understanding of such factors and how their interactions influence gene expression variation remains a challenge. The growth rates of yeast cells under several DNA-damaging conditions have been studied and a gene's toxicity degree is defined as the number of such conditions that the growth rate of the yeast deletion strain is significantly affected. Since toxicity degree reflects a gene's importance to cell survival under DNA-damaging conditions, we expect that it is negatively associated with gene expression variation. Mutations in both cis-regulatory elements and transcription factors (TF) regulating a gene affect the gene's expression and thus we study the relationship between gene expression variation and the number of TFs regulating a gene. Most importantly we study how these factors interact with each other influencing gene expression variation. Results Using yeast as a model system, we evaluated the effects of four separate factors and their interactions on gene expression variation: protein interaction degree, toxicity degree, number of TFs, and the presence of TATA box. Results showed that 1) gene expression variation is negatively correlated with the protein interaction degree in the protein interaction network, 2) essential genes tend to have less expression variation than non-essential genes and gene expression variation decreases with toxicity degree, and 3) the number of TFs regulating a gene is the most important factor influencing gene expression variation (R2 = 8–14%). In addition, the number of TFs regulating a gene was found to be an important factor influencing gene expression variation for both TATA-containing and non-TATA-containing genes, but with different association strength. Moreover, gene expression variation was significantly negatively correlated with toxicity degree only for TATA-containing genes. Conclusion The finding that distinct mechanisms may influence gene expression variation in TATA-containing and non-TATA-containing genes, provides new insights into the mechanisms that underlie the evolution of gene expression.

Zhou, Linqi; Ma, Xiaotu; Sun, Fengzhu

2008-01-01

67

Expression of Essential B Cell Development Genes in Horses with Common Variable Immunodeficiency  

PubMed Central

Common variable immunodeficiency (CVID) is a heterogeneous disorder of B cell differentiation or function with inadequate antibody production. Our laboratory studies a natural form of CVID in horses characterized by late-onset B cell lymphopenia due to impaired B cell production in the bone marrow. This study was undertaken to assess the status of B cell differentiation in the bone marrow of CVID-affected horses by measuring the expression of genes essential for early B cell commitment and development. Standard RT-PCR revealed that most of the transcription factors and key signaling molecules that directly regulate B cell differentiation in the bone marrow and precede PAX5 are expressed in the affected horses. Yet, the expression of PAX5 and relevant target genes was variable. Quantitative RT-PCR analysis confirmed that the mRNA expression of E2A, PAX5, CD19, and IGHD was significantly reduced in equine CVID patients when compared to healthy horses (p < 0.05). In addition, the PAX5/EBF1 and PAX5/B220 ratios were significantly reduced in CVID patients (p < 0.01). Immunohistochemical analysis confirmed the absence of PAX5-BSAP expression in the bone marrow of affected horses. Our data suggest that B cell development seems to be impaired at the transition between pre-pro-B cells and pro-B cells in equine CVID patients.

Tallmadge, R.L.; Such, K.A.; Miller, K.C.; Matychak, M.B.; Felippe, M.J.B.

2012-01-01

68

EXPRESSION OF A DNA REPLICATION GENE CLUSTER IN BACTERIOPHAGE T4: GENETIC LINKAGE AND THE CONTROL OF GENE PRODUCT INTERACTIONS  

Microsoft Academic Search

The results of this study bear on the relationship between genetic linkage and control of interactions between the protein products of different cistrons. In T4 bacteriophage, genes 45 and 44 encode essential components of the phage DNA replication multiprotein complex. T4 gene 45 maps directly up stream of gene 44 relative to the overall direction of reading of this region

W. L. GERALD; J. D. KARAM

69

Comparative Genomics of Mycoplasma: Analysis of Conserved Essential Genes and Diversity of the Pan-Genome  

Microsoft Academic Search

Mycoplasma, the smallest self-replicating organism with a minimal metabolism and little genomic redundancy, is expected to be a close approximation to the minimal set of genes needed to sustain bacterial life. This study employs comparative evolutionary analysis of twenty Mycoplasma genomes to gain an improved understanding of essential genes. By analyzing the core genome of mycoplasmas, we finally revealed the

Wei Liu; Liurong Fang; Mao Li; Sha Li; Shaohua Guo; Rui Luo; Zhixin Feng; Bin Li; Zhemin Zhou; Guoqing Shao; Huanchun Chen; Shaobo Xiao

2012-01-01

70

A genome-wide strategy for the identification of essential genes in Staphylococcus aureus  

Microsoft Academic Search

Summary To address the need for new approaches to antibiotic drug development, we have identified a large number of essential genes for the bacterial pathogen, Staphy- lococcus aureus, using a rapid shotgun antisense RNA method. Staphylococcus aureus chromosomal DNA fragments were cloned into a xylose-inducible expression plasmid and transformed into S. aureus. Homology comparisons between 658 S. aureus genes identified

R. Allyn Forsyth; Robert J. Haselbeck; Kari L. Ohlsen; Robert T. Yamamoto; Howard Xu; John D. Trawick; Daniel Wall; Liangsu Wang; Vickie Brown-Driver; Jamie M. Froelich; Paula King; Melissa McCarthy; Cheryl Malone; Brian Misiner; David Robbins; Zehui Tan; Zhan-yang Zhu; Grant Carr; Deborah A. Mosca; Carlos Zamudio; J. Gordon Foulkes; Judith W. Zyskind

2002-01-01

71

From essential to persistent genes: a functional approach to constructing synthetic life  

PubMed Central

A central undertaking in synthetic biology (SB) is the quest for the ‘minimal genome’. However, ‘minimal sets’ of essential genes are strongly context-dependent and, in all prokaryotic genomes sequenced to date, not a single protein-coding gene is entirely conserved. Furthermore, a lack of consensus in the field as to what attributes make a gene truly essential adds another aspect of variation. Thus, a universal minimal genome remains elusive. Here, as an alternative to defining a minimal genome, we propose that the concept of gene persistence can be used to classify genes needed for robust long-term survival. Persistent genes, although not ubiquitous, are conserved in a majority of genomes, tend to be expressed at high levels, and are frequently located on the leading DNA strand. These criteria impose constraints on genome organization, and these are important considerations for engineering cells and for creating cellular life-like forms in SB.

Acevedo-Rocha, Carlos G.; Fang, Gang; Schmidt, Markus; Ussery, David W.; Danchin, Antoine

2013-01-01

72

Comprehensive identification of essential Staphylococcus aureus genes using Transposon-Mediated Differential Hybridisation (TMDH)  

PubMed Central

Background In recent years there has been an increasing problem with Staphylococcus aureus strains that are resistant to treatment with existing antibiotics. An important starting point for the development of new antimicrobial drugs is the identification of "essential" genes that are important for bacterial survival and growth. Results We have developed a robust microarray and PCR-based method, Transposon-Mediated Differential Hybridisation (TMDH), that uses novel bioinformatics to identify transposon inserts in genome-wide libraries. Following a microarray-based screen, genes lacking transposon inserts are re-tested using a PCR and sequencing-based approach. We carried out a TMDH analysis of the S. aureus genome using a large random mariner transposon library of around a million mutants, and identified a total of 351 S. aureus genes important for survival and growth in culture. A comparison with the essential gene list experimentally derived for Bacillus subtilis highlighted interesting differences in both pathways and individual genes. Conclusion We have determined the first comprehensive list of S. aureus essential genes. This should act as a useful starting point for the identification of potential targets for novel antimicrobial compounds. The TMDH methodology we have developed is generic and could be applied to identify essential genes in other bacterial pathogens.

Chaudhuri, Roy R; Allen, Andrew G; Owen, Paul J; Shalom, Gil; Stone, Karl; Harrison, Marcus; Burgis, Timothy A; Lockyer, Michael; Garcia-Lara, Jorge; Foster, Simon J; Pleasance, Stephen J; Peters, Sarah E; Maskell, Duncan J; Charles, Ian G

2009-01-01

73

Cloning of hydrogenase genes and fine structure analysis of an operon essential for H2 metabolism in Escherichia coli.  

PubMed Central

Escherichia coli has two unlinked genes that code for hydrogenase synthesis and activity. The DNA fragments containing the two genes (hydA and hydB) were cloned into a plasmid vector, pBR322. The plasmids containing the hyd genes (pSE-290 and pSE-111 carrying the hydA and hydB genes, respectively) were used to genetically map a total of 51 mutant strains with defects in hydrogenase activity. A total of 37 mutants carried a mutation in the hydB gene, whereas the remaining 14 hyd were hydA. This complementation analysis also established the presence of two new genes, so far unidentified, one coding for formate dehydrogenase-2 (fdv) and another producing an electron transport protein (fhl) coupling formate dehydrogenase-2 to hydrogenase. Three of the four genes, hydB, fhl, and fdv, may constitute a single operon, and all three genes are carried by a 5.6-kilobase-pair chromosomal DNA insert in plasmid pSE-128. Plasmids carrying a part of this 5.6-kilobase-pair DNA (pSE-130) or fragments derived from this DNA in different orientations (pSE-126 and pSE-129) inhibited the production of active formate hydrogenlyase. This inhibition occurred even in a prototrophic E. coli, strain K-10, but only during an early induction period. These results, based on complementation analysis with cloned DNA fragments, show that both hydA and hydB genes are essential for the production of active hydrogenase. For the expression of active formate hydrogenlyase, two other gene products, fhl and fdv are also needed. All four genes map between 58 and 59 min in the E. coli chromosome.

Sankar, P; Lee, J H; Shanmugam, K T

1985-01-01

74

Identifying essential genes in bacterial metabolic networks with machine learning methods  

PubMed Central

Background Identifying essential genes in bacteria supports to identify potential drug targets and an understanding of minimal requirements for a synthetic cell. However, experimentally assaying the essentiality of their coding genes is resource intensive and not feasible for all bacterial organisms, in particular if they are infective. Results We developed a machine learning technique to identify essential genes using the experimental data of genome-wide knock-out screens from one bacterial organism to infer essential genes of another related bacterial organism. We used a broad variety of topological features, sequence characteristics and co-expression properties potentially associated with essentiality, such as flux deviations, centrality, codon frequencies of the sequences, co-regulation and phyletic retention. An organism-wise cross-validation on bacterial species yielded reliable results with good accuracies (area under the receiver-operator-curve of 75% - 81%). Finally, it was applied to drug target predictions for Salmonella typhimurium. We compared our predictions to the viability of experimental knock-outs of S. typhimurium and identified 35 enzymes, which are highly relevant to be considered as potential drug targets. Specifically, we detected promising drug targets in the non-mevalonate pathway. Conclusions Using elaborated features characterizing network topology, sequence information and microarray data enables to predict essential genes from a bacterial reference organism to a related query organism without any knowledge about the essentiality of genes of the query organism. In general, such a method is beneficial for inferring drug targets when experimental data about genome-wide knockout screens is not available for the investigated organism.

2010-01-01

75

Geptop: A Gene Essentiality Prediction Tool for Sequenced Bacterial Genomes Based on Orthology and Phylogeny  

PubMed Central

Integrative genomics predictors, which score highly in predicting bacterial essential genes, would be unfeasible in most species because the data sources are limited. We developed a universal approach and tool designated Geptop, based on orthology and phylogeny, to offer gene essentiality annotations. In a series of tests, our Geptop method yielded higher area under curve (AUC) scores in the receiver operating curves than the integrative approaches. In the ten-fold cross-validations among randomly upset samples, Geptop yielded an AUC of 0.918, and in the cross-organism predictions for 19 organisms Geptop yielded AUC scores between 0.569 and 0.959. A test applied to the very recently determined essential gene dataset from the Porphyromonas gingivalis, which belongs to a phylum different with all of the above 19 bacterial genomes, gave an AUC of 0.77. Therefore, Geptop can be applied to any bacterial species whose genome has been sequenced. Compared with the essential genes uniquely identified by the lethal screening, the essential genes predicted only by Gepop are associated with more protein-protein interactions, especially in the three bacteria with lower AUC scores (<0.7). This may further illustrate the reliability and feasibility of our method in some sense. The web server and standalone version of Geptop are available at http://cefg.uestc.edu.cn/geptop/ free of charge. The tool has been run on 968 bacterial genomes and the results are accessible at the website.

Wei, Wen; Ning, Lu-Wen; Ye, Yuan-Nong; Guo, Feng-Biao

2013-01-01

76

The Yield of Essential Oils in Melaleuca alternifolia (Myrtaceae) Is Regulated through Transcript Abundance of Genes in the MEP Pathway  

PubMed Central

Medicinal tea tree (Melaleuca alternifolia) leaves contain large amounts of an essential oil, dominated by monoterpenes. Several enzymes of the chloroplastic methylerythritol phosphate (MEP) pathway are hypothesised to act as bottlenecks to the production of monoterpenes. We investigated, whether transcript abundance of genes encoding for enzymes of the MEP pathway were correlated with foliar terpenes in M. alternifolia using a population of 48 individuals that ranged in their oil concentration from 39 -122 mg.g DM?1. Our study shows that most genes in the MEP pathway are co-regulated and that the expression of multiple genes within the MEP pathway is correlated with oil yield. Using multiple regression analysis, variation in expression of MEP pathway genes explained 87% of variation in foliar monoterpene concentrations. The data also suggest that sesquiterpenes in M. alternifolia are synthesised, at least in part, from isopentenyl pyrophosphate originating from the plastid via the MEP pathway.

Webb, Hamish; Lanfear, Robert; Hamill, John; Foley, William J.; Kulheim, Carsten

2013-01-01

77

A Survey of Essential Gene Function in the Yeast Cell Division Cycle  

PubMed Central

Mutations impacting specific stages of cell growth and division have provided a foundation for dissecting mechanisms that underlie cell cycle progression. We have undertaken an objective examination of the yeast cell cycle through flow cytometric analysis of DNA content in TetO7 promoter mutant strains representing 75% of all essential yeast genes. More than 65% of the strains displayed specific alterations in DNA content, suggesting that reduced function of an essential gene in most cases impairs progression through a specific stage of the cell cycle. Because of the large number of essential genes required for protein biosynthesis, G1 accumulation was the most common phenotype observed in our analysis. In contrast, relatively few mutants displayed S-phase delay, and most of these were defective in genes required for DNA replication or nucleotide metabolism. G2 accumulation appeared to arise from a variety of defects. In addition to providing a global view of the diversity of essential cellular processes that influence cell cycle progression, these data also provided predictions regarding the functions of individual genes: we identified four new genes involved in protein trafficking (NUS1, PHS1, PGA2, PGA3), and we found that CSE1 and SMC4 are important for DNA replication.

Yu, Lisa; Castillo, Lourdes Pena; Mnaimneh, Sanie

2006-01-01

78

The Snail Family Gene Snai3 Is Not Essential for Embryogenesis in Mice  

PubMed Central

The Snail gene family encodes zinc finger-containing transcriptional repressor proteins. Three members of the Snail gene family have been described in mammals, encoded by the Snai1, Snai2, and Snai3 genes. The function of the Snai1 and Snai2 genes have been studied extensively during both vertebrate embryogenesis and tumor progression and metastasis, and play critically important roles during these processes. However, little is known about the function of the Snai3 gene and protein. We describe here generation and analysis of Snai3 conditional and null mutant mice. We also generated an EYFP-tagged Snai3 null allele that accurately reflects endogenous Snai3 gene expression, with the highest levels of expression detected in thymus and skeletal muscle. Snai3 null mutant homozygous mice are viable and fertile, and exhibit no obvious phenotypic defects. These results demonstrate that Snai3 gene function is not essential for embryogenesis in mice.

Han, Xianghua; Booth, Carmen J.; Yoon, Jeong Kyo; Krebs, Luke T.; Gridley, Thomas

2013-01-01

79

Food production & availability - Essential prerequisites for sustainable food security.  

PubMed

Food and nutrition security are intimately interconnected, since only a food based approach can help in overcoming malnutrition in an economically and socially sustainable manner. Food production provides the base for food security as it is a key determinant of food availability. This paper deals with different aspects of ensuring high productivity and production without associated ecological harm for ensuring adequate food availability. By mainstreaming ecological considerations in technology development and dissemination, we can enter an era of evergreen revolution and sustainable food and nutrition security. Public policy support is crucial for enabling this. PMID:24135188

Swaminathan, M S; Bhavani, R V

2013-09-01

80

ORF18 Is a Transfactor That Is Essential for Late Gene Transcription of a Gammaherpesvirus  

PubMed Central

Lytic replication of the tumor-associated human gammaherpesviruses Epstein-Barr virus and Kaposi's sarcoma-associated herpesvirus has important implications in pathogenesis and tumorigenesis. Herpesvirus lytic genes have been temporally classified as exhibiting immediate-early (IE), early, and late expression kinetics. Though the regulation of IE and early gene expression has been studied extensively, very little is known regarding the regulation of late gene expression. Late genes, which primarily encode virion structural proteins, require viral DNA replication for their expression. We have identified a murine gammaherpesvirus 68 (MHV-68) early lytic gene, ORF18, essential for viral replication. ORF18 is conserved in both beta- and gammaherpesviruses. By generating an MHV-68 ORF18-null virus, we characterized the stage of the virus lytic cascade that requires the function of ORF18. Gene expression profiling and quantitation of viral DNA synthesis of the ORF18-null virus revealed that the expression of early genes and viral DNA replication were not affected; however, the transcription of late genes was abolished. Hence, we have identified a gammaherpesvirus-encoded factor essential for the expression of late genes independently of viral DNA synthesis.

Arumugaswami, Vaithilingaraja; Wu, Ting-Ting; Martinez-Guzman, DeeAnn; Jia, Qingmei; Deng, Hongyu; Reyes, Nichole; Sun, Ren

2006-01-01

81

Construction of a Bacterial Cell that Contains Only the Set of Essential Genes Necessary to Impart Life.  

National Technical Information Service (NTIS)

Work has continued on the Top Down approach to genome minimization. We have used the Essential (E), Non-essential (N), and Impaired (I) gene categories to make steady progress with gene and gene cluster deletions. To date, we have removed approximately 23...

A. Yee C. Hutchison H. Smith J. Glass

2013-01-01

82

Human herpesvirus 6 encoded glycoprotein Q1 gene is essential for virus growth  

Microsoft Academic Search

Human herpesvirus 6 (HHV-6) glycoprotein Q1 (gQ1), a unique gene in HHV-6, forms a complex with glycoproteinH (gH) and gL, which is the viral ligand for its cellular receptor, CD46. However, whether gQ1 is essential for virus growth is unknown, because a system is lacking for making gene knockouts for HHV-6. Recently, bacterial artificial chromosome (BAC) and E. coli mutagenesis

Huamin Tang; Akiko Kawabata; Mayumi Yoshida; Hiroko Oyaizu; Takahiro Maeki; Koichi Yamanishi; Yasuko Mori

2010-01-01

83

A genome-wide screen for essential yeast genes that affect telomere length maintenance  

PubMed Central

Telomeres are structures composed of repetitive DNA and proteins that protect the chromosomal ends in eukaryotic cells from fusion or degradation, thus contributing to genomic stability. Although telomere length varies between species, in all organisms studied telomere length appears to be controlled by a dynamic equilibrium between elongating mechanisms (mainly addition of repeats by the enzyme telomerase) and nucleases that shorten the telomeric sequences. Two previous studies have analyzed a collection of yeast deletion strains (deleted for nonessential genes) and found over 270 genes that affect telomere length (Telomere Length Maintenance or TLM genes). Here we complete the list of TLM by analyzing a collection of strains carrying hypomorphic alleles of most essential genes (DAmP collection). We identify 87 essential genes that affect telomere length in yeast. These genes interact with the nonessential TLM genes in a significant manner, and provide new insights on the mechanisms involved in telomere length maintenance. The newly identified genes span a variety of cellular processes, including protein degradation, pre-mRNA splicing and DNA replication.

Ungar, Lior; Yosef, Nir; Sela, Yael; Sharan, Roded; Ruppin, Eytan; Kupiec, Martin

2009-01-01

84

Identification of Linked Legionella pneumophila Genes Essential for Intracellular Growth and Evasion of the Endocytic Pathway  

Microsoft Academic Search

Legionella pneumophila replicates within a specialized phagosome in cultured cells, a function necessary for its pathogenicity. The replicative phagosome lacks membrane marker proteins, such as the glycoprotein LAMP-1, that are indicators of the normal endocytic pathway. We describe the isolation of several Legionella genes essential for intracellular growth and evasion of the endocytic pathway, using a genetic and cell biological

HELENE L. ANDREWS; JOSEPH P. VOGEL; RALPH R. ISBERG

85

Autographa californica multiple nucleopolyhedrovirus odv-e66 is an essential gene required for oral infectivity.  

PubMed

Autographa californica multiple nucleopolyhedrovirus (AcMNPV) odv-e66 is a core gene and encodes an occlusion-derived virus (ODV)-specific envelope protein, ODV-E66. The N-terminal 23 amino acid of the envelope protein ODV-E66 are sufficient to direct native and fusion proteins to induced membrane microvesicles and the viral envelope during infection with AcMNPV. In this study, an odv-e66-knockout bacmid can not express N-terminal hydrophobic domains was constructed via homologous recombination in Escherichia coli. The odv-e66 deletion had no effect on budded virus (BV) production and viral DNA replication in infected Sf9 cells. Larval bioassays demonstrated that injection of odv-e66 deletion BV into the hemocoel could kill P. xylostella larvae as efficiently as repaired and control viruses; however, odv-e66 deletion mutant resulted in a 50% lethal dose that was 10(3) higher than that of the repaired and control viruses when inoculated per os. These results indicated that ODV-E66 envelope protein most likely played an important role in the oral infectivity of AcMNPV, but is not essential for virus replication. PMID:21440017

Xiang, Xingwei; Chen, Lin; Hu, Xiaolong; Yu, Shaofang; Yang, Rui; Wu, Xiaofeng

2011-03-31

86

Identification of essential genes in the human fungal pathogen Aspergillus fumigatus by transposon mutagenesis.  

PubMed

The opportunistic pathogen Aspergillus fumigatus is the most frequent cause of deadly airborne fungal infections in developed countries. In order to identify novel antifungal-drug targets, we investigated the genome of A. fumigatus for genes that are necessary for efficient fungal growth. An artificial A. fumigatus diploid strain with one copy of an engineered impala160 transposon from Fusarium oxysporum integrated into its genome was used to generate a library of diploid strains by random in vivo transposon mutagenesis. Among 2,386 heterozygous diploid strains screened by parasexual genetics, 1.2% had a copy of the transposable element integrated into a locus essential for A. fumigatus growth. Comparison of genomic sequences flanking impala160 in these mutants with that of the genome of A. fumigatus allowed the characterization of 20 previously uncharacterized A. fumigatus genes. Among these, homologues of genes essential for Saccharomyces cerevisiae growth have been identified, as well as genes that do not have homologues in other fungal species. These results confirm that heterologous transposition using the transposable element impala is a powerful tool for functional genomics in ascomycota, and they pave the way for defining the complete set of essential genes in A. fumigatus, the first step toward target-based development of new antifungal drugs. PMID:12684374

Firon, Arnaud; Villalba, François; Beffa, Roland; D'Enfert, Christophe

2003-04-01

87

Functional requirements for bacteriophage growth: gene essentiality and expression in mycobacteriophage Giles.  

PubMed

Bacteriophages represent a majority of all life forms, and the vast, dynamic population with early origins is reflected in their enormous genetic diversity. A large number of bacteriophage genomes have been sequenced. They are replete with novel genes without known relatives. We know little about their functions, which genes are required for lytic growth, and how they are expressed. Furthermore, the diversity is such that even genes with required functions - such as virion proteins and repressors - cannot always be recognized. Here we describe a functional genomic dissection of mycobacteriophage Giles, in which the virion proteins are identified, genes required for lytic growth are determined, the repressor is identified, and the transcription patterns determined. We find that although all of the predicted phage genes are expressed either in lysogeny or in lytic growth, 45% of the predicted genes are non-essential for lytic growth. We also describe genes required for DNA replication, show that recombination is required for lytic growth, and that Giles encodes a novel repressor. RNAseq analysis reveals abundant expression of a small non-coding RNA in a lysogen and in late lytic growth, although it is non-essential for lytic growth and does not alter lysogeny. PMID:23560716

Dedrick, Rebekah M; Marinelli, Laura J; Newton, Gerald L; Pogliano, Kit; Pogliano, Joseph; Hatfull, Graham F

2013-04-08

88

whmD is an essential mycobacterial gene required for proper septation and cell division  

PubMed Central

A study of potential mycobacterial regulatory genes led to the isolation of the Mycobacterium smegmatis whmD gene, which encodes a homologue of WhiB, a Streptomyces coelicolor protein required for sporulation. Unlike its Streptomyces homologue, WhmD is essential in M. smegmatis. The whmD gene could be disrupted only in the presence of a plasmid supplying whmD in trans. A plasmid that allowed chemically regulated expression of the WhmD protein was used to generate a conditional whmD mutant. On withdrawal of the inducer, the conditional whmD mutant exhibited irreversible, filamentous, branched growth with diminished septum formation and aberrant septal placement, whereas WhmD overexpression resulted in growth retardation and hyperseptation. Nucleic acid synthesis and levels of the essential cell division protein FtsZ were unaltered by WhmD deficiency. Together, these phenotypes indicate a role for WhmD in mycobacterial septum formation and cell division.

Gomez, James E.; Bishai, William R.

2000-01-01

89

Construction and complementation of in-frame deletions of the essential Escherichia coli thymidylate kinase gene.  

PubMed

This work reports the construction of Escherichia coli in-frame deletion strains of tmk, which encodes thymidylate kinase, Tmk. The tmk gene is located at the third position of a putative five-gene operon at 24.9 min on the E. coli chromosome, which comprises the genes pabC, yceG, tmk, holB, and ycfH. To avoid potential polar effects on downstream genes of the operon, as well as recombination with plasmid-encoded tmk, the tmk gene was replaced by the kanamycin resistance gene kka1, encoding amino glycoside 3'-phosphotransferase kanamycin kinase. The kanamycin resistance gene is expressed under the control of the natural promoter(s) of the putative operon. The E. coli tmk gene is essential under any conditions tested. To show functional complementation in bacteria, the E. coli tmk gene was replaced by thymidylate kinases of bacteriophage T4 gp1, E. coli tmk, Saccharomyces cerevisiae cdc8, or the Homo sapiens homologue, dTYMK. Growth of these transgenic E. coli strains is completely dependent on thymidylate kinase activities of various origin expressed from plasmids. The substitution constructs show no polar effects on the downstream genes holB and ycfH with respect to cell viability. The presented transgenic bacteria could be of interest for testing of thymidylate kinase-specific phosphorylation of nucleoside analogues that are used in therapies against cancer and infectious diseases. PMID:16461678

Chaperon, David-Nicolas

2006-02-01

90

MAS1, a gene essential for yeast mitochondrial assembly, encodes a subunit of the mitochondrial processing protease.  

PubMed Central

We have previously described a yeast mutant (mas1) that accumulates mitochondrial precursor proteins at high temperature and is deficient in the activity of a matrix-localized protease which cleaves presequences from mitochondrial precursor proteins. We have now cloned and sequenced the wild-type MAS1 gene and found that it encodes a subunit of the mitochondrial processing protease, that it is essential for cell viability and that the protein product participates in its own cleavage during import into mitochondria. The MAS1 protein is thus the first genetically defined component of the mitochondrial protein import pathway. Images

Witte, C; Jensen, R E; Yaffe, M P; Schatz, G

1988-01-01

91

Identifying essential genes in Escherichia coli from a metabolic optimization principle  

PubMed Central

Understanding the organization of reaction fluxes in cellular metabolism from the stoichiometry and the topology of the underlying biochemical network is a central issue in systems biology. In this task, it is important to devise reasonable approximation schemes that rely on the stoichiometric data only, because full-scale kinetic approaches are computationally affordable only for small networks (e.g., red blood cells, ?50 reactions). Methods commonly used are based on finding the stationary flux configurations that satisfy mass-balance conditions for metabolites, often coupling them to local optimization rules (e.g., maximization of biomass production) to reduce the size of the solution space to a single point. Such methods have been widely applied and have proven able to reproduce experimental findings for relatively simple organisms in specific conditions. Here, we define and study a constraint-based model of cellular metabolism where neither mass balance nor flux stationarity are postulated and where the relevant flux configurations optimize the global growth of the system. In the case of Escherichia coli, steady flux states are recovered as solutions, although mass-balance conditions are violated for some metabolites, implying a nonzero net production of the latter. Such solutions furthermore turn out to provide the correct statistics of fluxes for the bacterium E. coli in different environments and compare well with the available experimental evidence on individual fluxes. Conserved metabolic pools play a key role in determining growth rate and flux variability. Finally, we are able to connect phenomenological gene essentiality with “frozen” fluxes (i.e., fluxes with smaller allowed variability) in E. coli metabolism.

Martelli, Carlotta; De Martino, Andrea; Marinari, Enzo; Marsili, Matteo; Perez Castillo, Isaac

2009-01-01

92

Analysis of essential Arabidopsis nuclear genes encoding plastid-targeted proteins.  

PubMed

The Chloroplast 2010 Project (http://www.plastid.msu.edu/) identified and phenotypically characterized homozygous mutants in over three thousand genes, the majority of which encode plastid-targeted proteins. Despite extensive screening by the community, no homozygous mutant alleles were available for several hundred genes, suggesting that these might be enriched for genes of essential function. Attempts were made to generate homozygotes in ~1200 of these lines and 521 of the homozygous viable lines obtained were deposited in the Arabidopsis Biological Resource Center (http://abrc.osu.edu/). Lines that did not yield a homozygote in soil were tested as potentially homozygous lethal due to defects either in seed or seedling development. Mutants were characterized at four stages of development: developing seed, mature seed, at germination, and developing seedlings. To distinguish seed development or seed pigment-defective mutants from seedling development mutants, development of seeds was assayed in siliques from heterozygous plants. Segregating seeds from heterozygous parents were sown on supplemented media in an attempt to rescue homozygous seedlings that could not germinate or survive in soil. Growth of segregating seeds in air and air enriched to 0.3% carbon dioxide was compared to discover mutants potentially impaired in photorespiration or otherwise responsive to CO2 supplementation. Chlorophyll fluorescence measurements identified CO2-responsive mutants with altered photosynthetic parameters. Examples of genes with a viable mutant allele and one or more putative homozygous-lethal alleles were documented. RT-PCR of homozygotes for potentially weak alleles revealed that essential genes may remain undiscovered because of the lack of a true null mutant allele. This work revealed 33 genes with two or more lethal alleles and 73 genes whose essentiality was not confirmed with an independent lethal mutation, although in some cases second leaky alleles were identified. PMID:24023856

Savage, Linda J; Imre, Kathleen M; Hall, David A; Last, Robert L

2013-09-04

93

A Novel Gene Essential for the Development of Single Positive Thymocytes?  

PubMed Central

A critical step during intrathymic T-cell development is the transition of CD4+ CD8+ double-positive (DP) cells to the major histocompatibility complex class I (MHC-I)-restricted CD4? CD8+ and MHC-II-restricted CD4+ CD8? single-positive (SP) cell stage. Here, we identify a novel gene that is essential for this process. Through the T-cell phenotype-based screening of N-ethyl-N-nitrosourea (ENU)-induced mutant mice, we established a mouse line in which numbers of CD4 and CD8 SP thymocytes as well as peripheral CD4 and CD8 T cells were dramatically reduced. Using linkage analysis and DNA sequencing, we identified a missense point mutation in a gene, E430004N04Rik (also known as themis), that does not belong to any known gene family. This orphan gene is expressed specifically in DP and SP thymocytes and peripheral T cells, whereas in mutant thymocytes the levels of protein encoded by this gene were drastically reduced. We generated E430004N04Rik-deficient mice, and their phenotype was virtually identical to that of the ENU mutant mice, thereby confirming that this gene is essential for the development of SP thymocytes.

Kakugawa , Kiyokazu; Yasuda, Takuwa; Miura, Ikuo; Kobayashi, Ayako; Fukiage, Hitomi; Satoh, Rumi; Matsuda, Masashi; Koseki, Haruhiko; Wakana, Shigeharu; Kawamoto, Hiroshi; Yoshida, Hisahiro

2009-01-01

94

The Wilms' Tumor Suppressor Gene (wt1) Product Regulates Dax-1 Gene Expression during Gonadal Differentiation  

Microsoft Academic Search

Gonadal differentiation is dependent upon a molecular cascade responsible for ovarian or testicular develop- ment from the bipotential gonadal ridge. Genetic analysis has implicated a number of gene products essential for this process, which include Sry, WT1, SF-1, and DAX-1. We have sought to better define the role of WT1 in this process by identifying downstream targets of WT1 during

JUNGHO KIM; DIRK PRAWITT; NABEEL BARDEESY; ELENA TORBAN; CAROLINE VICANER; PAUL GOODYER; BERNARD ZABEL; JERRY PELLETIER

95

The ispB gene encoding octaprenyl diphosphate synthase is essential for growth of Escherichia coli.  

PubMed

The Escherichia coli ispB gene encoding octaprenyl diphosphate synthase is responsible for the synthesis of the side chain of isoprenoid quinones. We tried to construct an E. coli ispB-disrupted mutant but could not isolate the chromosomal ispB disrupted mutant unless the ispB gene or its homolog was supplied on a plasmid. The chromosomal ispB disruptants that harbored plasmids carrying the ispB homologs from Haemophilus influenzae and Synechocystis sp. strain PCC6803 produced mainly ubiquinone 7 and ubiquinone 9, respectively. Our results indicate that the function of the ispB gene is essential for normal growth and that this function can be substituted for by homologs of the ispB gene from other organisms that produce distinct forms of ubiquinone. PMID:9139929

Okada, K; Minehira, M; Zhu, X; Suzuki, K; Nakagawa, T; Matsuda, H; Kawamukai, M

1997-05-01

96

The ispB gene encoding octaprenyl diphosphate synthase is essential for growth of Escherichia coli.  

PubMed Central

The Escherichia coli ispB gene encoding octaprenyl diphosphate synthase is responsible for the synthesis of the side chain of isoprenoid quinones. We tried to construct an E. coli ispB-disrupted mutant but could not isolate the chromosomal ispB disrupted mutant unless the ispB gene or its homolog was supplied on a plasmid. The chromosomal ispB disruptants that harbored plasmids carrying the ispB homologs from Haemophilus influenzae and Synechocystis sp. strain PCC6803 produced mainly ubiquinone 7 and ubiquinone 9, respectively. Our results indicate that the function of the ispB gene is essential for normal growth and that this function can be substituted for by homologs of the ispB gene from other organisms that produce distinct forms of ubiquinone.

Okada, K; Minehira, M; Zhu, X; Suzuki, K; Nakagawa, T; Matsuda, H; Kawamukai, M

1997-01-01

97

Genome-scale gene/reaction essentiality and synthetic lethality analysis  

PubMed Central

Synthetic lethals are to pairs of non-essential genes whose simultaneous deletion prohibits growth. One can extend the concept of synthetic lethality by considering gene groups of increasing size where only the simultaneous elimination of all genes is lethal, whereas individual gene deletions are not. We developed optimization-based procedures for the exhaustive and targeted enumeration of multi-gene (and by extension multi-reaction) lethals for genome-scale metabolic models. Specifically, these approaches are applied to iAF1260, the latest model of Escherichia coli, leading to the complete identification of all double and triple gene and reaction synthetic lethals as well as the targeted identification of quadruples and some higher-order ones. Graph representations of these synthetic lethals reveal a variety of motifs ranging from hub-like to highly connected subgraphs providing a birds-eye view of the avenues available for redirecting metabolism and uncovering complex patterns of gene utilization and interdependence. The procedure also enables the use of falsely predicted synthetic lethals for metabolic model curation. By analyzing the functional classifications of the genes involved in synthetic lethals, we reveal surprising connections within and across clusters of orthologous group functional classifications.

Suthers, Patrick F; Zomorrodi, Alireza; Maranas, Costas D

2009-01-01

98

The NGATHA Distal Organ Development Genes Are Essential for Style Specification in Arabidopsis[W  

PubMed Central

Floral organ identities are specified by a few transcription factors that act as master regulators. Subsequently, specification of organ axes programs the distribution of distinct tissue types within the organs that themselves develop unique identities. The C-class, AGAMOUS-clade MADS box genes are primary promoters of the gynoecium, which is divided into a distal style and a subtending ovary along the apical-basal axis. We show that members of a clade of B3 domain transcription factors, NGATHA1 (NGA1) to NGA4, are expressed distally in all lateral organs, and all four have a redundant and essential role in style development. Loss of all four genes results in gynoecia where style is replaced by valve-like projections and a reduction in style-specific SHATTERPROOF1 (SHP1) expression. In agreement, floral misexpression of NGA1 promotes ectopic style and SHP1 expression. STYLISH1, an auxin biosynthesis inducer, conditionally activated NGA genes, which in turn promoted distal expression of other STY genes in a putative positive feedback loop. Inhibited auxin transport or lack of YABBY1 gene activities resulted in a basally expanded style domain and broader expression of NGA genes. We speculate that early gynoecium factors delimit NGA gene response to an auxin-based signal, elicited by STY gene activity, to restrict the activation of style program to a late and distal carpel domain.

Alvarez, John Paul; Goldshmidt, Alexander; Efroni, Idan; Bowman, John L.; Eshed, Yuval

2009-01-01

99

Arabidopsis genes essential for seedling viability: isolation of insertional mutants and molecular cloning.  

PubMed Central

We have undertaken a large-scale genetic screen to identify genes with a seedling-lethal mutant phenotype. From screening approximately 38,000 insertional mutant lines, we identified >500 seedling-lethal mutants, completed cosegregation analysis of the insertion and the lethal phenotype for >200 mutants, molecularly characterized 54 mutants, and provided a detailed description for 22 of them. Most of the seedling-lethal mutants seem to affect chloroplast function because they display altered pigmentation and affect genes encoding proteins predicted to have chloroplast localization. Although a high level of functional redundancy in Arabidopsis might be expected because 65% of genes are members of gene families, we found that 41% of the essential genes found in this study are members of Arabidopsis gene families. In addition, we isolated several interesting classes of mutants and genes. We found three mutants in the recently discovered nonmevalonate isoprenoid biosynthetic pathway and mutants disrupting genes similar to Tic40 and tatC, which are likely to be involved in chloroplast protein translocation. Finally, we directly compared T-DNA and Ac/Ds transposon mutagenesis methods in Arabidopsis on a genome scale. In each population, we found only about one-third of the insertion mutations cosegregated with a mutant phenotype.

Budziszewski, G J; Lewis, S P; Glover, L W; Reineke, J; Jones, G; Ziemnik, L S; Lonowski, J; Nyfeler, B; Aux, G; Zhou, Q; McElver, J; Patton, D A; Martienssen, R; Grossniklaus, U; Ma, H; Law, M; Levin, J Z

2001-01-01

100

An essential gene of Saccharomyces cerevisiae coding for an actin-related protein.  

PubMed Central

Actin filaments provide the internal scaffold of eukaryotic cells; they are involved in maintenance of cell shape, cytokinesis, organelle movement, and cell motility. The major component of these filaments, actin, is one of the most well-conserved eukaryotic proteins. Recently genes more distantly related to the conventional actins were cloned from several organisms. In the budding yeast, Saccharomyces cerevisiae, one conventional actin gene, ACT1 (coding for the filament actin), and a so-called actin-like gene, ACT2 (of unknown function), have so far been identified. We report here the discovery of a third member of the actin gene family from this organism, which we named ACT3. The latter gene is essential for viability and codes for a putative polypeptide, Act3, of 489 amino acids (M(r) = 54,831). The deduced amino acid sequence of Act3 is less related to conventional actins than is the deduced amino acid sequence of Act2, mainly because of three unique hydrophilic [corrected] segments. These segments are found inserted into a part of the sequence corresponding to a surface loop of the known three-dimensional structure of the actin molecule. According to sequence comparison, the basal core structure of conventional actin may well be conserved in Act3. Our findings demonstrate that, unexpectedly, there exist three members of the diverse actin protein family in budding yeast that obviously provide different essential functions for survival. Images

Harata, M; Karwan, A; Wintersberger, U

1994-01-01

101

Survival of UV-irradiated mammalian cells correlates with efficient DNA repair in an essential gene  

SciTech Connect

The survival of UV-irradiated mammalian cells is not necessarily correlated with their overall capacity to carry out DNA repair. Human cells typically remove 80% of the pyrimidine dimers produced by a UV dose of 5 J/m2 within 24 hr. In contrast, a Chinese hamster ovary (CHO) cell line survives UV irradiation equally well while removing only 15% of the dimers. Using a newly developed technique to measure dimer frequencies in single-copy specific sequences, we find that the CHO cells remove 70% of the dimers from the essential dihydrofolate reductase (DHFR) gene but only 20% from sequences located 30 kilobases or more upstream from the 5' end of the gene in a 24-hr period. Repair-deficient human cells from xeroderma pigmentosum complementation group C (XPC) are similar to the CHO cells in overall repair levels, but they are extremely sensitive to killing by UV irradiation. In the XPC cells, we find little or no repair in the DHFR gene; in contrast, in normal human fibroblasts and epidermal keratinocytes, greater than 80% of the dimers induced in the gene by 20 J/m2 are removed in 24 hr. Since the CHO and normal human cells exhibit similar UV resistance, much higher than that of XPC cells, our findings suggest a correlation between efficient repair of essential genes and resistance to DNA-damaging agents such as UV light.

Bohr, V.A.; Okumoto, D.S.; Hanawalt, P.C.

1986-06-01

102

The Saccharomyces cerevisiae MYO2 gene encodes an essential myosin for vectorial transport of vesicles  

Microsoft Academic Search

Abstract. After the initiation of bud formation, cells of the yeast Saccharomyces,cerevisiae direct new growth,to the developing,bud. We show,here that this vectorial growth,is facilitated by activity of the MY02 gene. The wild-type MY02 gene encodes,an essential form of myosin,composed,of an NH2-terminal domain typical of the globular, actin-binding domain of other myosins. This NH:-terminal domain,is linked by what appears to be

G. C. Johnston; J. A. Prendergast; R. A. Singer

1991-01-01

103

Viable nonsense mutants for the essential gene SUP45 of Saccharomyces cerevisiae  

Microsoft Academic Search

BACKGROUND: Termination of protein synthesis in eukaryotes involves at least two polypeptide release factors (eRFs) – eRF1 and eRF3. The highly conserved translation termination factor eRF1 in Saccharomyces cerevisiae is encoded by the essential gene SUP45. RESULTS: We have isolated five sup45-n (n from nonsense) mutations that cause nonsense substitutions in the following amino acid positions of eRF1: Y53 ?

Svetlana E Moskalenko; Svetlana V Chabelskaya; Sergei G Inge-Vechtomov; Michel Philippe; Galina A Zhouravleva

2003-01-01

104

The organ-specific expression of terpene synthase genes contributes to the terpene hydrocarbon composition of chamomile essential oils  

PubMed Central

Background The essential oil of chamomile, one of the oldest and agronomically most important medicinal plant species in Europe, has significant antiphlogistic, spasmolytic and antimicrobial activities. It is rich in chamazulene, a pharmaceutically active compound spontaneously formed during steam distillation from the sesquiterpene lactone matricine. Chamomile oil also contains sesquiterpene alcohols and hydrocarbons which are produced by the action of terpene synthases (TPS), the key enzymes in constructing terpene carbon skeletons. Results Here, we present the identification and characterization of five TPS enzymes contributing to terpene biosynthesis in chamomile (Matricaria recutita). Four of these enzymes were exclusively expressed in above-ground organs and produced the common terpene hydrocarbons (?)-(E)-?-caryophyllene (MrTPS1), (+)-germacrene A (MrTPS3), (E)-?-ocimene (MrTPS4) and (?)-germacrene D (MrTPS5). A fifth TPS, the multiproduct enzyme MrTPS2, was mainly expressed in roots and formed several Asteraceae-specific tricyclic sesquiterpenes with (?)-?-isocomene being the major product. The TPS transcript accumulation patterns in different organs of chamomile were consistent with the abundance of the corresponding TPS products isolated from these organs suggesting that the spatial regulation of TPS gene expression qualitatively contribute to terpene composition. Conclusions The terpene synthases characterized in this study are involved in the organ-specific formation of essential oils in chamomile. While the products of MrTPS1, MrTPS2, MrTPS4 and MrTPS5 accumulate in the oils without further chemical alterations, (+)-germacrene A produced by MrTPS3 accumulates only in trace amounts, indicating that it is converted into another compound like matricine. Thus, MrTPS3, but also the other TPS genes, are good markers for further breeding of chamomile cultivars rich in pharmaceutically active essential oils.

2012-01-01

105

In vivo and in silico determination of essential genes of Campylobacter jejuni  

PubMed Central

Background In the United Kingdom, the thermophilic Campylobacter species C. jejuni and C. coli are the most frequent causes of food-borne gastroenteritis in humans. While campylobacteriosis is usually a relatively mild infection, it has a significant public health and economic impact, and possible complications include reactive arthritis and the autoimmune diseases Guillain-Barré syndrome. The rapid developments in "omics" technologies have resulted in the availability of diverse datasets allowing predictions of metabolism and physiology of pathogenic micro-organisms. When combined, these datasets may allow for the identification of potential weaknesses that can be used for development of new antimicrobials to reduce or eliminate C. jejuni and C. coli from the food chain. Results A metabolic model of C. jejuni was constructed using the annotation of the NCTC 11168 genome sequence, a published model of the related bacterium Helicobacter pylori, and extensive literature mining. Using this model, we have used in silico Flux Balance Analysis (FBA) to determine key metabolic routes that are essential for generating energy and biomass, thus creating a list of genes potentially essential for growth under laboratory conditions. To complement this in silico approach, candidate essential genes have been determined using a whole genome transposon mutagenesis method. FBA and transposon mutagenesis (both this study and a published study) predict a similar number of essential genes (around 200). The analysis of the intersection between the three approaches highlights the shikimate pathway where genes are predicted to be essential by one or more method, and tend to be network hubs, based on a previously published Campylobacter protein-protein interaction network, and could therefore be targets for novel antimicrobial therapy. Conclusions We have constructed the first curated metabolic model for the food-borne pathogen Campylobacter jejuni and have presented the resulting metabolic insights. We have shown that the combination of in silico and in vivo approaches could point to non-redundant, indispensable genes associated with the well characterised shikimate pathway, and also genes of unknown function specific to C. jejuni, which are all potential novel Campylobacter intervention targets.

2011-01-01

106

Plasma Hydrogen Peroxide Production in Human Essential Hypertension Role of Heredity, Gender, and Ethnicity  

Microsoft Academic Search

Oxygen free radicals, including hydrogen peroxide, may mediate oxidative stress in target organ tissues and contribute to cardiovascular complications in hypertension. To examine heritability of hydrogen peroxide production, we investigated this trait in a family-based cohort consisting of family members (n5236) ascertained through probands (n557) with essential hypertension. Significant effects on hydrogen peroxide production were found for gender and ethnicity,

Fred Lacy; Mala T. Kailasam; Daniel T. O'Connor; Geert W. Schmid-Schonbein; Robert J. Parmer

107

Plasmid selection in Escherichia coli using an endogenous essential gene marker  

Microsoft Academic Search

BACKGROUND: Antibiotic resistance genes are widely used for selection of recombinant bacteria, but their use risks contributing to the spread of antibiotic resistance. In particular, the practice is inappropriate for some intrinsically resistant bacteria and in vaccine production, and costly for industrial scale production. Non-antibiotic systems are available, but require mutant host strains, defined media or expensive reagents. An unexplored

Shan Goh; Liam Good

2008-01-01

108

The Schizosaccharomyces pombe cdc5+ gene encodes an essential protein with homology to c-Myb.  

PubMed Central

The Schizosaccharomyces pombe cdc5+ gene was identified in the first screen for cell division cycle mutants in this yeast. The cdc5+ gene was reported to be required for nuclear division but because of its modest elongation and leaky nature at the non-permissive temperature, it was not investigated further. Here, we report the characterization of the single allele of this gene, cdc5-120, in more detail. The mutant arrests with a 2N DNA content and a single interphase nucleus. Further genetic analyses suggest that cdc5+ gene function is essential in the G2 phase of the cell cycle. We have cloned and sequenced the cdc5+ gene. The deduced protein sequence predicts that Cdc5 is an 87 kDa protein and contains a region sharing significant homology with the DNA binding domain of the Myb family of transcription factors. Deletion mapping of the cdc5+ gene has shown that the N-terminal 232 amino acids of the protein, which contain the Myb-related region, are sufficient to complement the cdc5ts strain. A cdc5 null mutant was generated by homologous recombination. Haploid cells lacking cdc5+ are inviable, indicating that cdc5+ is an essential gene. A fusion protein consisting of bacterial glutathione S-transferase joined in-frame to the N-terminal 127 amino acids of the Cdc5 protein is able to bind to DNA cellulose at low salt concentrations. This evidence suggests that cdc5+ might encode a transcription factor whose activity is required for cell cycle progression and growth during G2. Images

Ohi, R; McCollum, D; Hirani, B; Den Haese, G J; Zhang, X; Burke, J D; Turner, K; Gould, K L

1994-01-01

109

A study of the production of essential oils in chamomile hairy root cultures  

Microsoft Academic Search

Summary  The active substances in chamomile (Matricaria recutita L.) belong to chemically different structural types. The largest group of medically important compounds forming the essential\\u000a oils are primarily chamazulene, (?)-?-bisabolol, bisabololoxides, bisabolonoxide A,trans-?-farnesene, ?-farnesene, spathulenol and thecis\\/trans-en-in-dicycloethers. Flavonoids, coumarins, mucilages, mono- and oligosaccharides also have pharmacological effects. We studied\\u000a the production of essential oils in genetically transformed cultures. Sterile juvenile chamomile

E. Máday; É. Szöke; Zs. Muskáth; É. Lemberkovics

1999-01-01

110

Targeted deletion of the murine corneodesmosin gene delineates its essential role in skin and hair physiology  

PubMed Central

Controlled proteolytic degradation of specialized junctional structures, corneodesmosomes, by epidermal proteases is an essential process for physiological desquamation of the skin. Corneodesmosin (CDSN) is an extracellular component of corneodesmosomes and, although considerable debate still exists, genetic studies have suggested that the CDSN gene in the major psoriasis-susceptibility locus (PSORS1) may be responsible for susceptibility to psoriasis, a human skin disorder characterized by excessive growth and aberrant differentiation of keratinocytes. CDSN is also expressed in the inner root sheath of hair follicles, and a heterozygous nonsense mutation of the CDSN gene in humans is associated with scalp-specific hair loss of poorly defined etiology. Here, we have investigated the pathogenetic roles of CDSN loss of function in the development of skin diseases by generating a mouse strain with targeted deletion of the Cdsn gene. Cdsn-deficient mouse skin showed detachment of the stratum corneum from the underlying granular layer and/or detachment within the upper granular layers due to the disrupted integrity of the corneodesmosomes. When grafted onto immunodeficient mice, Cdsn-deficient skin showed rapid hair loss together with epidermal abnormalities resembling psoriasis. These results underscore the essential roles of CDSN in hair physiology and suggest functional relevance of CDSN gene polymorphisms to psoriasis susceptibility.

Matsumoto, Mitsuru; Zhou, Yiqing; Matsuo, Shinji; Nakanishi, Hideki; Hirose, Kenji; Oura, Hajimu; Arase, Seiji; Ishida-Yamamoto, Akemi; Bando, Yoshimi; Izumi, Keisuke; Kiyonari, Hiroshi; Oshima, Naoko; Nakayama, Rika; Matsushima, Akemi; Hirota, Fumiko; Mouri, Yasuhiro; Kuroda, Noriyuki; Sano, Shigetoshi; Chaplin, David D.

2008-01-01

111

Regulation by dietary essential fatty acid balance of tumor necrosis factor production in mouse macrophages  

Microsoft Academic Search

Increasing the dietary a-linolenate (18:3n - 3)\\/linoleate (18:2n - 6) ratio results in an increase in lipopolysaccharide (LPS)-stimulated tumor necrosis fac- tor (TNF) production in mouse resident and casein-in- duced peritoneal macrophages (3). We found that pros- taglandin E2 (PGE2) production is inversely related to TNF production and that indomethacin abolishes the ef- fect of changing the essential fatty acid

Shiro Watanabe; Kikuo Onozaki; Shunsuke Yamamoto; Harumi Okuyama

112

UBA 1: an essential yeast gene encoding ubiquitin-activating enzyme.  

PubMed Central

All known functions of ubiquitin are mediated through its covalent attachment to other proteins. The post-translational formation of ubiquitin--protein conjugates is preceded by an ATP-requiring step in which the carboxyl terminus of ubiquitin is adenylated and subsequently joined, through a thiolester bond, to a cysteine residue in the ubiquitin-activating enzyme, also known as E1. We report the isolation and functional analysis of the gene (UBA1) for the ubiquitin-activating enzyme of the yeast Saccharomyces cerevisiae. UBA1 encodes a 114 kd protein whose amino acid sequence contains motifs characteristic of nucleotide-binding sites. Expression of catalytically active UBA1 protein in E. coli, which lacks the ubiquitin system, confirmed that the yeast UBA1 gene encodes a ubiquitin-activating enzyme. Deletion of the UBA1 gene is lethal, demonstrating that the formation of ubiquitin--protein conjugates is essential for cell viability. Images

McGrath, J P; Jentsch, S; Varshavsky, A

1991-01-01

113

The novel function of the Saccharomyces cerevisiae CBP2 gene as a splicing factor essential to excision of the Saccharomyces douglasii LSU intron in vivo.  

PubMed

In the yeast Saccharomyces cerevisiae, the product of the nuclear gene CBP2 is required exclusively for the splicing of the terminal intron of the mitochondrial cytochrome b gene. The homologous gene from the related yeast, Saccharomyces douglasii, has been shown to be essential for respiratory growth in the presence of a wild-type S. douglasii mitochondrial genome and dispensable in the presence of an intronless mitochondrial genome. The two CBP2 genes are functionally interchangeable although the target intron of the S. cerevisiae CBP2 gene is absent from the S. douglasii mitochondrial genome. To determine the function of the CBP2 gene in S. douglasii mitochondrial pre-RNA processing we have constructed and analyzed interspecific hybrid strains between the nuclear genome of S. cerevisiae carrying an inactive CBP2 gene and S. douglasii mitochondrial genomes with different intron contents. We have demonstrated that inactivation of the S. cerevisiae CBP2 gene affects the maturation of the S. douglasii LSU pre-RNA, leading to a respiratory-deficient phenotype in the hybrid strains. We have shown that the CBP2 gene is essential for excision of the S. douglasii LSU intron in vivo and that the gene is dispensable when this intron is deleted or replaced by the S. cerevisiae LSU intron. PMID:9613573

Tian, G L; Li, G Y; Slonimski, P P; Lazowska, J

1998-04-01

114

Mutations in the essential Escherichia coli gene, yqgF, and their effects on transcription.  

PubMed

The Escherichia coli yqgF gene is highly conserved across a broad spectrum of bacterial genomes. The gene was first identified as being essential for cell growth during screening for targets for broad-spectrum antibiotics. YqgF is structurally similar to RuvC, a Holliday junction resolvase, but its function has not been established. This study describes the isolation of a temperature-sensitive yqgF mutant, the growth of which was inhibited by rho or nusA multicopy plasmids, indicating that YqgF is involved in transcription. Rho is a global transcription termination factor that acts at Rho-dependent terminator sites, which exist not only at the ends of genes but also within genes. The transcription of genes possessing intragenic, or upstream, Rho-dependent terminators was reduced in temperature-sensitive yqgF mutants. This transcription inhibition was sensitive to the Rho inhibitor, bicyclomycin. In addition, the transcription of mutant tnaA genes defective for upstream Rho-dependent termination was not significantly affected by the yqgF mutation. Taken together, these results suggest that YqgF is involved in anti-termination at Rho-dependent terminators in vivo. PMID:22353788

Iwamoto, Akira; Osawa, Atsushi; Kawai, Makiko; Honda, Hirofumi; Yoshida, Saori; Furuya, Nobuhisa; Kato, Jun-Ichi

2012-02-21

115

The chloroplast ribosomal protein L21 gene is essential for plastid development and embryogenesis in Arabidopsis.  

PubMed

Embryogenesis in higher plants is controlled by a complex gene network. Identification and characterization of genes essential for embryogenesis will provide insights into the early events in embryo development. In this study, a novel mutant with aborted seed development (asd) was identified in Arabidopsis. The asd mutant produced about 25% of albino seeds at the early stage of silique development. The segregation of normal and albino seeds was inherited as a single recessive embryo-lethal trait. The gene disrupted in the asd mutant was isolated through map-based cloning. The mutated gene contains a single base change (A to C) in the coding region of RPL21C (At1g35680) that is predicted to encode the chloroplast 50S ribosomal protein L21. Allele test with other two T-DNA insertion lines in RPL21C and a complementation test demonstrated that the mutation in RPL21C was responsible for the asd phenotype. RPL21C exhibits higher expression in leaves and flowers compared with expression levels in roots and developing seeds. The RPL21C-GFP fusion protein was localized in chloroplasts. Cytological observations showed that the asd embryo development was arrested at the globular stage. There were no plastids with normal thylakoids and as a result no normal chloroplasts formed in mutant cells, indicating an indispensable role of the ASD gene in chloroplasts biogenesis. Our studies suggest that the chloroplast ribosomal protein L21 gene is required for chloroplast development and embryogenesis in Arabidopsis. PMID:22105802

Yin, Tuanzhang; Pan, Gang; Liu, Han; Wu, Jian; Li, Yongpeng; Zhao, Zhenxing; Fu, Tingdong; Zhou, Yongming

2011-11-22

116

Evolutionary changes in the expression pattern of a developmentally essential gene in three Drosophila species.  

PubMed Central

The hypothesis that morphological evolution may largely result from changes in gene regulation rather than gene structure has been difficult to test. Morphological differences among insects are often apparent in the cuticle structures produced. The dopa decarboxylase (Ddc) and alpha-methyldopa hypersensitive (amd) genes arose from an ancient gene duplication. In Drosophila, they have evolved nonoverlapping functions, including the production of distinct types of cuticle, and for Ddc, the production of the neurotransmitters, dopamine and serotonin. The amd gene is particularly active in the production of specialized flexible cuticles in the developing embryo. We have compared the pattern of amd expression in three Drosophila species. Several regions of expression conserved in all three species but, surprisingly, a unique domain of expression is found in Drosophila simulans that does occur in the closely related (2-5 million years) Drosophila melanogaster or in the more remote (40-50 million years) Drosophila virilis. The "sudden" appearance of a completely new and robust domain of expression provides a glimpse of evolutionary variation resulting from changes in regulation of structural gene expression. Images Fig. 1 Fig. 2 Fig. 3

Wang, D; Marsh, J L; Ayala, F J

1996-01-01

117

A risk of essential thrombocythemia in carriers of constitutional CHEK2 gene mutations.  

PubMed

Germline mutations of the CHEK2 gene have been reported in some myeloid and lymphoid malignancies, but their impact on development of essential thrombocythemia has not been studied. In 16 out of 106 (15.1%) consecutive patients, newly diagnosed with essential thrombocythemia, we found one of four analyzed CHEK2 mutations: I157T, 1100delC, IVS2+1G>A or del5395. They were associated with the increased risk of disease (OR=3.8; P=0.002). The median age at ET diagnosis among CHEK2+/JAK2V617F+ patients was seven years lower than that among CHEK2-/JAK2V617F+ (52 vs. 59 years; P=0.04), whereas there was no difference in the medians of hematologic parameters between these groups. The results obtained suggest that CHEK2 mutations could potentially contribute to the susceptibility to essential thrombocythemia. The germline inactivation of CHEK2, as it seems, has no direct impact on the development of disease, but it could cause disruption of cell cycle checkpoints and initiate or support the cancerogenic process of essential thrombocythemia at a younger age. PMID:22058216

Janiszewska, Hanna; Bak, Aneta; Pilarska, Maria; Heise, Marta; Junkiert-Czarnecka, Anna; Kuliszkiewicz-Janus, Ma?gorzata; Ca?becka, Ma?gorzata; Ja?wiec, Bozena; Wo?owiec, Dariusz; Kuliczkowski, Kazimierz; Haus, Olga

2011-11-04

118

An Arabidopsis tissue-specific RNAi method for studying genes essential to mitosis.  

PubMed

A large fraction of the genes in plants can be considered essential in the sense that when absent the plant fails to develop past the first few cell divisions. The fact that angiosperms pass through a haploid gametophyte stage can make it challenging to propagate such mutants even in the heterozygous condition. Here we describe a tissue-specific RNAi method that allows us to visualize cell division phenotypes in petals, which are large dispensable organs. Portions of the APETALA (AP3) and PISTILLATA (PI) promoters confer early petal-specific expression. We show that when either promoter is used to drive the expression of a beta-glucuronidase (GUS) RNAi transgene in plants uniformly expressing GUS, GUS expression is knocked down specifically in petals. We further tested the system by targeting the essential kinetochore protein CENPC and two different components of the Spindle Assembly Checkpoint (MAD2 and BUBR1). Plant lines expressing petal-specific RNAi hairpins targeting these genes exhibited an array of petal phenotypes. Cytological analyses of the affected flower buds confirmed that CENPC knockdown causes cell cycle arrest but provided no evidence that either MAD2 or BUBR1 are required for mitosis (although both genes are required for petal growth by this assay). A key benefit of the petal-specific RNAi method is that the phenotypes are not expressed in the lineages leading to germ cells, and the phenotypes are faithfully transmitted for at least four generations despite their pronounced effects on growth. PMID:23236491

Burgos-Rivera, Brunilís; Dawe, R Kelly

2012-12-07

119

Molecular genetic analysis of a vaccinia virus gene with an essential role in DNA replication  

SciTech Connect

The poxvirus, vaccinia, is large DNA virus which replicates in the cytoplasma of the host cell. The virus is believed to encode most or all of the functions required for the temporally regulated transcription and replication of its 186 kilobase genome. Physical and genetic autonomy from the host make vaccinia a useful eukaryotic organism in which to study replication genes and proteins, using a combination of biochemical and genetic techniques. Essential viral functions for replication are identified by conditional lethal mutants that fail to synthesize DNA at the non-permissive temperatures. One such group contains the non-complementing alleles ts17, ts24, ts69 (WR strain). Studies were undertaken to define the phenotype of ts mutants, and to identify and characterize the affected gene and protein. Mutant infection was essentially normal at 32{degree}C, but at 39{degree}C the mutants did not incorporate {sup 3}H-thymidine into nascent viral DNA or synthesize late viral proteins. If mutant cultures were shifted to non-permissive conditions at the height of replication, DNA synthesis was halted rapidly, implying that the mutants are defective in DNA elongation. The gene affected in the WR mutants and in ts6389, a DNA-minus mutant of the IHD strain, was mapped by marker rescue and corresponds to open reading frame 5 (orfD5) of the viral HindIII D fragment.

Evans, E.V.A.

1989-01-01

120

Application of an inducible system to engineer unmarked conditional mutants of essential genes of Pseudomonas aeruginosa.  

PubMed

The Phi CTX-based integration vector pYM101 harboring a tightly controlled modified phage T7 early gene promoter/LacI(q) repressor (T7/LacI) system was constructed for the generation of unmarked conditional mutants in Pseudomonas aeruginosa. Promoter activity of the T7/LacI system was demonstrated to be dependent on the presence of the inducer isopropyl -beta-D-1-thiogalactopyranoside (IPTG), as evaluated by measuring beta-galactosidase activity. In the absence of the inducer, the promoter was silent as its activity was lower than those of a promoter-less lacZ control. Unmarked conditional mutants of four predicted essential genes (lolCDE (PA2988-86), lpxC (PA4406), rho (PA5239), and def (PA0019)) were successfully constructed using this recombination system. In the absence of IPTG, the growth of all mutants was repressed; however, the addition of either 0.1 or 1mM IPTG restored growth rates to levels nearly identical to wild-type cells. It was therefore demonstrated that the inducible integration vector pYM101 is suitable for the creation of unmarked conditional mutants of P. aeruginosa, and is particularly useful for examining the function of essential genes. PMID:20538017

Morita, Yuji; Narita, Shin-ichiro; Tomida, Junko; Tokuda, Hajime; Kawamura, Yoshiaki

2010-06-09

121

Senataxin plays an essential role with DNA damage response proteins in meiotic recombination and gene silencing.  

PubMed

Senataxin, mutated in the human genetic disorder ataxia with oculomotor apraxia type 2 (AOA2), plays an important role in maintaining genome integrity by coordination of transcription, DNA replication, and the DNA damage response. We demonstrate that senataxin is essential for spermatogenesis and that it functions at two stages in meiosis during crossing-over in homologous recombination and in meiotic sex chromosome inactivation (MSCI). Disruption of the Setx gene caused persistence of DNA double-strand breaks, a defect in disassembly of Rad51 filaments, accumulation of DNA:RNA hybrids (R-loops), and ultimately a failure of crossing-over. Senataxin localised to the XY body in a Brca1-dependent manner, and in its absence there was incomplete localisation of DNA damage response proteins to the XY chromosomes and ATR was retained on the axial elements of these chromosomes, failing to diffuse out into chromatin. Furthermore persistence of RNA polymerase II activity, altered ubH2A distribution, and abnormal XY-linked gene expression in Setx?/? revealed an essential role for senataxin in MSCI. These data support key roles for senataxin in coordinating meiotic crossing-over with transcription and in gene silencing to protect the integrity of the genome. PMID:23593030

Becherel, Olivier J; Yeo, Abrey J; Stellati, Alissa; Heng, Evelyn Y H; Luff, John; Suraweera, Amila M; Woods, Rick; Fleming, Jean; Carrie, Dianne; McKinney, Kristine; Xu, Xiaoling; Deng, Chuxia; Lavin, Martin F

2013-04-11

122

An Arabidopsis Tissue-Specific RNAi Method for Studying Genes Essential to Mitosis  

PubMed Central

A large fraction of the genes in plants can be considered essential in the sense that when absent the plant fails to develop past the first few cell divisions. The fact that angiosperms pass through a haploid gametophyte stage can make it challenging to propagate such mutants even in the heterozygous condition. Here we describe a tissue-specific RNAi method that allows us to visualize cell division phenotypes in petals, which are large dispensable organs. Portions of the APETALA (AP3) and PISTILLATA (PI) promoters confer early petal-specific expression. We show that when either promoter is used to drive the expression of a beta-glucuronidase (GUS) RNAi transgene in plants uniformly expressing GUS, GUS expression is knocked down specifically in petals. We further tested the system by targeting the essential kinetochore protein CENPC and two different components of the Spindle Assembly Checkpoint (MAD2 and BUBR1). Plant lines expressing petal-specific RNAi hairpins targeting these genes exhibited an array of petal phenotypes. Cytological analyses of the affected flower buds confirmed that CENPC knockdown causes cell cycle arrest but provided no evidence that either MAD2 or BUBR1 are required for mitosis (although both genes are required for petal growth by this assay). A key benefit of the petal-specific RNAi method is that the phenotypes are not expressed in the lineages leading to germ cells, and the phenotypes are faithfully transmitted for at least four generations despite their pronounced effects on growth.

Burgos-Rivera, Brunilis; Dawe, R. Kelly

2012-01-01

123

Coupling mutagenesis and parallel deep sequencing to probe essential residues in a genome or gene.  

PubMed

The sequence of a protein determines its function by influencing its folding, structure, and activity. Similarly, the most conserved residues of orthologous and paralogous proteins likely define those most important. The detection of important or essential residues is not always apparent via sequence alignments because these are limited by the depth of any given gene's phylogeny, as well as specificities that relate to each protein's unique biological origin. Thus, there is a need for robust and comprehensive ways of evaluating the importance of specific amino acid residues of proteins of known or unknown function. Here we describe an approach called Mut-seq, which allows the identification of virtually all of the essential residues present in a whole genome through the application of limited chemical mutagenesis, selection for function, and deep parallel genomic sequencing. Here we have applied this method to T7 bacteriophage and T7-like virus JSF7 of Vibrio cholerae. PMID:23401533

Robins, William P; Faruque, Shah M; Mekalanos, John J

2013-02-11

124

RIM2, MSI1 and PGI1 are located within an 8 kb segment of Saccharomyces cerevisiae chromosome II, which also contains the putative ribosomal gene L21 and a new putative essential gene with a leucine zipper motif.  

PubMed

We report the DNA sequence of an 8 kb segment localized on the right arm of chromosome II from Saccharomyces cerevisiae. The sequence reveals the presence of eight open reading frames (ORFs). Three of them, YBR1402, YBR1405 and YBR1406 are previously sequenced genes, respectively the RIM2 (replication in mitochondria), MSI1 (multicopy suppressor of IRA1 gene) and PGI1 (phosphoglucoisomerase) genes. The predicted product of the ORF YBR1401 could be the putative yeast ribosomal protein L21. A new essential gene, YBR1403, has been identified by disruption; it possesses a leucine zipper motif. PMID:8346681

Démolis, N; Mallet, L; Bussereau, F; Jacquet, M

1993-06-01

125

Effect of commercially available plant-derived essential oil products on arthropod pests.  

PubMed

Plant-derived essential oil products, in general, are considered minimum-risk pesticides and are exempt from Environmental Protection Agency registration under section 25(b) of the Federal Insecticide Fungicide and Rodenticide Act. However, many of the plant-derived essential products available to consumers (homeowners) have not been judiciously evaluated for both efficacy and plant safety. In fact, numerous plant-derived essential oil products labeled for control of arthropod pests have not been subject to rigorous evaluation, and there is minimal scientific information or supporting data associated with efficacy against arthropod pests. We conducted a series of greenhouse experiments to determine the efficacy and phytotoxicity of an array of plant-derived essential oil products available to consumers on arthropod pests including the citrus mealybug, Planococcus citri (Risso); western flower thrips, Frankliniella occidentalis (Pergande); twospotted spider mite, Tetranychus urticae Koch; sweetpotato whitefly B-biotype, Bemisia tabaci (Gennadius); and green peach aphid, Myzus persicae (Sulzer). Although the products Flower Pharm (cottonseed, cinnamon, and rosemary oil) and Indoor Pharm (soybean, rosemary, and lavender oil) provided > 90% mortality of citrus mealybug, they were also the most phytotoxic to the coleus, Solenostemon scutellarioides (L.) Codd, plants. Both GC-Mite (cottonseed, clove, and garlic oil) and Bugzyme (citric acid) were most effective against the twospotted spider mite (> or = 90% mortality). However, SMC (canola, coriander oil, and triethanolamine), neem (clarified hydrophobic extract of neem oil), and Bug Assassin (eugenol, sodium lauryl sulfate, peppermint, and citronella oil) provided > 80% mortality. Monterey Garden Insect Spray, which contained 0.5% spinosad, was most effective against western flower thrips with 100% mortality. All the other products evaluated failed to provide sufficient control of western flower thrips with < 30% mortality. In addition, the products Pest Out (cottonseed, clove, and garlic oil), Bang (Pipereaceae), and Fruit & Vegetable Insect Spray (rosemary, cinnamon, clove oil, and garlic extract) had the highest flower (transvaal daisy, Gerberajamesonii [H. Bolus ex Hook.f]) phytotoxicity ratings (> or = 4.5 of 5) among all the products. None of the plant-derived essential oil products provided sufficient control of sweetpotato whitefly B-biotype or green peach aphid 7, 14, and 21 d after application. Furthermore, the products Bug Assassin (eugenol, sodium lauryl sulfate, peppermint, and citronella oil) and Sharpshooter (sodium lauryl sulfate and clove oil) were phytotoxic to the poinsettia, Euphorbia pulcherrima Willd. ex Klotzsch, plants. This study is one of the first to quantitatively demonstrate that commercially available plant-derived essential oil products vary in their effectiveness against certain arthropod pests stated on the label and are phytotoxic. PMID:19736770

Cloyd, Raymond A; Galle, Cindy L; Keith, Stephen R; Kalscheur, Nanette A; Kemp, Kenneth E

2009-08-01

126

Protein-network modeling of prostate cancer gene signatures reveals essential pathways in disease recurrence  

PubMed Central

Objective Uncovering the dominant molecular deregulation among the multitude of pathways implicated in aggressive prostate cancer is essential to intelligently developing targeted therapies. Paradoxically, published prostate cancer gene expression signatures of poor prognosis share little overlap and thus do not reveal shared mechanisms. The authors hypothesize that, by analyzing gene signatures with quantitative models of protein–protein interactions, key pathways will be elucidated and shown to be shared. Design The authors statistically prioritized common interactors between established cancer genes and genes from each prostate cancer signature of poor prognosis independently via a previously validated single protein analysis of network (SPAN) methodology. Additionally, they computationally identified pathways among the aggregated interactors across signatures and validated them using a similarity metric and patient survival. Measurement Using an information-theoretic metric, the authors assessed the mechanistic similarity of the interactor signature. Its prognostic ability was assessed in an independent cohort of 198 patients with high-Gleason prostate cancer using Kaplan–Meier analysis. Results Of the 13 prostate cancer signatures that were evaluated, eight interacted significantly with established cancer genes (false discovery rate <5%) and generated a 42-gene interactor signature that showed the highest mechanistic similarity (p<0.0001). Via parameter-free unsupervised classification, the interactor signature dichotomized the independent prostate cancer cohort with a significant survival difference (p=0.009). Interpretation of the network not only recapitulated phosphatidylinositol-3 kinase/NF-?B signaling, but also highlighted less well established relevant pathways such as the Janus kinase 2 cascade. Conclusions SPAN methodolgy provides a robust means of abstracting disparate prostate cancer gene expression signatures into clinically useful, prioritized pathways as well as useful mechanistic pathways.

Chen, James L; Li, Jianrong; Stadler, Walter M

2011-01-01

127

Genome-Wide Analysis Reveals Novel Genes Essential for Heme Homeostasis in Caenorhabditis elegans  

PubMed Central

Heme is a cofactor in proteins that function in almost all sub-cellular compartments and in many diverse biological processes. Heme is produced by a conserved biosynthetic pathway that is highly regulated to prevent the accumulation of heme—a cytotoxic, hydrophobic tetrapyrrole. Caenorhabditis elegans and related parasitic nematodes do not synthesize heme, but instead require environmental heme to grow and develop. Heme homeostasis in these auxotrophs is, therefore, regulated in accordance with available dietary heme. We have capitalized on this auxotrophy in C. elegans to study gene expression changes associated with precisely controlled dietary heme concentrations. RNA was isolated from cultures containing 4, 20, or 500 µM heme; derived cDNA probes were hybridized to Affymetrix C. elegans expression arrays. We identified 288 heme-responsive genes (hrgs) that were differentially expressed under these conditions. Of these genes, 42% had putative homologs in humans, while genomes of medically relevant heme auxotrophs revealed homologs for 12% in both Trypanosoma and Leishmania and 24% in parasitic nematodes. Depletion of each of the 288 hrgs by RNA–mediated interference (RNAi) in a transgenic heme-sensor worm strain identified six genes that regulated heme homeostasis. In addition, seven membrane-spanning transporters involved in heme uptake were identified by RNAi knockdown studies using a toxic heme analog. Comparison of genes that were positive in both of the RNAi screens resulted in the identification of three genes in common that were vital for organismal heme homeostasis in C. elegans. Collectively, our results provide a catalog of genes that are essential for metazoan heme homeostasis and demonstrate the power of C. elegans as a genetic animal model to dissect the regulatory circuits which mediate heme trafficking in both vertebrate hosts and their parasites, which depend on environmental heme for survival.

Rao, Anita U.; Cerqueira, Gustavo C.; Mitreva, Makedonka; El-Sayed, Najib M.; Krause, Michael; Hamza, Iqbal

2010-01-01

128

Discovery of genes essential for heme biosynthesis through large-scale gene expression analysis  

PubMed Central

Summary Heme biosynthesis consists of a series of eight enzymatic reactions that originate in mitochondria and continue in the cytosol before returning to mitochondria. Although these core enzymes are well studied, additional mitochondrial transporters and regulatory factors are predicted to be required. To discover such unknown components, we utilized a large-scale computational screen to identify mitochondrial proteins whose transcripts consistently co-express with the core machinery of heme biosynthesis. We identified SLC25A39, SLC22A4 and TMEM14C, which are putative mitochondrial transporters, as well as C1orf69 and ISCA1, which are iron-sulfur cluster proteins. Targeted knockdowns of all five genes in zebrafish resulted in profound anemia without impacting erythroid lineage specification. Moreover, silencing of Slc25a39 in murine erythroleukemia cells impaired iron incorporation into protoporphyrin IX, and vertebrate Slc25a39 complemented an iron homeostasis defect in the orthologous yeast mtm1? deletion mutant. Our results advance the molecular understanding of heme biosynthesis and offer promising candidate genes for inherited anemias.

Nilsson, Roland; Schultz, Iman J.; Pierce, Eric L.; Soltis, Kathleen A.; Naranuntarat, Amornrat; Ward, Diane M.; Baughman, Joshua; Paradkar, Prasad N.; Kingsley, Paul D.; Culotta, Valeria C.; Kaplan, Jerry; Palis, James; Paw, Barry H.; Mootha, Vamsi K.

2009-01-01

129

Urease and Nitrification Retardation Properties in Natural Essential Oils and Their By?products  

Microsoft Academic Search

A laboratory incubation experiment was conducted with the natural essential oil of peppermint (Mentha spicata, MS), dementholized oil (DMO), terpenes, and a chemical inhibitor dicyandiamide (DCD), as coating materials for retardation of soil urea hydrolysis and nitrification. Retardation of nitrification was highest with DCD. The urea hydrolysis and nitrification processes were inhibited by all three natural products. Intensity of inhibition

D. D. Patra; Usha Kiran; Preeti Pande

2006-01-01

130

The Murine Stanniocalcin 1 Gene Is Not Essential for Growth and Development  

PubMed Central

The stanniocalcin 1 (STC1) gene is expressed in a wide variety of tissues, including the kidney, prostate, thyroid, bone, and ovary. STC1 protein is considered to have roles in many physiological processes, including bone development, reproduction, wound healing, angiogenesis, and modulation of inflammatory response. In fish, STC1 is a hormone that is secreted by the corpuscles of Stannius and is involved in calcium and phosphate homeostasis. To determine the role of STC1 in mammals, we generated Stc1-null mice by gene targeting. The number of Stc1?/? mice obtained was in accordance with Mendelian ratios, and both males and females produced offspring normally. No anatomical or histological abnormalities were detected in any tissues. Our results demonstrated that Stc1 function is not essential for growth or reproduction in the mouse.

Chang, Andy C.-M.; Cha, Jeon; Koentgen, Frank; Reddel, Roger R.

2005-01-01

131

Integrating the Genetic and Physical Maps of Arabidopsis thaliana: Identification of Mapped Alleles of Cloned Essential (EMB) Genes  

Microsoft Academic Search

The classical genetic map of Arabidopsis includes more than 130 genes with an embryo-defective (emb) mutant phenotype. Many of these essential genes remain to be cloned. Hundreds of additional EMB genes have been cloned and catalogued (www.seedgenes.org) but not mapped. To facilitate EMB gene identification and assess the current level of saturation, we updated the classical map, compared the physical

David Meinke; Colleen Sweeney; Rosanna Muralla

2009-01-01

132

Relationships between proteasomes and viral gene products.  

PubMed

The interrelationships between proteasomes and viral gene products are very complex. 20S proteasomes associate with a number of viral mRNAs which are cleaved by proteasome's associated endonuclease activity. In addition proteasome's endopeptidase activities are involved in the presentation of viral antigens. Viral proteins of different origin associate with the 20S and 26S complexes and interfere with their enzymatic activities. A major part of this review deals with the interactions between 20S proteasomes and the gene products of the human immunodeficiency virus (HIV) which has been studied in detail by our group. PMID:10363656

Jarrousse, A S; Gautier, K; Apcher, S; Badaoui, S; Boissonnet, G; Dadet, M H; Henry, L; Bureau, J P; Schmid, H P; Petit, F

1999-04-01

133

Insecticidal activity of the essential oils from different plants against three stored-product insects.  

PubMed

This study was conducted to determine the insecticidal activity of essential oils from oregano, Origanum onites L. (Lamiales: Lamiaceae), savory, Satureja thymbra L. (Lamiales: Lamiaceae), and myrtle, Myrtus communis L. (Rosales: Myrtaceae) against three stored-product insects. Essential oils from three species of plants were obtained by Clevenger-type water distillation. The major compounds in these essential oils were identified using gas chromatography-mass spectrometry and their insecticidal activity was tested against adults of the Mediterranean flour moth Ephestia kuehniella Zeller (Lepidoptera: Pyralidae), the Indian meal moth Plodia interpunctella Hübner (Lepidoptera: Pyralidae) and the bean weevil Acanthoscelides obtectus Say (Coleoptera: Bruchidae). While the major compound found in oregano and savory was carvacrol, the main constituent of the myrtle was linalool. Among the tested insects, A. obtectus was the most tolerant species against the essential oils. However, the insecticidal activity of the myrtle oil was more pronounced than other oils tested against A. obtectus adults. The essential oils of oregano and savory were highly effective against P. interpunctella and E. kuehniella, with 100% mortality obtained after 24 h at 9 and 25 microl/l air for P. interpunctella and E. kuehniella, respectively. LC(50) and LC(99) values of each essential oil were estimated for each insect species. PMID:20578885

Ayvaz, Abdurrahman; Sagdic, Osman; Karaborklu, Salih; Ozturk, Ismet

2010-01-01

134

Insecticidal Activity of the Essential Oils from Different Plants Against Three Stored-Product Insects  

PubMed Central

This study was conducted to determine the insecticidal activity of essential oils from oregano, Origanum onites L. (Lamiales: Lamiaceae), savory, Satureja thymbra L. (Lamiales: Lamiaceae), and myrtle, Myrtus communis L. (Rosales: Myrtaceae) against three stored-product insects. Essential oils from three species of plants were obtained by Clevenger-type water distillation. The major compounds in these essential oils were identified using gas chromatography-mass spectrometry and their insecticidal activity was tested against adults of the Mediterranean flour moth Ephestia kuehniella Zeller (Lepidoptera: Pyralidae), the Indian meal moth Plodia interpunctella Hübner (Lepidoptera: Pyralidae) and the bean weevil Acanthoscelides obtectus Say (Coleoptera: Bruchidae). While the major compound found in oregano and savory was carvacrol, the main constituent of the myrtle was linalool. Among the tested insects, A. obtectus was the most tolerant species against the essential oils. However, the insecticidal activity of the myrtle oil was more pronounced than other oils tested against A. obtectus adults. The essential oils of oregano and savory were highly effective against P. interpunctella and E. kuehniella, with 100% mortality obtained after 24 h at 9 and 25 µl/l air for P. interpunctella and E. kuehniella, respectively. LC50 and LC99 values of each essential oil were estimated for each insect species.

Ayvaz, Abdurrahman; Sagdic, Osman; Karaborklu, Salih; Ozturk, Ismet

2010-01-01

135

Assessment of inhibitory potential of essential oils on natural mycoflora and Fusarium mycotoxins production in wheat  

PubMed Central

Background In the last years essential oils from different plants were used in the prevention of fungi and mycotoxins accumulation in cereals. The most attractive aspect derived from using of essential oils as seed grains protectants is due to their non-toxicity. This study was focused on assessment the inhibitory effect of some essential oils: Melissa officinalis (O1), Salvia officinalis (O2), Coriandrum sativum (O3), Thymus vulgaris (O4) Mentha piperita (O5) and Cinnamomum zeylanicum (O6) against natural mycoflora and Fusarium mycotoxins production correlated with their antioxidants properties. Results All essential oils showed inhibitory effect on fungal contamination of wheat seeds. This ability was dose-dependent. The highest inhibitory effect on Fusarium and Aspergillus fungi was recorded after 5?days of treatment. Fungi such as yeast (Pichia, Saccharomyces and Hyphopichia) were predominantly on seeds mycoflora after 22?days. Each treatment had a selective inhibitory effect on frequency of fungus genera. After 5?days of treatment the most fungicidal effect was recorder for O4, followed by O1. In terms of essential oils effect on mycotoxins development, the best control on fumonisins (FUMO) production was recorded for O6. The antioxidant properties of essential oils decreased in order: O4?>?O1?>?O6?>?O5?>?O2?>?O3. Also, our data suggested that there is a significant negative correlation between antioxidant properties and seed contamination index (SCI), but there was not recorded a good correlation between antioxidant properties and FUMO content. Conclusions Based on proven antifungal and antimycotoxin effects as well as their antioxidant properties, the essential oils could be recommended as natural preservatives for stored cereals. The highest inhibition of fungal growth was noted after 5?days of treatment and decreased after 22?days.

2013-01-01

136

The Essential Role of the Deinococcus radiodurans ssb Gene in Cell Survival and Radiation Tolerance.  

PubMed

Recent evidence has implicated single-stranded DNA-binding protein (SSB) expression level as an important factor in microbial radiation resistance. The genome of the extremely radiation resistant bacterium Deinococcus radiodurans contains genes for two SSB homologs: the homodimeric, canonical Ssb, encoded by the gene ssb, and a novel pentameric protein encoded by the gene ddrB. ddrB is highly induced upon exposure to radiation, and deletions result in decreased radiation-resistance, suggesting an integral role of the protein in the extreme resistance exhibited by this organism. Although expression of ssb is also induced after irradiation, Ssb is thought to be involved primarily in replication. In this study, we demonstrate that Ssb in D. radiodurans is essential for cell survival. The lethality of an ssb deletion cannot be complemented by providing ddrB in trans. In addition, the radiation-sensitive phenotype conferred by a ddrB deletion is not alleviated by providing ssb in trans. By altering expression of the ssb gene, we also show that lower levels of transcription are required for optimal growth than are necessary for high radiation resistance. When expression is reduced to that of E. coli, ionizing radiation resistance is similarly reduced. UV resistance is also decreased under low ssb transcript levels where growth is unimpaired. These results indicate that the expression of ssb is a key component of both normal cellular metabolism as well as pathways responsible for the high radiation tolerance of D. radiodurans. PMID:23951213

Lockhart, J Scott; Deveaux, Linda C

2013-08-09

137

Yeast PPA2 gene encodes a mitochondrial inorganic pyrophosphatase that is essential for mitochondrial function.  

PubMed

We have cloned a gene encoding a mitochondrial inorganic pyrophosphatase (PPase) in the yeast Saccharomyces cerevisiae by low stringency hybridization to PPA1, the yeast gene for cytoplasmic PPase. The new gene, PPA2, is located on chromosome 13 and encodes a protein whose sequence is 49% identical to the cytoplasmic enzyme. The protein differs from cytoplasmic PPase in that it has a leader sequence enriched in basic and hydroxylated residues, which is typically found in mitochondrial proteins. Yeast cells overproducing PPA2 had a 47-fold increase in mitochondrial PPase activity. This activity was further stimulated 3-fold by the uncoupler carbonyl cyanide p-trifluoromethoxyphenylhydrazone, which suggests that PPA2 is part of an energy-linked enzyme. Using gene disruptions, we found that PPA1 is required for cell growth. In contrast, cells disrupted for PPA2 are viable, but unable to grow on respiratory carbon sources. Fluorescence microscopy revealed that these cells have lost their mitochondrial DNA. We conclude that the mitochondrial PPase encoded by PPA2 is essential for mitochondrial function and maintenance of the mitochondrial genome. PMID:1648084

Lundin, M; Baltscheffsky, H; Ronne, H

1991-07-01

138

Non-essential and essential trace element concentrations in meat from cattle reared under organic, intensive or conventional production systems  

Microsoft Academic Search

We evaluated if differences in non-essential and essential trace element accumulation in beef-cattle reared under different systems (including organic, conventional and intensive management) were reflected in the meat derived from these animals. Diaphragm muscle from 166 calves from nine farms were analysed. Muscle cadmium concentrations were low (<10 µg\\/kg wet weight) and muscle arsenic, mercury and lead levels were below

I. Blanco-Penedo; M. López-Alonso; M. Miranda; J. Hernández; F. Prieto; R. F. Shore

2010-01-01

139

Non-essential and essential trace element concentrations in meat from cattle reared under organic, intensive or conventional production systems.  

PubMed

We evaluated if differences in non-essential and essential trace element accumulation in beef-cattle reared under different systems (including organic, conventional and intensive management) were reflected in the meat derived from these animals. Diaphragm muscle from 166 calves from nine farms were analysed. Muscle cadmium concentrations were low (<10 microg/kg wet weight) and muscle arsenic, mercury and lead levels were below the limits of detection (<12, 2 and 3 microg/kg, respectively) in most (77-97%) samples; there were no significant differences between farms. Essential trace element concentrations in muscle were generally within adequate physiological ranges and, although they varied significantly between farms, this was not apparently related to management practices. There were no significant correlations in element concentrations between muscle and liver or kidney (organ concentrations that better reflect exposure), except for cobalt (positive association) and zinc (negative association). Non-essential and essential trace element concentrations in muscle in the studied animals did not generally reflect differences in exposure. This is particularly relevant for animals reared in systems (such as organic farms) where cattle are exposed to higher levels of non-essential elements (probably due to soil ingestion when grazing) but also can suffer from mineral deficiencies. PMID:19750401

Blanco-Penedo, I; López-Alonso, M; Miranda, M; Hernández, J; Prieto, F; Shore, R F

2010-01-01

140

A gene homologous to chloroplast carbonic anhydrase (icfA) is essential to photosynthetic carbon dioxide fixation by Synechococcus PCC7942.  

PubMed Central

To understand the CO2-concentrating mechanism in cyanobacteria, a genomic DNA fragment that complements a temperature-sensitive high-CO2 (5%)-requiring mutant of Synechococcus PCC7942 has been isolated. An open reading frame (ORF272) encoding a polypeptide of 272 amino acids (Mr, 30,184) was found within the genomic region located 20 kilobases downstream from the genes for ribulose-1,5-bisphosphate carboxylase/oxygenase (rbcLS). Insertion of a kanamycin-resistance gene cartridge within the ORF272 in wild-type cells led to a high-CO2-requiring phenotype. Strains carrying a gene disabled by insertional mutagenesis accumulated inorganic carbon in the cells, but they could not fix it efficiently, even though ribulose-1,5-bisphosphate carboxylase activity was comparable to that of the wild-type strain. Therefore, the ORF272 was designated as a gene icfA, which is essential to inorganic carbon fixation. Furthermore, the predicted icfA gene product shared significant sequence similarities with plant chloroplast carbonic anhydrases (CAs) from pea (22%) and spinach (22%) and also with the Escherichia coli cynT gene product (31%), which was recently identified to be E. coli CA. These results indicate that the putative CA encoded by icfA is essential to photosynthetic carbon dioxide fixation in cyanobacteria and that plant chloroplast CAs may have evolved from a common ancestor of the prokaryotic CAs, which are distinct from mammalian CAs and Chlamydomonas periplasmic CAs.

Fukuzawa, H; Suzuki, E; Komukai, Y; Miyachi, S

1992-01-01

141

The rev gene of visna virus is required for productive infection.  

PubMed

To assess the role of the animal lentivirus rev regulatory gene product in the life cycle of visna virus, we introduced mutations into the functional fourth exon of the rev gene of visna virus. Cultured cells transfected with a full-length clone of visna DNA produce infectious virus but visna DNA with mutations in rev does not. The documented requirement for a functional rev gene in productive infections establishes the essential role of this gene in the replication of an animal lentivirus. PMID:7510438

Toohey, K L; Haase, A T

1994-04-01

142

High risk of essential hypertension in males with intron 4 VNTR polymorphism of eNOS gene.  

PubMed

In this study 250 patients with essential hypertension were investigated in comparison to 218 normotensives for association with epidemiological parameters. Of these DNA samples from 176 patients and 168 controls were analyzed for intron 4 27bp repeat polymorphism of eNOS gene. The study revealed significantly high risk of essential hypertension for individuals who were obese, with a positive family history and with non-vegetarian food habits. Though the intron 4b/a polymorphism of eNOS gene did not reveal any association with essential hypertension in general, males with a/a genotype of the polymorphism did show significantly high risk for developing hypertension. PMID:20680151

Patkar, Sushma; Charita, B H; Ramesh, C; Padma, T

2009-05-01

143

Complex Formation and Interactions between Transcription Factors Essential for Human Prolactin Receptor Gene Transcription ?  

PubMed Central

The protein association of estrogen receptor ? ER? with DNA-bound SP1 and C/EBP? is essential for the 17?-estradiol (E2)-induced activation of human prolactin receptor (hPRLR) gene transcription. Protein-protein interaction and complex formation at the hPIII promoter of hPRLR was investigated. The basic region and leucine zipper (bZIP) of C/EBP?, zinc finger (ZF) motifs of SP1, and the DNA binding domain of ER? were identified as regions responsible for the interactions between transfactors. The E2-induced interaction was confirmed by bioluminescence resonance energy transfer (BRET) assays of live cells. The combination of BRET/bimolecular luminescence complementation assay revealed that ER? exists as a constitutive homodimer, and E2 induced a change(s) in ER? homodimer conformation favorable for its association with C/EBP? and SP1. Chromatin immunoprecipitation and small interfering RNA knockdown of members of the complex in breast cancer cells demonstrated the endogenous recruitment of components of the complex onto the hPIII promoter of the hPRLR gene. SP1 is the preferred transfactor for the recruitment of ER? to the complex that facilitates the C/EBP? association. The E2/ER?-induced hPRLR transcription was demonstrated in ER?-negative breast cancer cells. This study indicates that the enhanced complex formation of ER? dimer with SP1 and C/EBP? by E2 has an essential role in the transcriptional activation of the hPRLR gene.

Kang, Jung-Hoon; Tsai-Morris, Chon-Hwa; Dufau, Maria L.

2011-01-01

144

The Streptomyces venezuelae pikAV gene contains a transcription unit essential for expression of enzymes involved in glycosylation of narbonolide and 10-deoxymethynolide.  

PubMed

In Streptomyces venezuelae, four polyketide synthase (PKS) polypeptides encoded by pikAI-pikAIV are used to generate 10 and 12-membered macrocyclic structures, narbonolide and 10-deoxymethynolide. Sequence analysis suggests these genes are translationally coupled with downstream genes, pikAV (encoding a type II thioesterase), desVIII-desVI (encoding enzymes responsible for production of the final glycosylated products pikromycin, narbomycin, methymycin and neomethymycin) and desR (a resistance gene). Type II thioesterases have been suggested to have an editing function in polyketide biosynthesis and deletion of the corresponding genes often leads to decreased levels of polyketide production. Surprisingly an in-frame deletion of 687 bp of the 843 bp pikAV ORF led to a strain SC1022 that produced normal yields of polyketide products, but only in the aglycone form. Plasmid-based expression of the desVIII-VI and desR in the SC1022 strain completely restored production of glycosylated products, despite the absence of a functional pikAV gene product. Under these conditions the PikAV TEII therefore does not play an important role in polyketide biosynthesis, and its function remains an enigma. These observations also demonstrate that the region of pikAV DNA deleted in strain SC1022 contains a transcription unit essential for expression of the des genes. A sequence alignment of PikAV with members of the highly conserved type II thioesterases revealed a short divergent region at the carboxy terminus, suggesting a region of pikAV that might contain such a transcription unit. DNA containing this region of pikAV was shown to be able to increase plasmid-based expression of both crotonyl CoA reductase gene (ccr) and the erythromycin resistance gene (ermE) in S. venezuelae. PMID:11223265

Chen, S; Roberts, J B; Xue, Y; Sherman, D H; Reynolds, K A

2001-01-24

145

NAT2, an essential gene encoding methionine N alpha-acetyltransferase in the yeast Saccharomyces cerevisiae.  

PubMed

N alpha-Acetylation is catalyzed by N alpha-acetyltransferases, which transfer acetyl groups from acetyl coenzyme A to the N termini of most eukaryotic proteins co-translationally. NAT1 and ARD1 from the yeast Saccharomyces cerevisiae (Mullen, J. R., Kayne, P. S., Moerschell, R. P., Tsunasawa, S., Gribskov, M., Colavito-Shepanski, M., Grunstein, M., Sherman, F., and Sternglanz, R. (1989) EMBO J. 8, 2067-2075) were previously shown to encode the major N alpha-acetyltransferase, which act on certain proteins having serine, glycine, and alanine but not methionine termini (Sherman, F., Moerschell, R. P., Tsunasawa, S., and Sternglanz, R. (1993) in Methods in Protein Sequence Analysis (Imahori, K., and Sakiyama, F., eds) pp. 173-181, Plenum Publishing Corp., New York). We have identified a second gene, NAT2, that may correspond to the N alpha-acetyltransferase acting on a subset of proteins having methionine termini. Crude extracts of a series of heat-sensitive mutants (Ts-) were screened for acetylation of a 24-amino acid synthetic peptide Met-Asn-Asn- in vitro. One mutant, nat2-1, out of 115 strains examined, lacked acetyltransferase activity, and the mutation co-segregated as a single gene with the heat-sensitive phenotype. The nat2-1 mutants were deficient in the ability to acetylate Met-Asn-Asn- and Met-Glu-Arg-peptides but were able to N alpha-acetylate Ser-Glu-Phe- and Ser-Tyr-Ser- peptides in vitro. The NAT2 wild-type gene was cloned by complementation of the nat2-1 mutant, and the DNA sequence revealed an open reading frame of 288 amino acids. Gene disruption demonstrated that NAT2 is an essential gene, and hybridization analysis indicated that it is located on chromosome VII. Furthermore, there was limited, but significant identities between the yeast N alpha-acetyltransferases Nat1, Ard1, Nat2, and Mak3, although no common motifs could be identified. We propose that NAT2 encodes the major N alpha-acetyl-transferase acting on certain proteins with only methionine termini, and that N alpha-acetylation of some of these proteins is essential for viability. PMID:8175741

Kulkarni, M S; Sherman, F

1994-05-01

146

Regulated Ectopic Expression and Allelic-Replacement Mutagenesis as a Method for Gene Essentiality Testing in Staphylococcus aureus  

Microsoft Academic Search

Conditional expression systems were utilized for the ectopic induction of essential genes in Staphylococcus aureus. Resulting strains were then subjected to allelic-replacement mutagenesis of the native allele under inducing conditions for expression of the ectopic copy of the gene. This strategy produced test strains whereby cellular viability was uniquely dependent on the presence of inducer and provided a direct and

Frank Fan; R. Dwayne Lunsford; Daniel Sylvester; Jing Fan; Helena Celesnik; Serban Iordanescu; Martin Rosenberg; Damien McDevitt

2001-01-01

147

Acidic pH induced STM1485 gene is essential for intracellular replication of Salmonella  

PubMed Central

During the course of infection, Salmonella has to face several potentially lethal environmental conditions, one such being acidic pH. The ability to sense and respond to the acidic pH is crucial for the survival and replication of Salmonella. The physiological role of one gene (STM1485) involved in this response, which is upregulated inside the host cells (by 90- to 113-fold) is functionally characterized in Salmonella pathogenesis. In vitro, the ?STM1485 neither exhibited any growth defect at pH 4.5 nor any difference in the acid tolerance response. The ?STM1485 was compromised in its capacity to proliferate inside the host cells and complementation with STM1485 gene restored its virulence. We further demonstrate that the surface translocation of Salmonella pathogenicity island-2 (SPI-2) encoded translocon proteins, SseB and SseD were reduced in the ?STM1485. The increase in co-localization of this mutant with lysosomes was also observed. In addition, the ?STM1485 displayed significantly reduced competitive indices (CI) in spleen, liver and mesenteric lymph nodes in murine typhoid model when infected by intra-gastric route. Based on these results, we conclude that the acidic pH induced STM1485 gene is essential for intracellular replication of Salmonella.

Allam, Uday Sankar; Krishna, M. Gopala; Sen, Minakshi; Thomas, Rony; Lahiri, Amit; Gnanadhas, Divya Prakash; Chakravortty, Dipshikha

2012-01-01

148

ICER induced by hyperglycemia represses the expression of genes essential for insulin exocytosis  

PubMed Central

The GTPases Rab3a and Rab27a and their effectors Granuphilin/Slp4 and Noc2 are essential regulators of neuroendocrine secretion. Chronic exposure of pancreatic ?-cells to supraphysiological glucose levels decreased selectively the expression of these proteins. This glucotoxic effect was mimicked by cAMP-raising agents and blocked by PKA inhibitors. We demonstrate that the transcriptional repressor ICER, which is induced in a PKA-dependent manner by chronic hyperglycemia and cAMP-raising agents, is responsible for the decline of the four genes. ICER overexpression diminished the level of Granuphilin, Noc2, Rab3a and Rab27a by binding to cAMP responsive elements located in the promoters of these genes and inhibited exocytosis of ?-cells in response to secretagogues. Moreover, the loss in the expression of the genes of the secretory machinery caused by glucose and cAMP-raising agents was prevented by an antisense construct that reduces ICER levels. We propose that induction of inappropriate ICER levels lead to defects in the secretory process of pancreatic ?-cells possibly contributing, in conjunction with other known deleterious effects of hyperglycemia, to defective insulin release in type 2 diabetes.

Abderrahmani, Amar; Cheviet, Severine; Ferdaoussi, Mourad; Coppola, Thierry; Waeber, Gerard; Regazzi, Romano

2006-01-01

149

Gene PA2449 Is Essential for Glycine Metabolism and Pyocyanin Biosynthesis in Pseudomonas aeruginosa PAO1  

PubMed Central

Many pseudomonads produce redox active compounds called phenazines that function in a variety of biological processes. Phenazines are well known for their toxicity against non-phenazine-producing organisms, which allows them to serve as crucial biocontrol agents and virulence factors during infection. As for other secondary metabolites, conditions of nutritional stress or limitation stimulate the production of phenazines, but little is known of the molecular details underlying this phenomenon. Using a combination of microarray and metabolite analyses, we demonstrate that the assimilation of glycine as a carbon source and the biosynthesis of pyocyanin in Pseudomonas aeruginosa PAO1 are both dependent on the PA2449 gene. The inactivation of the PA2449 gene was found to influence the transcription of a core set of genes encoding a glycine cleavage system, serine hydroxymethyltransferase, and serine dehydratase. PA2449 also affected the transcription of several genes that are integral in cell signaling and pyocyanin biosynthesis in P. aeruginosa PAO1. This study sheds light on the unexpected relationship between the utilization of an unfavorable carbon source and the production of pyocyanin. PA2449 is conserved among pseudomonads and might be universally involved in the assimilation of glycine among this metabolically diverse group of bacteria.

Lundgren, Benjamin R.; Thornton, William; Dornan, Mark H.; Villegas-Penaranda, Luis Roberto; Boddy, Christopher N.

2013-01-01

150

[Substantiation studies for a method of enriching sour milk products with essential fatty acids].  

PubMed

Comparative investigations were conducted to produce arguments in favour of technology employed in preparing special acidified dairy products with an elevated proportion of polyunsaturated essential fatty acids for baby and dietetic nutrition. These investigations included a study into oxidative changes occurring in the vegetable oil during preparation of acidified milk by following two technological patterns that provide for introduction of the vegetable oil into the milk prior to its souring and into sour milk. PMID:580662

Pokrovski?, A A; Korobkina, G S; Brents, M Ia; Biriukova, Z A; Kurbatova, N Ia

151

Genes essential to iron transport in the cyanobacterium Synechocystis sp. strain PCC 6803.  

PubMed

Genes encoding polypeptides of an ATP binding cassette (ABC)-type ferric iron transporter that plays a major role in iron acquisition in Synechocystis sp. strain PCC 6803 were identified. These genes are slr1295, slr0513, slr0327, and recently reported sll1878 (Katoh et al., J. Bacteriol. 182:6523-6524, 2000) and were designated futA1, futA2, futB, and futC, respectively, for their involvement in ferric iron uptake. Inactivation of these genes individually or futA1 and futA2 together greatly reduced the activity of ferric iron uptake in cells grown in complete medium or iron-deprived medium. All the fut genes are expressed in cells grown in complete medium, and expression was enhanced by iron starvation. The futA1 and futA2 genes appear to encode periplasmic proteins that play a redundant role in iron binding. The deduced products of futB and futC genes contain nucleotide-binding motifs and belong to the ABC transporter family of inner-membrane-bound and membrane-associated proteins, respectively. These results and sequence similarities among the four genes suggest that the Fut system is related to the Sfu/Fbp family of iron transporters. Inactivation of slr1392, a homologue of feoB in Escherichia coli, greatly reduced the activity of ferrous iron transport. This system is induced by intracellular low iron concentrations that are achieved in cells exposed to iron-free medium or in the fut-less mutants grown in complete medium. PMID:11292796

Katoh, H; Hagino, N; Grossman, A R; Ogawa, T

2001-05-01

152

Early estrogen-induced gene 1, a novel RANK signaling component, is essential for osteoclastogenesis.  

PubMed

The receptor activator of NF-?B (RANK) and immunoreceptor tyrosine-based activation motif (ITAM)-containing adaptors are essential factors involved in regulating osteoclast formation and bone remodeling. Here, we identify early estrogen-induced gene 1 (EEIG1) as a novel RANK ligand (RANKL)-inducible protein that physically interacts with RANK and further associates with Gab2, PLC?2 and Tec/Btk kinases upon RANKL stimulation. EEIG1 positively regulates RANKL-induced osteoclast formation, likely due to its ability to facilitate RANKL-stimulated PLC?2 phosphorylation and NFATc1 induction. In addition, an inhibitory peptide designed to block RANK-EEIG1 interaction inhibited RANKL-induced bone destruction by reducing osteoclast formation. Together, our results identify EEIG1 as a novel RANK signaling component controlling RANK-mediated osteoclast formation, and suggest that targeting EEIG1 might represent a new therapeutic strategy for the treatment of pathological bone resorption. PMID:23478294

Choi, Han Kyoung; Kang, Hye Ri; Jung, Eutteum; Kim, Tae Eon; Lin, Jing Jing; Lee, Soo Young

2013-03-12

153

Human PTCHD3 nulls: rare copy number and sequence variants suggest a non-essential gene  

PubMed Central

Background Copy number variations (CNVs) can contribute to variable degrees of fitness and/or disease predisposition. Recent studies show that at least 1% of any given genome is copy number variable when compared to the human reference sequence assembly. Homozygous deletions (or CNV nulls) that are found in the normal population are of particular interest because they may serve to define non-essential genes in human biology. Results In a genomic screen investigating CNV in Autism Spectrum Disorders (ASDs) we detected a heterozygous deletion on chromosome 10p12.1, spanning the Patched-domain containing 3 (PTCHD3) gene, at a frequency of ~1.4% (6/427). This finding seemed interesting, given recent discoveries on the role of another Patched-domain containing gene (PTCHD1) in ASD. Screening of another 177 ASD probands yielded two additional heterozygous deletions bringing the frequency to 1.3% (8/604). The deletion was found at a frequency of ~0.73% (27/3,695) in combined control population from North America and Northern Europe predominately of European ancestry. Screening of the human genome diversity panel (HGDP-CEPH) covering worldwide populations yielded deletions in 7/1,043 unrelated individuals and those detected were confined to individuals of European/Mediterranean/Middle Eastern ancestry. Breakpoint mapping yielded an identical 102,624 bp deletion in all cases and controls tested, suggesting a common ancestral event. Interestingly, this CNV occurs at a break of synteny between humans and mouse. Considering all data, however, no significant association of these rare PTCHD3 deletions with ASD was observed. Notwithstanding, our RNA expression studies detected PTCHD3 in several tissues, and a novel shorter isoform for PTCHD3 was characterized. Expression in transfected COS-7 cells showed PTCHD3 isoforms colocalize with calnexin in the endoplasmic reticulum. The presence of a patched (Ptc) domain suggested a role for PTCHD3 in various biological processes mediated through the Hedgehog (Hh) signaling pathway. However, further investigation yielded one individual harboring a homozygous deletion (PTCHD3 null) without ASD or any other overt abnormal phenotype. Exon sequencing of PTCHD3 in other individuals with deletions revealed compound point mutations also resulting in a null state. Conclusion Our data suggests that PTCHD3 may be a non-essential gene in some humans and characterization of this novel CNV at 10p12.1 will facilitate population and disease studies.

2011-01-01

154

The Arabidopsis COW1 gene encodes a phosphatidylinositol transfer protein essential for root hair tip growth.  

PubMed

Root hairs are a major site for the uptake of water and nutrients into plants, and they form an increasingly important model system for the study of development in higher plants. We now report on the molecular genetic analysis of the srh1 mutant in Arabidopsis thaliana impaired in root hair tip growth. We show that srh1 is a new allele of cow1 (can of worms1) and we identified the COW1 gene using a positional cloning strategy. The N-terminus of the COW1 protein is 32% identical to an essential phosphatidylinositol transfer protein (PITP), the yeast Sec14 protein (sec14p) while the C-terminus is 34.5% identical to a late nodulin of Lotus japonicus, Nlj16. We show that expression of the COW1 lipid-binding domain complements the growth defect associated with Sec14p dysfunction in yeast. In addition, we show that GFP fused to the COW1 protein specifically accumulates at the site of root hair outgrowth. We conclude that the COW1 protein is a PITP, essential for proper root hair growth. PMID:15546352

Böhme, Karen; Li, Yong; Charlot, Florence; Grierson, Claire; Marrocco, Katia; Okada, Kyotaka; Laloue, Michel; Nogué, Fabien

2004-12-01

155

The TgsGP Gene Is Essential for Resistance to Human Serum in Trypanosoma brucei gambiense  

PubMed Central

Trypanosoma brucei gambiense causes 97% of all cases of African sleeping sickness, a fatal disease of sub-Saharan Africa. Most species of trypanosome, such as T. b. brucei, are unable to infect humans due to the trypanolytic serum protein apolipoprotein-L1 (APOL1) delivered via two trypanosome lytic factors (TLF-1 and TLF-2). Understanding how T. b. gambiense overcomes these factors and infects humans is of major importance in the fight against this disease. Previous work indicated that a failure to take up TLF-1 in T. b. gambiense contributes to resistance to TLF-1, although another mechanism is required to overcome TLF-2. Here, we have examined a T. b. gambiense specific gene, TgsGP, which had previously been suggested, but not shown, to be involved in serum resistance. We show that TgsGP is essential for resistance to lysis as deletion of TgsGP in T. b. gambiense renders the parasites sensitive to human serum and recombinant APOL1. Deletion of TgsGP in T. b. gambiense modified to uptake TLF-1 showed sensitivity to TLF-1, APOL1 and human serum. Reintroducing TgsGP into knockout parasite lines restored resistance. We conclude that TgsGP is essential for human serum resistance in T. b. gambiense.

DeJesus, Eric; Kieft, Rudo; Hajduk, Stephen; Veitch, Nicola; Steketee, Pieter C.; Cooper, Anneli; Weir, William; MacLeod, Annette

2013-01-01

156

Characterization of an SNR gene locus in Saccharomyces cerevisiae that specifies both dispensible and essential small nuclear RNAs  

SciTech Connect

A genetic locus is described that specifies two Saccharomyces cerevisiae small nuclear RNAs (snRNAs). The genes specifying the two snRNAs are separated by only 67 base pairs and are transcribed in the same direction. The product RNAs contain 128 and 190 nucleotides and are designated snR128 and snR190, respectively. These RNAs resemble snRNAs of other eucaryotes in nuclear localization and possession of a 5' trimethylguanosine cap. Neither snRNA is related in sequence to previously described vertebrate or yeast snRNAs. Both RNAs exhibit properties consistent with nucleolar organization and hydrogen bonding to pre-rRNA species, suggesting possible roles in ribosome biogenesis. The snR128 species cosediments with deproteinized 27S pre-rRNA, whereas snR190 is associated with a 20S intermediate. Gene disruption in vitro followed by replacement of the chromosomal alleles reveals that SNR128 is essential, whereas SNR190 is not.

Zagorski, J.; Tollervey, D.; Fournier, M.J.

1988-08-01

157

Isolation and characterization of SYN1, a RAD21-like gene essential for meiosis in Arabidopsis.  

PubMed Central

The proper pairing, recombination, and segregation of chromosomes are central to meiosis and sexual reproduction. The syn1 mutation was previously identified as a synaptic mutant in a T-DNA-tagged population of plants. SYN1 has been isolated and found to exhibit similarity to Schizosaccharomyces pombe RAD21 and RAD21-like proteins, which are required for chromosome condensation and sister chromatid cohesion during mitosis. Plants homozygous for syn1 are male and female sterile and show defects in chromosome condensation and pairing beginning at leptonema of meiosis I. Fragmentation of the chromosomes was observed at metaphase I. Alternative promoters produced two SYN1 transcripts. One transcript was expressed at low levels in most tissues, whereas the other was expressed only in prebolting buds. DNA blot analyses suggest that Arabidopsis contains a small RAD21 gene family. Consistent with the DNA blot data, a second Arabidopsis RAD21-like gene has been identified. These results suggest that different RAD21-like proteins play essential roles in chromosome condensation and pairing during both meiosis and mitosis.

Bai, X; Peirson, B N; Dong, F; Xue, C; Makaroff, C A

1999-01-01

158

Arabidopsis FAD2 gene encodes the enzyme that is essential for polyunsaturated lipid synthesis.  

PubMed Central

The polyunsaturated fatty acids linoleate and alpha-linolenate are important membrane components and are the essential fatty acids of human nutrition. The major enzyme responsible for the synthesis of these compounds is the plant oleate desaturase of the endoplasmic reticulum, and its activity is controlled in Arabidopsis by the fatty acid desaturation 2 (fad2) locus. A fad2 allele was identified in a population of Arabidopsis in which mutations had been created by T-DNA insertions. Genomic DNA flanking the T-DNA was cloned by plasmid rescue and used to isolate cDNA and genomic clones of FAD2. A cDNA containing the entire FAD2 coding sequence was expressed in fad2 mutant plants and shown to complement the mutant fatty acid phenotype. The deduced amino acid sequence from the cDNA showed homology to other plant desaturases, and this confirmed that FAD2 is the structural gene for the desaturase. Gel blot analyses of FAD2 mRNA levels showed that the gene is expressed throughout the plant and suggest that transcript levels are in excess of the amount needed to account for oleate desaturation. Sequence analysis identified histidine-rich motifs that could contribute to an iron binding site in the cytoplasmic domain of the protein. Such a position would facilitate interaction between the desaturase and cytochrome b5, which is the direct source of electrons for the desaturation reaction, but would limit interaction of the active site with the fatty acyl substrate.

Okuley, J; Lightner, J; Feldmann, K; Yadav, N; Lark, E; Browse, J

1994-01-01

159

The Ornithine Decarboxylase Gene Is Essential for Cell Survival during Early Murine Development  

PubMed Central

Overexpression and inhibitor studies have suggested that the c-Myc target gene for ornithine decarboxylase (ODC), the enzyme which converts ornithine to putrescine, plays an important role in diverse biological processes, including cell growth, differentiation, transformation, and apoptosis. To explore the physiological function of ODC in mammalian development, we generated mice harboring a disrupted ODC gene. ODC-heterozygous mice were viable, normal, and fertile. Although zygotic ODC is expressed throughout the embryo prior to implantation, loss of ODC did not block normal development to the blastocyst stage. Embryonic day E3.5 ODC-deficient embryos were capable of uterine implantation and induced maternal decidualization yet failed to develop substantially thereafter. Surprisingly, analysis of ODC-deficient blastocysts suggests that loss of ODC does not affect cell growth per se but rather is required for survival of the pluripotent cells of the inner cell mass. Therefore, ODC plays an essential role in murine development, and proper homeostasis of polyamine pools appears to be required for cell survival prior to gastrulation.

Pendeville, Helene; Carpino, Nick; Marine, Jean-Christophe; Takahashi, Yutaka; Muller, Marc; Martial, Joseph A.; Cleveland, John L.

2001-01-01

160

The you Gene Encodes an EGF-CUB Protein Essential for Hedgehog Signaling in Zebrafish  

PubMed Central

Hedgehog signaling is required for many aspects of development in vertebrates and invertebrates. Misregulation of the Hedgehog pathway causes developmental abnormalities and has been implicated in certain types of cancer. Large-scale genetic screens in zebrafish have identified a group of mutations, termed you-class mutations, that share common defects in somite shape and in most cases disrupt Hedgehog signaling. These mutant embryos exhibit U-shaped somites characteristic of defects in slow muscle development. In addition, Hedgehog pathway mutations disrupt spinal cord patterning. We report the positional cloning of you, one of the original you-class mutations, and show that it is required for Hedgehog signaling in the development of slow muscle and in the specification of ventral fates in the spinal cord. The you gene encodes a novel protein with conserved EGF and CUB domains and a secretory pathway signal sequence. Epistasis experiments support an extracellular role for You upstream of the Hedgehog response mechanism. Analysis of chimeras indicates that you mutant cells can appropriately respond to Hedgehog signaling in a wild-type environment. Additional chimera analysis indicates that wild-type you gene function is not required in axial Hedgehog-producing cells, suggesting that You is essential for transport or stability of Hedgehog signals in the extracellular environment. Our positional cloning and functional studies demonstrate that You is a novel extracellular component of the Hedgehog pathway in vertebrates.

2005-01-01

161

The you gene encodes an EGF-CUB protein essential for Hedgehog signaling in zebrafish.  

PubMed

Hedgehog signaling is required for many aspects of development in vertebrates and invertebrates. Misregulation of the Hedgehog pathway causes developmental abnormalities and has been implicated in certain types of cancer. Large-scale genetic screens in zebrafish have identified a group of mutations, termed you-class mutations, that share common defects in somite shape and in most cases disrupt Hedgehog signaling. These mutant embryos exhibit U-shaped somites characteristic of defects in slow muscle development. In addition, Hedgehog pathway mutations disrupt spinal cord patterning. We report the positional cloning of you, one of the original you-class mutations, and show that it is required for Hedgehog signaling in the development of slow muscle and in the specification of ventral fates in the spinal cord. The you gene encodes a novel protein with conserved EGF and CUB domains and a secretory pathway signal sequence. Epistasis experiments support an extracellular role for You upstream of the Hedgehog response mechanism. Analysis of chimeras indicates that you mutant cells can appropriately respond to Hedgehog signaling in a wild-type environment. Additional chimera analysis indicates that wild-type you gene function is not required in axial Hedgehog-producing cells, suggesting that You is essential for transport or stability of Hedgehog signals in the extracellular environment. Our positional cloning and functional studies demonstrate that You is a novel extracellular component of the Hedgehog pathway in vertebrates. PMID:15660164

Woods, Ian G; Talbot, William S

2005-03-01

162

Cloning of the RNA8 gene of Saccharomyces cerevisiae, detection of the RNA8 protein, and demonstration that it is essential for nuclear pre-mRNA splicing  

SciTech Connect

Strains of Saccharomyces cerevisiae that bear the temperature-sensitive mutation rna8-1 are defective in nuclear pre-mRNA splicing at the restrictive temperature (36/sup 0/C), suggesting that the RNA8 gene encodes a component of the splicing machinery. The RNA8 gene was cloned by complementation of the temperature-sensitive growth defect of an rna8-1 mutant strain. Integrative transformation and gene disruption experiments confirmed the identify of the cloned DNA and demonstrated that the RNA8 gene encodes an essential function. The RNA8 gene was shown to be represented once per S. cerevisia haploid genome and to encode a low-abundance transcript of approximately 7.4 kilobases. By using antisera raised against BETA-galactosidase-RNA8 fusion proteins, the RNA8 gene product was identified in S. cerevisiae cell extracts as a low-abundance protein of approximately 260 kilodaltons. Immunodepletion of the RNA8 protein specifically abolished the activity of S. cerevisia in vitro splicing extract, confirming that RNA8 plays an essential role in splicing.

Jackson, S.P.; Lossky, M.; Beggs, J.D.

1988-03-01

163

The Fusarium graminearum MAP1 gene is essential for pathogenicity and development of perithecia.  

PubMed

SUMMARY Fusarium graminearum is the causal agent of ear blight disease of cereals. Infection occurs at anthesis when ascospores and/or conidia directly penetrate exposed anther and ovary tissue. The hemibiotrophic hyphae colonize floral tissues and developing grains to cause premature ear senescence. During infection, Fusarium hyphae can also produce hazardous trichothecene mycotoxins, thereby posing a threat to human and animal health and safety. The Fusarium MAP1 gene was identified using a PCR approach by its homology to a known pathogenicity gene of Magnaporthe grisea, the mitogen-activated protein kinase gene PMK1. Gene replacement F. graminearum map1 mutants were non-pathogenic on wheat flowers and roots, and also could not infect wounded wheat floral tissue or tomato fruits. Unlike the wild-type strain, map1 mutant inoculations did not compromise grain yield. Map1 mutants lost their ability to form perithecia in vitro, but their rate of asexual conidiation was unaffected. DON mycotoxin production in planta was still detected. Collectively, the observed phenotypes suggest that the Map1 signalling protein controls multiple events in disease establishment and propagation. Novel approaches to control Fusarium ear blight disease by blocking perithecial development are discussed. PMID:20569395

Urban, Martin; Mott, Ellie; Farley, Tom; Hammond-Kosack, Kim

2003-09-01

164

The gene for a tRNA modifying enzyme, m5U54-methyltransferase, is essential for viability in Escherichia coli.  

PubMed Central

One of the most abundant modified nucleosides in tRNA is 5-methyluridine (m5U or rT, ribothymidine). The enzyme tRNA(m5U54)methyltransferase [S-adenosyl-L-methionine:tRNA (uracil-5-)-methyltransferase, EC 2.1.1.35] (the trmA gene product) catalyzes S-adenosylmethionine-dependent methylation of the uracil in position 54 (T psi C loop) in all Escherichia coli tRNAs to form m5U. Hitherto no modified nucleoside in tRNA has been shown to be essential for growth, although their importance in fine tuning the function of tRNA is well established. In this paper, we show that the structural gene trmA is essential for viability, although the known catalytic activity of the tRNA(m5U54)methyltransferase is not. Images

Persson, B C; Gustafsson, C; Berg, D E; Bjork, G R

1992-01-01

165

The Haemophilus influenzae adenylate cyclase gene: cloning, sequence, and essential role in competence.  

PubMed Central

Competence for transformation in Haemophilus influenzae is stimulated by cyclic AMP (cAMP) and requires the cAMP-dependent catabolite regulatory protein CRP. Thus, understanding the control of competence will require understanding how cAMP levels are regulated. As a first step, we have cloned the H. influenzae adenylate cyclase gene (cya) by complementing the Lac- phenotype of delta cya Escherichia coli. Its sequence specifies an 843-amino-acid protein which has significant identity to other known bacterial adenylate cyclases (41 to 43% and 61% identical to the cya genes of enteric bacteria and of Pasteurella multocida, respectively). As seen in other bacterial cya genes, there is evidence for regulation similar to that demonstrated for E. coli: the presence of a strong consensus CRP binding site within the promoter of the gene may provide feedback control of cAMP levels by repressing cya transcription, and translation may be limited by the weak ribosome binding site and by initiation of protein synthesis with GUG rather than AUG or the UUG used in other bacterial cya genes. We confirmed the essential role of cAMP in competence by constructing and characterizing H. influenzae cya mutants. This strain failed to develop competence either spontaneously or after transfer to a competence-inducing medium. However, it became as competent as its wild-type parent in the presence of exogenous cAMP. This result suggests that the failure of exogenously added cAMP to induce optimum competence in wild-type cells is not due to a limitation to the entry of cAMP into the cells. Rather, it strongly favors models in which competence induction requires both an increase in intracellular cAMP and a second as yet unidentified regulatory event. H. influenzae strains mutant in cya or crp were unable to ferment xylose or ribose. This confirms that influenzae, like E. coli, uses cAMP and CRP to regulate nutrient uptake and utilization and lends increasing support to the hypothesis that DNA uptake is mechanism of nutrient acquisition.

Dorocicz, I R; Williams, P M; Redfield, R J

1993-01-01

166

Vapyrin, a gene essential for intracellular progression of arbuscular mycorrhizal symbiosis, is also essential for infection by rhizobia in the nodule symbiosis of Medicago truncatula.  

PubMed

Intracellular invasion of root cells is required for the establishment of successful endosymbioses in legumes of both arbuscular mycorrhizal (AM) fungi and rhizobial bacteria. In both interactions a requirement for successful entry is the activation of a common signalling pathway that includes five genes required to generate calcium oscillations and two genes required for the perception of the calcium response. Recently, it has been discovered that in Medicago truncatula, the Vapyrin (VPY) gene is essential for the establishment of the arbuscular mycorrhizal symbiosis. Here, we show by analyses of mutants that the same gene is also required for rhizobial colonization and nodulation. VPY encodes a protein featuring a Major Sperm Protein domain, typically featured on proteins involved in membrane trafficking and biogenesis, and a series of ankyrin repeats. Plants mutated in this gene have abnormal rhizobial infection threads and fewer nodules, and in the case of interactions with AM fungi, epidermal penetration defects and aborted arbuscule formation. Calcium spiking in root hairs in response to supplied Nod factors is intact in the vpy-1 mutant. This, and the elevation of VPY transcripts upon application of Nod factors which we show to be dependent on NFP, DMI1, and DMI3, indicates that VPY acts downstream of the common signalling pathway. PMID:21223389

Murray, Jeremy D; Muni, RajaSekhara Reddy Duvvuru; Torres-Jerez, Ivone; Tang, Yuhong; Allen, Stacy; Andriankaja, Megan; Li, Guangming; Laxmi, Ashverya; Cheng, Xiaofei; Wen, Jiangqi; Vaughan, David; Schultze, Michael; Sun, Jongho; Charpentier, Myriam; Oldroyd, Giles; Tadege, Million; Ratet, Pascal; Mysore, Kirankumar S; Chen, Rujin; Udvardi, Michael K

2010-11-29

167

Conserved and essential transcription factors for cellulase gene expression in ascomycete fungi  

PubMed Central

Rational engineering of filamentous fungi for improved cellulase production is hampered by our incomplete knowledge of transcriptional regulatory networks. We therefore used the model filamentous fungus Neurospora crassa to search for uncharacterized transcription factors associated with cellulose deconstruction. A screen of a N. crassa transcription factor deletion collection identified two uncharacterized zinc binuclear cluster transcription factors (clr-1 and clr-2) that were required for growth and enzymatic activity on cellulose, but were not required for growth or hemicellulase activity on xylan. Transcriptional profiling with next-generation sequencing methods refined our understanding of the N. crassa transcriptional response to cellulose and demonstrated that clr-1 and clr-2 were required for the bulk of that response, including induction of all major cellulase and some major hemicellulase genes. Functional CLR-1 was necessary for expression of clr-2 and efficient cellobiose utilization. Phylogenetic analyses showed that CLR-1 and CLR-2 are conserved in the genomes of most filamentous ascomycete fungi capable of degrading cellulose. In Aspergillus nidulans, a strain carrying a deletion of the clr-2 homolog (clrB) failed to induce cellulase gene expression and lacked cellulolytic activity on Avicel. Further manipulation of this control system in industrial production strains may significantly improve yields of cellulases for cellulosic biofuel production.

Coradetti, Samuel T.; Craig, James P.; Xiong, Yi; Shock, Teresa; Tian, Chaoguang; Glass, N. Louise

2012-01-01

168

Identification of a Gene Essential for the First Committed Step in the Biosynthesis of Bacteriochlorophyll c*  

PubMed Central

Bacteriochlorophylls (BChls) c, d, and e are the major chlorophylls in chlorosomes, which are the largest and one of the most efficient antennae produced by chlorophototrophic organisms. In the biosynthesis of these three BChls, a C-132-methylcarboxyl group found in all other chlorophylls (Chls) must be removed. This reaction is postulated to be the first committed step in the synthesis of these BChls. Analyses of gene neighborhoods of (B)Chl biosynthesis genes and distribution patterns in organisms producing chlorosomes helped to identify a gene (bciC) that appeared to be a good candidate to produce the enzyme involved in this biochemical reaction. To confirm that this was the case, a deletion mutant of an open reading frame orthologous to bciC, CT1077, was constructed in Chlorobaculum tepidum, a genetically tractible green sulfur bacterium. The CT1077 deletion mutant was unable to synthesize BChl c but still synthesized BChl a and Chl a. The deletion mutant accumulated large amounts of various (bacterio)pheophorbides, all of which still had C-132-methylcarboxyl groups. A C. tepidum strain in which CT1077 was replaced by an orthologous gene, Cabther_B0031 from “Candidatus Chloracidobacterium thermophilum” was constructed. Although the product of Cabther_B0031 was only 28% identical to the product of CT1077, this strain synthesized BChl c, BChl a, and Chl a in amounts similar to wild-type C. tepidum cells. To indicate their roles in the first committed step of BChl c, d, and e biosynthesis, open reading frames CT1077 and Cabther_B0031 have been redesignated bciC. The potential mechanism by which BciC removes the C-132-methylcarboxyl moiety of chlorophyllide a is discussed.

Liu, Zhenfeng; Bryant, Donald A.

2011-01-01

169

Functional Analysis of Epstein-Barr Virus SM Protein: Identification of Amino Acids Essential for Structure, Transactivation, Splicing Inhibition, and Virion Production  

PubMed Central

The Epstein-Barr virus (EBV) SM protein is a posttranscriptional regulator of cellular and viral gene expression that binds and stabilizes target mRNAs and shuttles from nucleus to cytoplasm. SM enhances expression of several EBV genes required for lytic replication and is essential for virion production. SM increases accumulation of specific mRNAs but also inhibits expression of several intron-containing transcripts. The mechanism by which SM inhibits gene expression is poorly understood. The experiments described here had several aims: to determine whether specific domains of SM were responsible for activation or inhibition function; whether these functions could be separated; and whether one or more of these functions were essential for virion production. A mutational analysis of SM was performed, focusing on amino acids in SM that are evolutionarily conserved among SM homologs in other herpesviruses. Mutation of the carboxy-terminal region of SM revealed a region that is likely to be structurally important for SM protein conformation. In addition, several amino acids were identified that are critical for activation and inhibition function. A specific mutation of a highly conserved cysteine residue revealed that it was essential for gene inhibition but not for transactivation, indicating that these two functions operate through independent mechanisms. Furthermore, the ability of wild-type SM and the inability of the mutant to inhibit gene expression were shown to correlate with the ability to inhibit splicing of a human target gene and thereby prevent accumulation of its processed mRNA. Surprisingly, some mutations which preserved both activation and inhibition functions in vitro nevertheless abolished virion production, suggesting that other SM functions or protein-protein interactions are also required for lytic replication.

Ruvolo, Vivian; Sun, Liang; Howard, Karilynn; Sung, Seung; Delecluse, Henri-Jacques; Hammerschmidt, Wolfgang; Swaminathan, Sankar

2004-01-01

170

Bifunctional gene cluster lnqBCDEF mediates bacteriocin production and immunity with differential genetic requirements.  

PubMed

A comprehensive gene disruption of lacticin Q biosynthetic cluster lnqQBCDEF was carried out. The results demonstrated the necessity of the complete set of lnqQBCDEF for lacticin Q production, whereas immunity was flexible, with LnqEF (ABC transporter) being essential for and LnqBCD partially contributing to immunity. PMID:23335763

Iwatani, Shun; Horikiri, Yuko; Zendo, Takeshi; Nakayama, Jiro; Sonomoto, Kenji

2013-01-18

171

Bifunctional Gene Cluster lnqBCDEF Mediates Bacteriocin Production and Immunity with Differential Genetic Requirements  

PubMed Central

A comprehensive gene disruption of lacticin Q biosynthetic cluster lnqQBCDEF was carried out. The results demonstrated the necessity of the complete set of lnqQBCDEF for lacticin Q production, whereas immunity was flexible, with LnqEF (ABC transporter) being essential for and LnqBCD partially contributing to immunity.

Iwatani, Shun; Horikiri, Yuko; Zendo, Takeshi; Nakayama, Jiro

2013-01-01

172

SuhB Is a Regulator of Multiple Virulence Genes and Essential for Pathogenesis of Pseudomonas aeruginosa  

PubMed Central

ABSTRACT During initial colonization and chronic infection, pathogenic bacteria encounter distinct host environments. Adjusting gene expression accordingly is essential for the pathogenesis. Pseudomonas aeruginosa has evolved complicated regulatory networks to regulate different sets of virulence factors to facilitate colonization and persistence. The type III secretion system (T3SS) and motility are associated with acute infections, while biofilm formation and the type VI secretion system (T6SS) are associated with chronic persistence. To identify novel regulatory genes required for pathogenesis, we screened a P. aeruginosa transposon (Tn) insertion library and found suhB to be an essential gene for the T3SS gene expression. The expression of suhB was upregulated in a mouse acute lung infection model, and loss of suhB resulted in avirulence. Suppression of T3SS gene expression in the suhB mutant is linked to a defective translation of the T3SS master regulator, ExsA. Further studies demonstrated that suhB mutation led to the upregulation of GacA and its downstream small RNAs, RsmY and RsmZ, triggering T6SS expression and biofilm formation while inhibiting the T3SS. Our results demonstrate that an in vivo-inducible gene, suhB, reciprocally regulates genes associated with acute and chronic infections and plays an essential role in the pathogenesis of P. aeruginosa.

Li, Kewei; Xu, Chang; Jin, Yongxin; Sun, Ziyu; Liu, Chang; Shi, Jing; Chen, Gukui; Chen, Ronghao; Jin, Shouguang; Wu, Weihui

2013-01-01

173

Analysis of Sequence, Map Position, and Gene Expression Reveals Conserved Essential Genes for Iron Uptake in Arabidopsis and Tomato1[w  

PubMed Central

Arabidopsis (Arabidopsis thaliana) and tomato (Lycopersicon esculentum) show similar physiological responses to iron deficiency, suggesting that homologous genes are involved. Essential gene functions are generally considered to be carried out by orthologs that have remained conserved in sequence and map position in evolutionarily related species. This assumption has not yet been proven for plant genomes that underwent large genome rearrangements. We addressed this question in an attempt to deduce functional gene pairs for iron reduction, iron transport, and iron regulation between Arabidopsis and tomato. Iron uptake processes are essential for plant growth. We investigated iron uptake gene pairs from tomato and Arabidopsis, namely sequence, conserved gene content of the regions containing iron uptake homologs based on conserved orthologous set marker analysis, gene expression patterns, and, in two cases, genetic data. Compared to tomato, the Arabidopsis genome revealed more and larger gene families coding for the iron uptake functions. The number of possible homologous pairs was reduced if functional expression data were taken into account in addition to sequence and map position. We predict novel homologous as well as partially redundant functions of ferric reductase-like and iron-regulated transporter-like genes in Arabidopsis and tomato. Arabidopsis nicotianamine synthase genes encode a partially redundant family. In this study, Arabidopsis gene redundancy generally reflected the presumed genome duplication structure. In some cases, statistical analysis of conserved gene regions between tomato and Arabidopsis suggested a common evolutionary origin. Although involvement of conserved genes in iron uptake was found, these essential genes seem to be of paralogous rather than orthologous origin in tomato and Arabidopsis.

Bauer, Petra; Thiel, Thomas; Klatte, Marco; Bereczky, Zsolt; Brumbarova, Tzvetina; Hell, Rudiger; Grosse, Ivo

2004-01-01

174

The Unconserved Groucho Central Region Is Essential for Viability and Modulates Target Gene Specificity  

PubMed Central

Groucho (Gro) is a Drosophila corepressor required by numerous DNA-binding repressors, many of which are distributed in gradients and provide positional information during development. Gro contains well-conserved domains at its N- and C-termini, and a poorly conserved central region that includes the GP, CcN, and SP domains. All lethal point mutations in gro map to the conserved regions, leading to speculation that the unconserved central domains are dispensable. However, our sequence analysis suggests that the central domains are disordered leading us to suspect that the lack of lethal mutations in this region reflects a lack of order rather than an absence of essential functions. In support of this conclusion, genomic rescue experiments with Gro deletion variants demonstrate that the GP and CcN domains are required for viability. Misexpression assays using these same deletion variants show that the SP domain prevents unrestrained and promiscuous repression by Gro, while the GP and CcN domains are indispensable for repression. Deletion of the GP domain leads to loss of nuclear import, while deletion of the CcN domain leads to complete loss of repression. Changes in Gro activity levels reset the threshold concentrations at which graded repressors silence target gene expression. We conclude that co-regulators such as Gro are not simply permissive components of the repression machinery, but cooperate with graded DNA-binding factors in setting borders of gene expression. We suspect that disorder in the Gro central domains may provide the flexibility that allows this region to mediate multiple interactions required for repression.

Turki-Judeh, Wiam; Courey, Albert J.

2012-01-01

175

The Histone-Like Protein Hlp Is Essential for Growth of Streptococcus pyogenes: Comparison of Genetic Approaches To Study Essential Genes?†  

PubMed Central

Selection of possible targets for vaccine and drug development requires an understanding of the physiology of bacterial pathogens, for which the ability to manipulate expression of essential genes is critical. For Streptococcus pyogenes (the group A streptococcus [GAS]), an important human pathogen, the lack of genetic tools for such studies has seriously hampered research. To address this problem, we characterized variants of the inducible Ptet cassette, in both sense and antisense contexts, as tools to regulate transcription from GAS genes. We found that although the three-operator Ptet construct [Ptet(O)3] had low uninduced expression, its induction level was low, while the two-operator construct [Ptet(O)2] was inducible to a high level but showed significant constitutive expression. Use of Ptet(O)3 in the chromosome allowed us to demonstrate previously that RNases J1 and J2 are required for growth of GAS. Here we report that the uninduced level from the chromosomally inserted Ptet(O)2 construct was too high for us to observe differential growth. For the highly expressed histone-like protein (Hlp) of GAS, neither chromosomal insertion of Ptet(O)2 or Ptet(O)3 nor their use on a high-copy-number plasmid to produce antisense RNA specific to hlp resulted in adequate differential expression. However, by replacing the ribosome binding site of hlp with an engineered riboswitch to control translation of Hlp, we demonstrated for the first time that this protein is essential for GAS growth.

Bugrysheva, Julia V.; Froehlich, Barbara J.; Freiberg, Jeffrey A.; Scott, June R.

2011-01-01

176

The vif gene is essential for efficient replication of caprine arthritis encephalitis virus in goat synovial membrane cells and affects the late steps of the virus replication cycle.  

PubMed Central

Complex retrovirus genomes contain a variable number of accessory genes, among which is the vif gene. We investigated in vitro the role of the vif gene of caprine arthritis encephalitis virus (CAEV) by studying the phenotype of five vif mutants after infection of primary goat synovial membrane (GSM) cells and blood-derived monocytes/macrophages. Any deletion introduced into the vif gene resulted in slow and low viral replication and production of virions with an infectious titer lower than that of wild-type viral particles. The wild-type phenotype could be restored by the trans expression of the vif gene in a complementation assay. Quantitative PCR and reverse transcription-PCR analyses were performed in order to determine which stage of the replicative cycle was impaired by the vif deletion. Our results demonstrated that CAEV Vif did not act at the level of reverse transcription or transcription but rather at the late stage of virus formation and/or release, as lower amounts of virus were produced after a single replicative cycle. The vif-deleted CAEV produced after 24 h of infection was still able to infect GSM cells, indicating that the vif gene is not essential for virus infectivity but is required for efficient virus production.

Harmache, A; Bouyac, M; Audoly, G; Hieblot, C; Peveri, P; Vigne, R; Suzan, M

1995-01-01

177

A Vitamin B12-Based System for Conditional Expression Reveals dksA To Be an Essential Gene in Myxococcus xanthus ?  

PubMed Central

Myxococcus xanthus is a prokaryotic model system for the study of multicellular development and the response to blue light. The previous analyses of these processes and the characterization of new genes would benefit from a robust system for controlled gene expression, which has been elusive so far for this bacterium. Here, we describe a system for conditional expression of genes in M. xanthus based on our recent finding that vitamin B12 and CarH, a MerR-type transcriptional repressor, together downregulate a photoinducible promoter. Using this system, we confirmed that M. xanthus rpoN, encoding ?54, is an essential gene, as reported earlier. We then tested it with ftsZ and dksA. In most bacteria, ftsZ is vital due to its role in cell division, whereas null mutants of dksA, whose product regulates the stringent response via transcriptional control of rRNA and amino acid biosynthesis promoters, are viable but cause pleiotropic effects. As with rpoN, it was impossible to delete endogenous ftsZ or dksA in M. xanthus except in a merodiploid background carrying another functional copy, which indicates that these are essential genes. B12-based conditional expression of ftsZ was insufficient to provide the high intracellular FtsZ levels required. With dksA, as with rpoN, cells were viable under permissive but not restrictive conditions, and depletion of DksA or ?54 produced filamentous, aberrantly dividing cells. dksA thus joins rpoN in a growing list of genes dispensable in many bacteria but essential in M. xanthus.

Garcia-Moreno, Diana; Polanco, Maria Carmen; Navarro-Aviles, Gloria; Murillo, Francisco J.; Padmanabhan, S.; Elias-Arnanz, Montserrat

2009-01-01

178

The zebrafish detour gene is essential for cranial but not spinal motor neuron induction.  

PubMed

The zebrafish detour (dtr) mutation generates a novel neuronal phenotype. In dtr mutants, most cranial motor neurons, especially the branchiomotor, are missing. However, spinal motor neurons are generated normally. The loss of cranial motor neurons is not due to aberrant hindbrain patterning, failure of neurogenesis, increased cell death or absence of hh expression. Furthermore, activation of the Hh pathway, which normally induces branchiomotor neurons, fails to induce motor neurons in the dtr hindbrain. Despite this, not all Hh-mediated regulation of hindbrain development is abolished since the regulation of a neural gene by Hh is intact in the dtr hindbrain. Finally, dtr can function cell autonomously to induce branchiomotor neurons. These results suggest that detour encodes a component of the Hh signaling pathway that is essential for the induction of motor neurons in the hindbrain but not in the spinal cord and that dtr function is required for the induction of only a subset of Hh-mediated events in the hindbrain. PMID:10331983

Chandrasekhar, A; Schauerte, H E; Haffter, P; Kuwada, J Y

1999-06-01

179

Ewing sarcoma gene EWS is essential for meiosis and B lymphocyte development  

PubMed Central

Ewing sarcoma gene EWS encodes a putative RNA-binding protein with proposed roles in transcription and splicing, but its physiological role in vivo remains undefined. Here, we have generated Ews-deficient mice and demonstrated that EWS is required for the completion of B cell development and meiosis. Analysis of Ews–/– lymphocytes revealed a cell-autonomous defect in precursor B lymphocyte (pre–B lymphocyte) development. During meiosis, Ews-null spermatocytes were deficient in XY bivalent formation and showed reduced meiotic recombination, resulting in massive apoptosis and complete arrest in gamete maturation. Inactivation of Ews in mouse embryonic fibroblasts resulted in premature cellular senescence, and the mutant animals showed hypersensitivity to ionizing radiation. Finally, we showed that EWS interacts with lamin A/C and that loss of EWS results in a reduced lamin A/C expression. Our findings reveal essential functions for EWS in pre–B cell development and meiosis, with proposed roles in DNA pairing and recombination/repair mechanisms. Furthermore, we demonstrate a novel role of EWS in cellular senescence, possibly through its interaction and modulation of lamin A/C.

Li, Hongjie; Watford, Wendy; Li, Cuiling; Parmelee, Alissa; Bryant, Mark A.; Deng, Chuxia; O'Shea, John; Lee, Sean Bong

2007-01-01

180

Moderate expression of Wnt signaling genes is essential for porcine parthenogenetic embryo development.  

PubMed

Parthenogenetic embryos are invariably lost in mid-gestation, possibly due to the lack of the paternal genome and the consequent induction of aberrant gene expression. Wnt signaling is essential for embryonic development; however, the studies of this pathway in porcine parthenogenetic embryos have been limited. Here, the role of Wnt signaling in porcine parthenogenetic embryos was studied. In vivo embryos were used as controls. Single cell quantitative real-time PCR showed that Wnt signaling was down-regulated in porcine parthenogenetic embryos. Furthermore, immunofluorescence staining and real-time PCR demonstrated that porcine parthenogenetic embryo development was largely unaffected by the inhibition of Wnt signaling with IWP-2, but blastocyst hatching and trophectoderm development was blocked. In addition, parthenogenetic blastocyst hatching was improved by the activation of Wnt signaling by BIO. However, the developmental competency of porcine embryos, including blastocyst hatching, was impaired and apoptosis was induced upon the excessive activation of Wnt signaling. These findings constitute novel evidence that Wnt signaling is important for porcine pre-implantation development and that its down-regulation may lead to the low hatching rate of porcine parthenogenetic blastocysts. PMID:23333243

Huang, Yongye; Ouyang, Hongsheng; Xie, Wanhua; Chen, Xianju; Yao, Chaogang; Han, Yang; Han, Xiaolei; Song, Qi; Pang, Daxin; Tang, Xiaochun

2013-01-16

181

Antisense Phosphorodiamidate Morpholino Oligomers Targeted to an Essential Gene Inhibit Burkholderia cepacia complex  

PubMed Central

Background: Members of the Burkholderia cepacia complex (Bcc) cause significant morbidity and mortality in patients with chronic granulomatous disease (CGD) and cystic fibrosis (CF). Many Bcc strains are antibiotic resistant requiring the exploration of novel antimicrobial approaches including antisense technologies, such as phosphorodiamidate morpholino oligomers (PMOs). Methods: Peptide-conjugated PMOs (PPMOs) were developed to target the acpP gene, encoding an acyl carrier protein thought to be essential for growth. Their antimicrobial activities were tested against different strains of Bcc in vitro and in infection models. Results: PPMOs targeting acpP were bactericidal against clinical isolates of Bcc (> 4 log reduction), whereas a PPMO with a scrambled base sequence (Scr) had no effect on growth. Human neutrophils (PMN) were infected with B. multivorans, and treated with AcpP PPMO. AcpP PPMO augmented killing compared to PMN alone ± Scr PPMO. CGD mice infected with B. multivorans were treated with AcpP PPMO, Scr PPMO or water at 0, 3 and 6 hours post-infection. Compared to water treated controls, the AcpP PPMO treated mice showed a ~80% reduction in the risk of dying by day 30 and relatively little pathology. Conclusions: AcpP PPMO is active against Bcc infections in vitro and in vivo.

Greenberg, David E.; Marshall-Batty, Kimberly R.; Brinster, Lauren R.; Zarember, Kol A.; Shaw, Pamela A.; Mellbye, Brett L.; Iversen, Patrick L.; Holland, Steven M.; Geller, Bruce L.

2010-01-01

182

Twist is an essential regulator of the skeletogenic gene regulatory network in the sea urchin embryo  

PubMed Central

Recent work on the sea urchin endomesoderm gene regulatory network (GRN) offers many opportunities to study the specification and differentiation of each cell type during early development at a mechanistic level. The mesoderm lineages consist of two cell populations, primary and secondary mesenchyme cells (PMCs and SMCs). The micromere-PMC GRN governs the development of the larval skeleton, which is the exclusive fate of PMCs, and SMCs diverge into four lineages, each with its own GRN state. Here we identify a sea urchin ortholog of the Twist transcription factor, and show that it plays an essential role in the PMC GRN and later is involved in SMC formation. Perturbations of Twist either by morpholino knockdown or by overexpression result in defects in progressive phases of PMC development, including specification, ingression/EMT, differentiation and skeletogenesis. Evidence is presented that Twist expression is required for the maintenance of the PMC specification state, and a reciprocal regulation between Alx1 and Twist offers stability for the subsequent processes, such as PMC differentiation and skeletogenesis. These data illustrate the significance of regulatory state maintenance and continuous progression during cell specification, and the dynamics of the sequential events that depend on those earlier regulatory states.

Wu, Shu-Yu; Yang, Yu-Ping; McClay, David R.

2008-01-01

183

Influence of Origanum vulgare L. essential oil on enterotoxin production, membrane permeability and surface characteristics of Staphylococcus aureus.  

PubMed

This study evaluated the influence of the essential oil from Origanum vulgare L. on the enterotoxin production, membrane permeability and cell surface characteristics of Staphylococcus aureus. The suppression of enterotoxin production occurred totally in the broth added with the essential oil at subinhibitory concentrations (0.3 and 0.15 microL/mL). Loss of 260-nm-absorbing material and potassium ions occurred immediately after addition of the essential oil at 0.6 and 1.2 microL/mL and followed up to 120 min. Electron microscopy of essential oil-treated cells revealed the formation of roles in the cell surfaces and loss of cytoplasm material. According to these results, O. vulgare essential oil could be rationally applied in food products both to inhibit the growth of S. aureus and to suppress the synthesis of staphylococcal enterotoxins. PMID:20015563

de Souza, Evandro Leite; de Barros, Jefferson Carneiro; de Oliveira, Carlos Eduardo Vasconcelos; da Conceição, Maria Lúcia

2009-12-04

184

Essential Role for IKK? in Production of Type 1 Interferons by Plasmacytoid Dendritic Cells*  

PubMed Central

Plasmacytoid dendritic cells (pDCs) are characterized by their ability to produce high levels of type 1 interferons in response to ligands that activate TLR7 and TLR9, but the signaling pathways required for IFN production are incompletely understood. Here we exploit the human pDC cell line Gen2.2 and improved pharmacological inhibitors of protein kinases to address this issue. We demonstrate that ligands that activate TLR7 and TLR9 require the TAK1-IKK? signaling pathway to induce the production of IFN? via a pathway that is independent of the degradation of I?B?. We also show that IKK? activity, as well as the subsequent IFN?-stimulated activation of the JAK-STAT1/2 signaling pathway, are essential for the production of IFN? by TLR9 ligands. We further show that TLR7 ligands CL097 and R848 fail to produce significant amounts of IFN? because the activation of IKK? is not sustained for a sufficient length of time. The TLR7/9-stimulated production of type 1 IFNs is inhibited by much lower concentrations of IKK? inhibitors than those needed to suppress the production of NF?B-dependent proinflammatory cytokines, such as IL-6, suggesting that drugs that inhibit IKK? may have a potential for the treatment of forms of lupus that are driven by self-RNA and self-DNA-induced activation of TLR7 and TLR9, respectively.

Pauls, Eduardo; Shpiro, Natalia; Peggie, Mark; Young, Erick R.; Sorcek, Ronald J.; Tan, Li; Choi, Hwan Geun; Cohen, Philip

2012-01-01

185

THE ELR MOTIF IN VIL8 GENE OF THE MAREK'S DISEASE VIRUS IS ESSENTIAL FOR THE ANGIOGENESIS ACTIVITY  

Technology Transfer Automated Retrieval System (TEKTRAN)

Marek's disease virus (MDV) encodes a chemokine-like gene, vIL-8, in serotype 1 MDV. VIL8 is not essential for replication in cell culture but is critical for the in vivo replication of MDV in the lymphoid organs. MDV vIL-8 shares significant homology to cellular CXC chemokines such as IL-8 and GRO...

186

A novel class of evolutionarily conserved genes defined by piwi are essential for stem cell self-renewal  

Microsoft Academic Search

Germ-line stem cells (GSCs) serve as the source for gametogenesis in diverse organisms. We cloned and characterized the Drosophila piwi gene and showed that it is required for the asymmetric division of GSCs to produce and maintain a daughter GSC but is not essential for the further differentiation of the committed daughter cell. Genetic mosaic and RNA in situ analyses

Daniel N. Cox; Anna Chao; Jeff Baker; Lisa Chang; Dan Qiao; Haifan Lin

1998-01-01

187

ROS Production Is Essential for the Apoptotic Function of E2F1 in Pheochromocytoma and Neuroblastoma Cell Lines  

PubMed Central

In this study we demonstrate that accumulation of reactive oxygen species (ROS) is essential for E2F1 mediated apoptosis in ER-E2F1 PC12 pheochromocytoma, and SH-SY5Y and SK-N-JD neuroblastoma stable cell lines. In these cells, the ER-E2F1 fusion protein is expressed in the cytosol; the addition of 4-hydroxytamoxifen (OHT) induces its translocation to the nucleus and activation of E2F1target genes. Previously we demonstrated that, in ER-E2F1 PC12 cells, OHT treatment induced apoptosis through activation of caspase-3. Here we show that caspase-8 activity did not change upon treatment with OHT. Moreover, over-expression of Bcl-xL arrested OHT-induced apoptosis; by contrast, over-expression of c-FLIP, did not have any effect on OHT-induced apoptosis. OHT addition induces BimL expression, its translocation to mitochondria and activation of Bax, which is paralleled by diminished mitochondrial enrichment of Bcl-xL. Treatment with a Bax-inhibitory peptide reduced OHT-induced apoptosis. These results point out the essential role of mitochondria on the apoptotic process driven by E2F1. ROS accumulation followed E2F1 induction and treatment with the antioxidant N-acetylcysteine, inhibited E2F1-induced Bax translocation to mitochondria and subsequent apoptosis. The role of ROS in mediating OHT-induced apoptosis was also studied in two neuroblastoma cell lines, SH-SY5Y and SK-N-JD. In SH-SY5Y cells, activation of E2F1 by the addition of OHT induced ROS production and apoptosis, whereas over-expression of E2F1 in SK-N-JD cells failed to induce either response. Transcriptional profiling revealed that many of the genes responsible for scavenging ROS were down-regulated following E2F1-induction in SH-SY5Y, but not in SK-N-JD cells. Finally, inhibition of GSK3? blocked ROS production, Bax activation and the down regulation of ROS scavenging genes. These findings provide an explanation for the apparent contradictory role of E2F1 as an apoptotic agent versus a cell cycle activator.

Espada, Lilia; Meo-Evoli, Nathalie; Sancho, Patricia; Real, Sebastian; Fabregat, Isabel; Ambrosio, Santiago; Tauler, Albert

2012-01-01

188

Transposon Mutagenesis Identified Chromosomal and Plasmid Genes Essential for Adaptation of the Marine Bacterium Dinoroseobacter shibae to Anaerobic Conditions.  

PubMed

Anaerobic growth and survival are integral parts of the life cycle of many marine bacteria. To identify genes essential for the anoxic life of Dinoroseobacter shibae, a transposon library was screened for strains impaired in anaerobic denitrifying growth. Transposon insertions in 35 chromosomal and 18 plasmid genes were detected. The essential contribution of plasmid genes to anaerobic growth was confirmed with plasmid-cured D. shibae strains. A combined transcriptome and proteome approach identified oxygen tension-regulated genes. Transposon insertion sites of a total of 1,527 mutants without an anaerobic growth phenotype were determined to identify anaerobically induced but not essential genes. A surprisingly small overlap of only three genes (napA, phaA, and the Na(+)/Pi antiporter gene Dshi_0543) between anaerobically essential and induced genes was found. Interestingly, transposon mutations in genes involved in dissimilatory and assimilatory nitrate reduction (napA, nasA) and corresponding cofactor biosynthesis (genomic moaB, moeB, and dsbC and plasmid-carried dsbD and ccmH) were found to cause anaerobic growth defects. In contrast, mutation of anaerobically induced genes encoding proteins required for the later denitrification steps (nirS, nirJ, nosD), dimethyl sulfoxide reduction (dmsA1), and fermentation (pdhB1, arcA, aceE, pta, acs) did not result in decreased anaerobic growth under the conditions tested. Additional essential components (ferredoxin, cccA) of the anaerobic electron transfer chain and central metabolism (pdhB) were identified. Another surprise was the importance of sodium gradient-dependent membrane processes and genomic rearrangements via viruses, transposons, and insertion sequence elements for anaerobic growth. These processes and the observed contributions of cell envelope restructuring (lysM, mipA, fadK), C4-dicarboxylate transport (dctM1, dctM3), and protease functions to anaerobic growth require further investigation to unravel the novel underlying adaptation strategies. PMID:23974024

Ebert, Matthias; Laaß, Sebastian; Burghartz, Melanie; Petersen, Jörn; Koßmehl, Sebastian; Wöhlbrand, Lars; Rabus, Ralf; Wittmann, Christoph; Tielen, Petra; Jahn, Dieter

2013-08-23

189

Dependence Relationships between Gene Ontology Terms based on TIGR Gene Product Annotations  

Microsoft Academic Search

The Gene Ontology is an important tool for the representation and processing of information about gene products and functions. It provides controlled vocabularies for the designations of cellular components, molecular functions, and biological processes used in the annotation of genes and gene products. These constitute three separate ontologies, of cellular com- ponents), molecular functions and biological processes, respectively. The question

Anand KUMAR; Barry SMITH; Christian BORGELT

190

The tax gene of human T-cell leukemia virus type 2 is essential for transformation of human T lymphocytes.  

PubMed Central

The mechanism of human T-cell leukemia virus (HTLV)-mediated transformation and induction of malignancy is unknown; however, several studies have implicated the viral gene product, Tax. Conclusive evidence for the role of Tax in the HTLV malignant process has been impeded by the inability to mutate tax in the context of an infectious virus and dissociate viral replication from cellular transformation. To circumvent this problem we constructed a mutant of HTLV type 2 (HTLV-2) that replicates by a Tax-independent mechanism. For these studies, the Tax response element in the viral long terminal repeat was replaced with the cytomegalovirus immediate-early promoter enhancer (C-enh). Transcription of the chimeric HTLV-2 (HTLVC-enh) was efficiently directed by this heterologous promoter. Also, the chimeric virus transformed primary human T lymphocytes with an efficiency similar to that of wild-type HTLV-2. A tax-knockout virus, termed HTLVC-enhDeltaTax, was constructed to directly assess the importance of Tax in cellular transformation. Transfection and infection studies indicated that HTLVC-enhDeltaTax was replication competent; however, HTLVC-enhDeltaTax failed to transform primary human T lymphocytes. We conclude that Tax is essential for HTLV-mediated transformation of human T lymphocytes. Furthermore, this chimeric HTLV, that replicates in the absence of Tax, should facilitate studies to determine the precise mechanism of T-lymphocyte transformation by HTLV.

Ross, T M; Pettiford, S M; Green, P L

1996-01-01

191

29 CFR 776.17 - Employment in a âclosely related process or occupation directly essential toâ production of goods.  

Code of Federal Regulations, 2013 CFR

...directly essential to the production thereof...77 Prior to the Fair Labor Standards Amendments of 1949, this was...occupation necessary to the productionâ of...The legislative history shows that the...3(j) of the Act is intended...

2013-07-01

192

The Hansenula polymorpha PEX14 gene encodes a novel peroxisomal membrane protein essential for peroxisome biogenesis.  

PubMed Central

We have cloned the Hansenula polymorpha PEX14 gene by functional complementation of the chemically induced pex14-1 mutant, which lacked normal peroxisomes. The sequence of the PEX14 gene predicts a novel protein product (Pex14p) of 39 kDa which showed no similarity to any known protein and lacked either of the two known peroxisomal targeting signals. Biochemical and electron microscopical analysis indicated that Pex14p is a component of the peroxisomal membrane. The synthesis of Pex14p is induced by peroxisome-inducing growth conditions. In cells of both pex14-1 and a PEX14 disruption mutant, peroxisomal membrane remnants were evident; these contained the H.polymorpha peroxisomal membrane protein Pex3p together with a small amount of the major peroxisomal matrix proteins alcohol oxidase, catalase and dihydroxyacetone synthase, the bulk of which resided in the cytosol. Unexpectedly, overproduction of Pex14p in wild-type H. polymorpha cells resulted in a peroxisome-deficient phenotype typified by the presence of numerous small vesicles which lacked matrix proteins; these were localized in the cytosol. Apparently, the stoichiometry of Pex14p relative to one or more other components of the peroxisome biogenesis machinery appears to be critical for protein import.

Komori, M; Rasmussen, S W; Kiel, J A; Baerends, R J; Cregg, J M; van der Klei, I J; Veenhuis, M

1997-01-01

193

Transducin Beta-Like Gene FTL1 Is Essential for Pathogenesis in Fusarium graminearum ?  

PubMed Central

Fusarium head blight caused by Fusarium graminearum is an important disease of wheat and barley. In a previous study, we identified several mutants with reduced virulence by insertional mutagenesis. A transducin beta-like gene named FTL1 was disrupted in one of these nonpathogenic mutants. FTL1 is homologous to Saccharomyces cerevisiae SIF2, which is a component of the Set3 complex involved in late stages of ascospore formation. The ?ftl1 mutant was significantly reduced in conidiation and failed to cause typical disease symptoms. It failed to colonize the vascular tissues of rachis or cause necrosis on the rachis of inoculated wheat heads. The ?ftl1 mutant also was defective in spreading from infected anthers to ovaries and more sensitive than the wild type to plant defensins MsDef1 and osmotin. However, the activation of two mitogen-activated protein kinases, Mgv1 and Gpmk1, production of deoxynivalenol, and expression of genes known to be important for plant infection in F. graminearum were not affected, indicating that the defect of the ?ftl1 mutant in plant infection is unrelated to known virulence factors in this pathogen and may involve novel mechanisms. The ?ftl1 deletion mutant was significantly reduced in histone deacetylation, and many members of the yeast Set3 complex are conserved in F. graminearum. FTL1 appears to be a component of this well-conserved protein complex that plays a critical role in the penetration and colonization of wheat tissues.

Ding, Shengli; Mehrabi, Rahim; Koten, Cornelia; Kang, Zhensheng; Wei, Yangdou; Seong, Kyeyong; Kistler, H. Corby; Xu, Jin-Rong

2009-01-01

194

Gene repression by Pax5 in B cells is essential for blood cell homeostasis and is reversed in plasma cells.  

PubMed

The transcription factor Pax5 represses lineage-inappropriate genes and activates B cell-specific genes in B lymphocytes. By identifying 110 Pax5-repressed genes, we now demonstrate that Pax5 downregulates diverse biological activities including receptor signaling, cell adhesion, migration, transcriptional control, and cellular metabolism at B cell commitment. The T lymphoid or myeloid expression of these genes demonstrates that Pax5(-/-) pro-B cells and common lymphoid progenitors display lymphoid and myeloid promiscuity of gene expression. These lineage-inappropriate genes require continuous Pax5 activity for their repression, as they are reactivated in committed pro-B cells and mature B cells following conditional Pax5 deletion. Pax5-repressed genes are also reexpressed in plasma cells, which depend for normal function on Cd28 and Ccr2 reactivation. The loss of Pax5 during terminal differentiation thus contributes to the plasma cell transcription program. Finally, ectopic expression of the Pax5-repressed chemokine gene Ccl3 in B cells results in increased osteoclast formation and bone loss, demonstrating that Pax5-mediated gene repression is essential for normal homeostasis of hematopoietic development. PMID:16546096

Delogu, Alessio; Schebesta, Alexandra; Sun, Qiong; Aschenbrenner, Katharina; Perlot, Thomas; Busslinger, Meinrad

2006-03-01

195

The Am-tra2 gene is an essential regulator of female splice regulation at two levels of the sex determination hierarchy of the honeybee.  

PubMed

Heteroallelic and homo- or hemiallelic Complementary sex determiner (Csd) proteins determine sexual fate in the honeybee (Apis mellifera) by controlling the alternative splicing of the downstream gene fem (feminizer). Thus far, we have little understanding of how heteroallelic Csd proteins mediate the splicing of female fem messenger RNAs (mRNAs) or how Fem proteins direct the splicing of honeybee dsx (Am-dsx) pre-mRNAs. Here, we report that Am-tra2, which is an ortholog of Drosophila melanogaster tra2, is an essential component of female splicing of the fem and Am-dsx transcripts in the honeybee. The Am-tra2 transcripts are alternatively (but non-sex-specifically) spliced, and they are translated into six protein isoforms that all share the basic RNA-binding domain/RS (arginine/serine) domain structure. Knockdown studies showed that the Am-tra2 gene is required to splice fem mRNAs into the productive female and nonproductive male forms. We suggest that the Am-Tra2 proteins are essential regulators of fem pre-mRNA splicing that, together with heteroallelic Csd proteins and/or Fem proteins, implement the female pathway. In males, the Am-Tra2 proteins may enhance the switch of fem transcripts into the nonproductive male form when heteroallelic Csd proteins are absent. This dual function of Am-Tra2 proteins possibly enhances and stabilizes the binary decision process of male/female splicing. Our knockdown studies also imply that the Am-Tra2 protein is an essential regulator for Am-dsx female splice regulation, suggesting an ancestral role in holometabolous insects. We also provide evidence that the Am-tra2 gene has an essential function in honeybee embryogenesis that is unrelated to sex determination. PMID:22942126

Nissen, Inga; Müller, Miriam; Beye, Martin

2012-08-31

196

The Am-tra2 Gene Is an Essential Regulator of Female Splice Regulation at Two Levels of the Sex Determination Hierarchy of the Honeybee  

PubMed Central

Heteroallelic and homo- or hemiallelic Complementary sex determiner (Csd) proteins determine sexual fate in the honeybee (Apis mellifera) by controlling the alternative splicing of the downstream gene fem (feminizer). Thus far, we have little understanding of how heteroallelic Csd proteins mediate the splicing of female fem messenger RNAs (mRNAs) or how Fem proteins direct the splicing of honeybee dsx (Am-dsx) pre-mRNAs. Here, we report that Am-tra2, which is an ortholog of Drosophila melanogaster tra2, is an essential component of female splicing of the fem and Am-dsx transcripts in the honeybee. The Am-tra2 transcripts are alternatively (but non-sex-specifically) spliced, and they are translated into six protein isoforms that all share the basic RNA-binding domain/RS (arginine/serine) domain structure. Knockdown studies showed that the Am-tra2 gene is required to splice fem mRNAs into the productive female and nonproductive male forms. We suggest that the Am-Tra2 proteins are essential regulators of fem pre-mRNA splicing that, together with heteroallelic Csd proteins and/or Fem proteins, implement the female pathway. In males, the Am-Tra2 proteins may enhance the switch of fem transcripts into the nonproductive male form when heteroallelic Csd proteins are absent. This dual function of Am-Tra2 proteins possibly enhances and stabilizes the binary decision process of male/female splicing. Our knockdown studies also imply that the Am-Tra2 protein is an essential regulator for Am-dsx female splice regulation, suggesting an ancestral role in holometabolous insects. We also provide evidence that the Am-tra2 gene has an essential function in honeybee embryogenesis that is unrelated to sex determination.

Nissen, Inga; Muller, Miriam; Beye, Martin

2012-01-01

197

Uptake of N,N?-Diacetylchitobiose [(GlcNAc)2] via the Phosphotransferase System Is Essential for Chitinase Production by Serratia marcescens 2170  

PubMed Central

The chiR gene of Serratia marcescens 2170, encoding a LysR-type transcriptional activator, was identified previously as an essential factor for expression of chitinases and a chitin-binding protein, CBP21. To identify other genes that are essential for chitinase production, transposon mutagenesis with mini-Tn5Km1 was carried out, and 25 mutants that were unable to produce chitinases and CBP21 were obtained. Analysis of the mutated gene of one of the mutants, N22, revealed the presence of a pts operon in this bacterium, and a mutation was found in ptsI in the operon. In addition to its inability to produce chitinase, N22 did not grow well on N-acetyl-d-glucosamine (GlcNAc), (GlcNAc)2, and some other carbon sources, most of which were phosphotransferase system (PTS) sugars. Thus, the inability to produce chitinase was assumed to be caused by the defect in uptake of (GlcNAc)2 via the PTS, considering that (GlcNAc)2 is the minimal substrate for chitinase induction and the major product of chitin hydrolysis by chitinases of this bacterium. To confirm this assumption, the chb operon, encoding the (GlcNAc)2-specific enzyme II permease, was cloned by reference to its Escherichia coli counterpart, and the Serratia chb operon was shown to comprise chbB, chbC, bglA, chbR, and chbG. Disruption of chbC drastically reduced production of chitinases and CBP21 and impaired growth on colloidal chitin. These results indicate that uptake of (GlcNAc)2 is mediated by the PTS and that the (GlcNAc)2-specific enzyme II permease constitutes its major pathway. Since (GlcNAc)2 uptake is essential for induction of chitinases and CBP21 production, (GlcNAc)2 appears to be the key molecule in recognition and utilization of chitin by S. marcescens.

Uchiyama, Taku; Kaneko, Ryousuke; Yamaguchi, Junko; Inoue, Akane; Yanagida, Takahiro; Nikaidou, Naoki; Regue, Miguel; Watanabe, Takeshi

2003-01-01

198

The Tumor Suppressor Gene, RASSF1A, Is Essential for Protection against Inflammation -Induced Injury.  

PubMed

Ras association domain family protein 1A (RASSF1A) is a tumor suppressor gene silenced in cancer. Here we report that RASSF1A is a novel regulator of intestinal inflammation as Rassf1a(+/-) , Rassf1a(-/-) and an intestinal epithelial cell specific knockout mouse (Rassf1a (IEC-KO) ) rapidly became sick following dextran sulphate sodium (DSS) administration, a chemical inducer of colitis. Rassf1a knockout mice displayed clinical symptoms of inflammatory bowel disease including: increased intestinal permeability, enhanced cytokine/chemokine production, elevated nuclear factor of kappa light polypeptide gene enhancer in B-cells (NF?B) activity, elevated colonic cell death and epithelial cell injury. Furthermore, epithelial restitution/repair was inhibited in DSS-treated Rassf1a(-/-) mice with reduction of several makers of proliferation including Yes associated protein (YAP)-driven proliferation. Surprisingly, tyrosine phosphorylation of YAP was detected which coincided with increased nuclear p73 association, Bax-driven epithelial cell death and p53 accumulation resulting in enhanced apoptosis and poor survival of DSS-treated Rassf1a knockout mice. We can inhibit these events and promote the survival of DSS-treated Rassf1a knockout mice with intraperitoneal injection of the c-Abl and c-Abl related protein tyrosine kinase inhibitor, imatinib/gleevec. However, p53 accumulation was not inhibited by imatinib/gleevec in the Rassf1a(-/-) background which revealed the importance of p53-dependent cell death during intestinal inflammation. These observations suggest that tyrosine phosphorylation of YAP (to drive p73 association and up-regulation of pro-apoptotic genes such as Bax) and accumulation of p53 are consequences of inflammation-induced injury in DSS-treated Rassf1a(-/-) mice. Mechanistically, we can detect robust associations of RASSF1A with membrane proximal Toll-like receptor (TLR) components to suggest that RASSF1A may function to interfere and restrict TLR-driven activation of NF?B. Failure to restrict NF?B resulted in the inflammation-induced DNA damage driven tyrosine phosphorylation of YAP, subsequent p53 accumulation and loss of intestinal epithelial homeostasis. PMID:24146755

Gordon, Marilyn; El-Kalla, Mohamed; Zhao, Yuewen; Fiteih, Yahya; Law, Jennifer; Volodko, Natalia; Mohamed, Anwar; El-Kadi, Ayman O S; Liu, Lei; Odenbach, Jeff; Thiesen, Aducio; Onyskiw, Christina; Ghazaleh, Haya Abu; Park, Jikyoung; Lee, Sean Bong; Yu, Victor C; Fernandez-Patron, Carlos; Alexander, R Todd; Wine, Eytan; Baksh, Shairaz

2013-10-16

199

The Tumor Suppressor Gene, RASSF1A, Is Essential for Protection against Inflammation -Induced Injury  

PubMed Central

Ras association domain family protein 1A (RASSF1A) is a tumor suppressor gene silenced in cancer. Here we report that RASSF1A is a novel regulator of intestinal inflammation as Rassf1a+/?, Rassf1a?/? and an intestinal epithelial cell specific knockout mouse (Rassf1a IEC-KO) rapidly became sick following dextran sulphate sodium (DSS) administration, a chemical inducer of colitis. Rassf1a knockout mice displayed clinical symptoms of inflammatory bowel disease including: increased intestinal permeability, enhanced cytokine/chemokine production, elevated nuclear factor of kappa light polypeptide gene enhancer in B-cells (NF?B) activity, elevated colonic cell death and epithelial cell injury. Furthermore, epithelial restitution/repair was inhibited in DSS-treated Rassf1a?/? mice with reduction of several makers of proliferation including Yes associated protein (YAP)-driven proliferation. Surprisingly, tyrosine phosphorylation of YAP was detected which coincided with increased nuclear p73 association, Bax-driven epithelial cell death and p53 accumulation resulting in enhanced apoptosis and poor survival of DSS-treated Rassf1a knockout mice. We can inhibit these events and promote the survival of DSS-treated Rassf1a knockout mice with intraperitoneal injection of the c-Abl and c-Abl related protein tyrosine kinase inhibitor, imatinib/gleevec. However, p53 accumulation was not inhibited by imatinib/gleevec in the Rassf1a?/? background which revealed the importance of p53-dependent cell death during intestinal inflammation. These observations suggest that tyrosine phosphorylation of YAP (to drive p73 association and up-regulation of pro-apoptotic genes such as Bax) and accumulation of p53 are consequences of inflammation-induced injury in DSS-treated Rassf1a?/? mice. Mechanistically, we can detect robust associations of RASSF1A with membrane proximal Toll-like receptor (TLR) components to suggest that RASSF1A may function to interfere and restrict TLR-driven activation of NF?B. Failure to restrict NF?B resulted in the inflammation-induced DNA damage driven tyrosine phosphorylation of YAP, subsequent p53 accumulation and loss of intestinal epithelial homeostasis.

Fiteih, Yahya; Law, Jennifer; Volodko, Natalia; Mohamed, Anwar; El-Kadi, Ayman O. S.; Liu, Lei; Odenbach, Jeff; Thiesen, Aducio; Onyskiw, Christina; Ghazaleh, Haya Abu; Park, Jikyoung; Lee, Sean Bong; Yu, Victor C.; Fernandez-Patron, Carlos; Alexander, R. Todd; Wine, Eytan; Baksh, Shairaz

2013-01-01

200

Two Closely Related Genes of Arabidopsis Encode Plastidial Cytidinediphosphate Diacylglycerol Synthases Essential for Photoautotrophic Growth1[C  

PubMed Central

Cytidinediphosphate diacylglycerol synthase (CDS) catalyzes the formation of cytidinediphosphate diacylglycerol, an essential precursor of anionic phosphoglycerolipids like phosphatidylglycerol or -inositol. In plant cells, CDS isozymes are located in plastids, mitochondria, and microsomes. Here, we show that these isozymes are encoded by five genes in Arabidopsis (Arabidopsis thaliana). Alternative translation initiation or alternative splicing of CDS2 and CDS4 transcripts can result in up to 10 isoforms. Most of the cDNAs encoding the various plant isoforms were functionally expressed in yeast and rescued the nonviable phenotype of the mutant strain lacking CDS activity. The closely related genes CDS4 and CDS5 were found to encode plastidial isozymes with similar catalytic properties. Inactivation of both genes was required to obtain Arabidopsis mutant lines with a visible phenotype, suggesting that the genes have redundant functions. Analysis of these Arabidopsis mutants provided further independent evidence for the importance of plastidial phosphatidylglycerol for structure and function of thylakoid membranes and, hence, for photoautotrophic growth.

Haselier, Andre; Akbari, Hana; Weth, Agnes; Baumgartner, Werner; Frentzen, Margrit

2010-01-01

201

Trm2p-dependent derepression is essential for methanol-specific gene activation in the methylotrophic yeast Candida boidinii.  

PubMed

We identified a gene, designated TRM2, responsible for methanol-inducible gene expression in the methylotrophic yeast Candida boidinii. The encoded protein Trm2p contains two C(2)H(2)-type zinc finger motifs near the N terminus and shows high similarity to Saccharomyces cerevisiae Adr1p and Pichia pastoris Mxr1p. A C. boidinii gene-disrupted strain (trm2Delta) could not grow on methanol or oleate, but could grow on glucose or ethanol. Trm2p was necessary for the activation of five methanol-inducible promoters tested. Trm2p was localized to the nucleus during growth on nonfermentable carbon sources, but to the cytosol during growth on glucose. A chromatin immunoprecipitation assay revealed that Trm2p specifically bound to the promoters of the alcohol oxidase gene (AOD1) and the dihydroxyacetone synthase gene in cells grown on methanol or oleate, but did not bind to these promoters in cells grown on glucose. The derepressed level of expression of AOD1, which was observed in the trm1Delta strain (the TRM1 gene encodes a transcription factor responsible for methanol-specific gene activation), was decreased in the trm1Deltatrm2Delta strain to a level similar to that observed in the trm2Delta strain. These results suggest that Trm2p-dependent derepression is essential for the Trm1p-dependent methanol-specific gene activation in C. boidinii. PMID:20491943

Sasano, Yu; Yurimoto, Hiroya; Kuriyama, Masamitsu; Sakai, Yasuyoshi

2010-04-29

202

Molecular cloning and expression of the spsB gene encoding an essential type I signal peptidase from Staphylococcus aureus.  

PubMed Central

The gene, spsB, encoding a type I signal peptidase has been cloned from the gram-positive eubacterium Staphylococcus aureus. The gene encodes a protein of 191 amino acid residues with a calculated molecular mass of 21,692 Da. Comparison of the protein sequence with those of known type I signal peptidases indicates conservation of amino acid residues known to be important or essential for catalytic activity. The enzyme has been expressed to high levels in Escherichia coli and has been demonstrated to possess enzymatic activity against E. coli preproteins in vivo. Experiments whereby the spsB gene was transferred to a plasmid that is temperature sensitive for replication indicate that spsB is an essential gene. We identified an open reading frame immediately upstream of the spsB gene which encodes a type I signal peptidase homolog of 174 amino acid residues with a calculated molecular mass of 20,146 Da that is predicted to be devoid of catalytic activity.

Cregg, K M; Wilding, I; Black, M T

1996-01-01

203

Gene regulatory networks controlling hematopoietic progenitor niche cell production and differentiation in the Drosophila lymph gland.  

PubMed

Hematopoiesis occurs in two phases in Drosophila, with the first completed during embryogenesis and the second accomplished during larval development. The lymph gland serves as the venue for the final hematopoietic program, with this larval tissue well-studied as to its cellular organization and genetic regulation. While the medullary zone contains stem-like hematopoietic progenitors, the posterior signaling center (PSC) functions as a niche microenvironment essential for controlling the decision between progenitor maintenance versus cellular differentiation. In this report, we utilize a PSC-specific GAL4 driver and UAS-gene RNAi strains, to selectively knockdown individual gene functions in PSC cells. We assessed the effect of abrogating the function of 820 genes as to their requirement for niche cell production and differentiation. 100 genes were shown to be essential for normal niche development, with various loci placed into sub-groups based on the functions of their encoded protein products and known genetic interactions. For members of three of these groups, we characterized loss- and gain-of-function phenotypes. Gene function knockdown of members of the BAP chromatin-remodeling complex resulted in niche cells that do not express the hedgehog (hh) gene and fail to differentiate filopodia believed important for Hh signaling from the niche to progenitors. Abrogating gene function of various members of the insulin-like growth factor and TOR signaling pathways resulted in anomalous PSC cell production, leading to a defective niche organization. Further analysis of the Pten, TSC1, and TSC2 tumor suppressor genes demonstrated their loss-of-function condition resulted in severely altered blood cell homeostasis, including the abundant production of lamellocytes, specialized hemocytes involved in innate immune responses. Together, this cell-specific RNAi knockdown survey and mutant phenotype analyses identified multiple genes and their regulatory networks required for the normal organization and function of the hematopoietic progenitor niche within the lymph gland. PMID:22911822

Tokusumi, Yumiko; Tokusumi, Tsuyoshi; Shoue, Douglas A; Schulz, Robert A

2012-07-24

204

The cycHJKL gene cluster plays an essential role in the biogenesis of c -type cytochromes in Bradyrhizobium japonicum  

Microsoft Academic Search

We present an extended genetic analysis of the previously identified cycH locus in Bradyrhizobium japonicum. Three new open reading frames found in an operon-like structure immediately adjacent to the 3' end of cycH were termed cycJ, cycK and cycL. A deletion mutant (?cycHJKL) and biochemical analysis of its phenotype showed that the genes of the cluster are essential for the

Daniel Ritz; Linda Thöny-Meyer; Hauke Hennecke

1995-01-01

205

Methods of double-stranded RNA-mediated gene inactivation in Arabidopsis and their use to define an essential gene in methionine biosynthesis.  

PubMed

Controlled down-regulation of endogenous plant gene expression is a useful tool, but antisense and sense silencing lack predictability. Recent studies show that expression of both antisense and sense RNA together is an effective means of inactivating reporter and viral genes in plants. We created transgenic plants expressing antisense and sense RNA together in a single 'double-stranded RNA' (dsRNA) transcript. This approach shows great promise as a highly effective means for reducing gene function. With this approach, we demonstrated that the Arabidopsis cystathionine beta-lyase gene, which encodes a methionine biosynthetic enzyme, is essential for viability. Inactivation of this gene was rescued by the addition of methionine to the growth medium. Compared to antisense and sense constructs, the dsRNA construct showed a much more consistent and complete suppression of gene activity. Additionally, expression of a transcript with a spacer sequence containing an unrelated gene between antisense and sense luciferase gene fragments led to stronger inactivation of a second luciferase transgene than did constructs with a minimal spacing between sense and antisense fragments. However, the gene in the spacer region was neither functionally expressed nor functional in silencing a second, unlinked homologous transgene. PMID:11202438

Levin, J Z; de Framond, A J; Tuttle, A; Bauer, M W; Heifetz, P B

2000-12-01

206

PLASMA PROTEIN PRODUCTION INFLUENCED BY AMINO ACID MIXTURES AND LACK OF ESSENTIAL AMINO ACIDS  

PubMed Central

When blood plasma proteins are depleted by bleeding with return of red cells suspended in saline (plasmapheresis) it is possible to bring dogs to a steady state of hypoproteinemia and a constant level of plasma protein production if the diet nitrogen intake is controlled and limited. Such dogs are outwardly normal but have a lowered resistance to infection and intoxication and probably to vitamin deficiency. When the diet nitrogen is provided by certain mixtures of the ten growth essential amino acids plus glycine, given intravenously at a rapid rate, plasma protein production is good. The same mixture absorbed subcutaneously at a slower rate may be slightly better utilized. Fed orally the same mixture is better utilized and associated with a lower urinary nitrogen excretion. An ample amino acid mixture for the daily intake of a 10 kilo dog may contain in grams dl-threonine 1.4, dl-valine 3, dl-leucine 3, dl-isoleucine 2, l(+)-lysine·HCl·H2O 2.2, dl-tryptophane 0.3, dl-phenylalanine 2, dl-methionine 1.2, l(+)-histidine·HCl·H2O 1, l(+)-arginine·HCl 1, and glycine 2. Half this quantity is inadequate and not improved by addition of a mixture of alanine, serine, norleucine, proline, hydroxyproline, and tyrosine totalling 1.4 gm. Aspartic acid appears to induce vomiting when added to a mixture of amino acids. The same response has been reported for glutamic acid (8). Omission from the intake of leucine or of leucine and isoleucine results in negative nitrogen balance and rapid weight loss but plasma protein production may be temporarily maintained. It is possible that leucine may be captured from red blood cell destruction. Tryptophane deficiency causes an abrupt decline in plasma protein production. No decline occurred during 2 weeks of histidine deficiency but the urinary nitrogen increased to negative balance. Plasma protein production may be impaired during conditions of dietary deficiency not related to the protein or amino acid intake. Skin lesions and liver function impairment are described. Unidentified factors present in liver and yeast appear to be involved.

Madden, S. C.; Anderson, F. W.; Donovan, J. C.; Whipple, G. H.

1945-01-01

207

The ZIC gene family encodes multi-functional proteins essential for patterning and morphogenesis.  

PubMed

The zinc finger of the cerebellum gene (ZIC) discovered in Drosophila melanogaster (odd-paired) has five homologs in Xenopus, chicken, mice, and humans, and seven in zebrafish. This pattern of gene copy expansion is accompanied by a divergence in gene and protein structure, suggesting that Zic family members share some, but not all, functions. ZIC genes are implicated in neuroectodermal development and neural crest cell induction. All share conserved regions encoding zinc finger domains, however their heterogeneity and specification remain unexplained. In this review, the evolution, structure, and expression patterns of the ZIC homologs are described; specific functions attributable to individual family members are supported. A review of data from functional studies in Xenopus and murine models suggest that ZIC genes encode multifunctional proteins operating in a context-specific manner to drive critical events during embryogenesis. The identification of ZIC mutations in congenital syndromes highlights the relevance of these genes in human development. PMID:23443491

Houtmeyers, Rob; Souopgui, Jacob; Tejpar, Sabine; Arkell, Ruth

2013-02-27

208

Effect of different liming levels on the biomass production and essential oil extraction yield of Cunila galioides Benth.  

PubMed

Poejo is an aromatic and medicinal plant native to highland areas of south Brazil, in acid soils with high Al3+ concentration. The main objective of the present work was to evaluate the effect of liming on the extraction yield of essential oil of three chemotypes of poejo (Cunila galioides Benth). For this purpose, the experiments were performed in a greenhouse, using 8-litre pots. The treatments were four dosages of limestone (0, 3.15, 12.5, and 25 g.L(-1)) and a completely random experimental design was used, with four replications and three chemotypes, set up in a 3 × 4 factorial arrangement. The parameters evaluated were dry weight of aerial parts, essential oil content and chemical composition of essential oil. Results showed that liming affects the biomass production, essential oil yield and chemical composition, with cross interaction verified between chemotype and limestone dosage. For the higher dosage lower biomass production, lower yield of essential oil as well as the lowest content of citral (citral chemotype) and limonene (menthene chemotype) was observed. In the ocimene chemotype, no liming influence was observed on the essential oil yield and on the content of major compounds. The dosage of 3.15 g.L(-1) can be considered the best limestone dosage for the production of poejo for the experimental conditions evaluated. PMID:23295505

Mossi, A J; Pauletti, G F; Rota, L; Echeverrigaray, S; Barros, I B I; Oliveira, J V; Paroul, N; Cansian, R L

2012-11-01

209

The phs gene and hydrogen sulfide production by Salmonella typhimurium.  

PubMed

Salmonella typhimurium produces H2S from thiosulfate or sulfite. The respective pathways for the two reductions must be distinct as mutants carrying motations in phs, chlA, and menB reduced sulfite, but not thiosulfate, to H2S, and glucose repressed the production of H2S from thiosulfate while it stimulated its production from sulfite. The phs and chlA mutants also lacked a methyl viologen-linked thiosulfate reductase activity present in anaerobically grown wild-type cultures. A number of hydroxylamine, transposon Tn10 insertion, and Mu d1(Apr lac) operon fusion mutants defective in phs were characterized. One of the hydroxylamine mutants was an amber mutant, as indicated by suppression of its mutation in a supD background. The temperature-sensitive phs mutants produced H2S and methyl viologen-linked thiosulfate reductase at 30 degrees C but not at 42 degrees C. The reductases in all such mutants grown at 30 degrees C were as thermostable as the wild-type enzyme and did not differ in electrophoretic relative mobility, suggesting that phs is not the structural gene for thiosulfate reductase. Expression of beta-galactosidase in phs::Mu d1(Apr lac) mutants was dependent on anaerobiosis and the presence of reduced sulfur. It was also strongly influenced by carbon source and growth stage. The results are consistent with a model in which the phs gene encodes a regulatory protein essential for the reduction of thiosulfate to hydrogen sulfide. PMID:3108233

Clark, M A; Barrett, E L

1987-06-01

210

A Yeast Gene that is Essential for Release from Glucose Repression Encodes a Protein Kinase  

Microsoft Academic Search

The SNF1 gene plays a central role in carbon catabolite repression in the yeast Saccharomyces cerevisiae, namely that SNF1 function is required for expression of glucose-repressible genes. The nucleotide sequence of the cloned SNF1 gene was determined, and the predicted amino acid sequence shows that SNF1 encodes a 72,040-dalton polypeptide that has significant homology to the conserved catalytic domain of

John L. Celenza; Marian Carlson

1986-01-01

211

Insertional mutagenesis in zebrafish rapidly identifies genes essential for early vertebrate development  

Microsoft Academic Search

To rapidly identify genes required for early vertebrate development, we are carrying out a large-scale, insertional mutagenesis screen in zebrafish, using mouse retroviral vectors as the mutagen. We will obtain mutations in 450 to 500 different genes—roughly 20% of the genes that can be mutated to produce a visible embryonic phenotype in this species—and will clone the majority of the

Gregory Golling; Adam Amsterdam; Zhaoxia Sun; Marcelo Antonelli; Ernesto Maldonado; Wenbiao Chen; Shawn Burgess; Maryann Haldi; Karen Artzt; Sarah Farrington; Shuh-Yow Lin; Robert M. Nissen; Nancy Hopkins

2002-01-01

212

Usher syndrome IIIA gene clarin-1 is essential for hair cell function and associated neural activation.  

PubMed

Usher syndrome 3A (USH3A) is an autosomal recessive disorder characterized by progressive loss of hearing and vision due to mutation in the clarin-1 (CLRN1) gene. Lack of an animal model has hindered our ability to understand the function of CLRN1 and the pathophysiology associated with USH3A. Here we report for the first time a mouse model for ear disease in USH3A. Detailed evaluation of inner ear phenotype in the Clrn1 knockout mouse (Clrn1(-/-)) coupled with expression pattern of Clrn1 in the inner ear are presented here. Clrn1 was expressed as early as embryonic day 16.5 in the auditory and vestibular hair cells and associated ganglionic neurons, with its expression being higher in outer hair cells (OHCs) than inner hair cells. Clrn1(-/-) mice showed early onset hearing loss that rapidly progressed to severe levels. Two to three weeks after birth (P14-P21), Clrn1(-/-) mice showed elevated auditory-evoked brainstem response (ABR) thresholds and prolonged peak and interpeak latencies. By P21, approximately 70% of Clrn1(-/-) mice had no detectable ABR and by P30 these mice were deaf. Distortion product otoacoustic emissions were not recordable from Clrn1(-/-) mice. Vestibular function in Clrn1(-/-) mice mirrored the cochlear phenotype, although it deteriorated more gradually than cochlear function. Disorganization of OHC stereocilia was seen as early as P2 and by P21 OHC loss was observed. In sum, hair cell dysfunction and prolonged peak latencies in vestibular and cochlear evoked potentials in Clrn1(-/-) mice strongly indicate that Clrn1 is necessary for hair cell function and associated neural activation. PMID:19414487

Geng, Ruishuang; Geller, Scott F; Hayashi, Toshinori; Ray, Catherine A; Reh, Thomas A; Bermingham-McDonogh, Olivia; Jones, Sherri M; Wright, Charles G; Melki, Sami; Imanishi, Yoshikazu; Palczewski, Krzysztof; Alagramam, Kumar N; Flannery, John G

2009-05-03

213

Effects of essential oils on rumen fermentation, milk production, and feeding behavior in lactating dairy cows.  

PubMed

Seven ruminally cannulated lactating Holstein dairy cows were used in an incomplete Latin rectangle design to assess the effects of 2 commercial essential oil (EO) products on rumen fermentation, milk production, and feeding behavior. Cows were fed a total mixed ration with a 42:58 forage:concentrate ratio (DM basis). Treatments included addition of 0.5 g/d of CE Lo (85 mg of cinnamaldehyde and 140 mg of eugenol), 10 g/d of CE Hi (1,700 mg of cinnamaldehyde and 2,800 mg of eugenol), 0.25 g/d of CAP (50mg of capsicum), or no oil (CON). Cows were fed ad libitum twice daily for 21 d per period. Dry matter intake, number of meals/d, h eating/d, mean meal length, rumination events/d, h ruminating/d, and mean rumination length were not affected by EO. However, length of the first meal after feeding decreased with addition of CE Hi (47.2 min) and CAP (49.4 min) compared with CON (65.4 min). Total volatile fatty acids, individual volatile fatty acids, acetate:propionate ratio, and ammonia concentration were not affected by EO. Mean rumen pH as well as bouts, total h, mean bout length, total area, and mean bout area under pH 5.6 did not differ among treatments. Total tract digestibility of organic matter, dry matter, neutral detergent fiber, acid detergent fiber, crude protein, and starch were not affected by EO. Milk yield and composition did not change with EO. In situ dry matter disappearance of ground soybean hulls was not affected by EO. However, organic matter disappearance of soybean hulls with CE Hi tended to decrease compared with CON. Compared with CON, neutral detergent fiber disappearance (41.5 vs. 37.6%) and acid detergent fiber disappearance (44.5 vs. 38.8%) decreased with addition of CE Hi. The CE Lo had no effect on rumen fermentation, milk production, or feeding behavior but CAP shortened the length of the first meal without changing rumen fermentation or production, making it a possible additive for altering feeding behavior. The CE Hi negatively affected rumen fermentation and shortened the length of the first meal, suggesting that a dose of 10 g/d is not beneficial to lactating dairy cows. PMID:21524537

Tager, L R; Krause, K M

2011-05-01

214

Mammalian Polycomb-mediated repression of Hox genes requires the essential spliceosomal protein Sf3b1  

PubMed Central

Polycomb group (PcG) proteins are responsible for the stable repression of homeotic (Hox) genes by forming multimeric protein complexes. We show (1) physical interaction between components of the U2 small nuclear ribonucleoprotein particle (U2 snRNP), including Sf3b1 and PcG proteins Zfp144 and Rnf2; and (2) that Sf3b1 heterozygous mice exhibit skeletal transformations concomitant with ectopic Hox expressions. These alterations are enhanced by Zfp144 mutation but repressed by Mll mutation (a trithorax-group gene). Importantly, the levels of Sf3b1 in PcG complexes were decreased in Sf3b1-heterozygous embryos. These findings suggest that Sf3b1-PcG protein interaction is essential for true PcG-mediated repression of Hox genes.

Isono, Kyoichi; Mizutani-Koseki, Yoko; Komori, Toshihisa; Schmidt-Zachmann, Marion S.; Koseki, Haruhiko

2005-01-01

215

A novel class of evolutionarily conserved genes defined by piwi are essential for stem cell self-renewal  

PubMed Central

Germ-line stem cells (GSCs) serve as the source for gametogenesis in diverse organisms. We cloned and characterized the Drosophila piwi gene and showed that it is required for the asymmetric division of GSCs to produce and maintain a daughter GSC but is not essential for the further differentiation of the committed daughter cell. Genetic mosaic and RNA in situ analyses suggest that piwi expression in adjacent somatic cells regulates GSC division. piwi encodes a highly basic novel protein, well conserved during evolution. We isolated piwi homologs in Caenorhabditis elegans and humans and also identified Arabidopsis piwi-like genes known to be required for meristem cell maintenance. Decreasing C. elegans piwi expression reduces the proliferation of GSC-equivalent cells. Thus, piwi represents a novel class of genes required for GSC division in diverse organisms.

Cox, Daniel N.; Chao, Anna; Baker, Jeff; Chang, Lisa; Qiao, Dan; Lin, Haifan

1998-01-01

216

An essential Saccharomyces cerevisiae gene homologous to SNF2 encodes a helicase-related protein in a new family.  

PubMed Central

The Saccharomyces cerevisiae SNF2 gene affects the expression of many diversely regulated genes and has been implicated in transcriptional activation. We report here the cloning and characterization of STH1, a gene that is homologous to SNF2. STH1 is essential for mitotic growth and is functionally distinct from SNF2. A bifunctional STH1-beta-galactosidase protein is located in the nucleus. The predicted 155,914-Da STH1 protein is 72% identical to SNF2 over 661 amino acids and 46% identical over another stretch of 66 amino acids. Both STH1 and SNF2 contain a putative nucleoside triphosphate-binding site and sequences resembling the consensus helicase motifs. The large region of homology shared by STH1 and SNF2 is conserved among other eukaryotic proteins, and STH1 and SNF2 appear to define a novel family of proteins related to helicases. Images

Laurent, B C; Yang, X; Carlson, M

1992-01-01

217

Regulation of dev, an Operon That Includes Genes Essential for Myxococcus xanthus Development and CRISPR-Associated Genes and Repeats  

Microsoft Academic Search

Received 5 February 2007\\/Accepted 6 March 2007 Expression of dev genes is important for triggering spore differentiation inside Myxococcus xanthus fruiting bodies. DNA sequence analysis suggested that dev and cas (CRISPR-associated) genes are cotranscribed at the dev locus, which is adjacent to CRISPR (clustered regularly interspaced short palindromic repeats). Analysis of RNA from developing M. xanthus confirmed that dev and

Poorna Viswanathan; Kimberly Murphy; Bryan Julien; Anthony G. Garza; Lee Kroos

2007-01-01

218

Aerosolized essential oils and individual natural product compounds as brown treesnake repellents.  

PubMed

Chemical irritants useful as repellents for brown treesnakes (Boiga irregularis) were identified. Exposure to various compounds produced a range of intensities for locomotory behavior in snakes. Essential oils comprised of 10 g liter-1 solutions of cedarwood, cinnamon, sage, juniper berry, lavender and rosemary each were potent snake irritants. Brown treesnakes exposed to a 2-s burst of aerosol of these oils exhibited prolonged, violent undirected locomotory behavior. In contrast, exposure to a 10 g liter-1 concentration of ginger oil aerosol caused snakes to locomote, but in a deliberate, directed manner. We also tested specific compounds, all derivative of food and flavor ingredients. 10 g liter-1 solutions delivered as aerosols of m-anisaldehyde, trans-anethole, cineole, cinnamaldehyde, citral, ethyl phenylacetate, eugenol, geranyl acetate or methyl salicylate all acted as potent irritants for brown treesnakes. The individual ingredients were classified using cluster analysis into groups that promoted different levels of response by snakes. This study is the first to systematically investigate the irritant potential of natural products for snakes. These data will be useful in the development of practical pest management tools for snakes. PMID:12192901

Clark, Larry; Shivik, John

2002-08-01

219

Targeted Comparative RNA Interference Analysis Reveals Differential Requirement of Genes Essential for Cell Proliferation  

PubMed Central

Differences in the genetic and epigenetic make up of cell lines have been very useful for dissecting the roles of specific genes in the biology of a cell. Targeted comparative RNAi (TARCOR) analysis uses high throughput RNA interference (RNAi) against a targeted gene set and rigorous quantitation of the phenotype to identify genes with a differential requirement for proliferation between cell lines of different genetic backgrounds. To demonstrate the utility of such an analysis, we examined 257 growth-regulated genes in parallel in a breast epithelial cell line, MCF10A, and a prostate cancer cell line, PC3. Depletion of an unexpectedly high number of genes (25%) differentially affected proliferation of the two cell lines. Knockdown of many genes that spare PC3 (p53?) but inhibit MCF10A (p53+) proliferation induces p53 in MCF10A cells. EBNA1BP2, involved in ribosome biogenesis, is an example of such a gene, with its depletion arresting MCF10A at G1/S in a p53-dependent manner. TARCOR is thus useful for identifying cell type–specific genes and pathways involved in proliferation and also for exploring the heterogeneity of cell lines. In particular, our data emphasize the importance of considering the genetic status, when performing siRNA screens in mammalian cells.

Machida, Yuichi J.; Chen, Yuefeng; Machida, Yuka; Malhotra, Ankit; Sarkar, Sukumar

2006-01-01

220

laminin alpha 1 gene is essential for normal lens development in zebrafish  

Microsoft Academic Search

BACKGROUND: Laminins represent major components of basement membranes and play various roles in embryonic and adult tissues. The functional laminin molecule consists of three chains, alpha, beta and gamma, encoded by separate genes. There are twelve different laminin genes identified in mammals to date that are highly homologous in their sequence but different in their tissue distribution. The laminin alpha

Natalya S Zinkevich; Dmitry V Bosenko; Brian A Link; Elena V Semina

2006-01-01

221

Housekeeping genes essential for pantothenate biosynthesis are plasmid-encoded in Rhizobium etli and Rhizobium leguminosarum  

PubMed Central

Background A traditional concept in bacterial genetics states that housekeeping genes, those involved in basic metabolic functions needed for maintenance of the cell, are encoded in the chromosome, whereas genes required for dealing with challenging environmental conditions are located in plasmids. Exceptions to this rule have emerged from genomic sequence data of bacteria with multipartite genomes. The genome sequence of R. etli CFN42 predicts the presence of panC and panB genes clustered together on the 642 kb plasmid p42f and a second copy of panB on plasmid p42e. They encode putative pantothenate biosynthesis enzymes (pantoate-?-alanine ligase and 3-methyl-2-oxobutanoate hydroxymethyltransferase, respectively). Due to their ubiquitous distribution and relevance in the central metabolism of the cell, these genes are considered part of the core genome; thus, their occurrence in a plasmid is noteworthy. In this study we investigate the contribution of these genes to pantothenate biosynthesis, examine whether their presence in plasmids is a prevalent characteristic of the Rhizobiales with multipartite genomes, and assess the possibility that the panCB genes may have reached plasmids by horizontal gene transfer. Results Analysis of mutants confirmed that the panC and panB genes located on plasmid p42f are indispensable for the synthesis of pantothenate. A screening of the location of panCB genes among members of the Rhizobiales showed that only R. etli and R. leguminosarum strains carry panCB genes in plasmids. The panCB phylogeny attested a common origin for chromosomal and plasmid-borne panCB sequences, suggesting that the R. etli and R. leguminosarum panCB genes are orthologs rather than xenologs. The panCB genes could not totally restore the ability of a strain cured of plasmid p42f to grow in minimal medium. Conclusions This study shows experimental evidence that core panCB genes located in plasmids of R. etli and R. leguminosarum are indispensable for the synthesis of pantothenate. The unusual presence of panCB genes in plasmids of Rhizobiales may be due to an intragenomic transfer from chromosome to plasmid. Plasmid p42f encodes other functions required for growth in minimal medium. Our results support the hypothesis of cooperation among different replicons for basic cellular functions in multipartite rhizobia genomes.

2011-01-01

222

The Role of the Parkinson's Disease Gene PARK9 in Essential Cellular Pathways and the Manganese Homeostasis Network in Yeast  

PubMed Central

YPK9 (Yeast PARK9; also known as YOR291W) is a non-essential yeast gene predicted by sequence to encode a transmembrane P-type transport ATPase. However, its substrate specificity is unknown. Mutations in the human homolog of YPK9, ATP13A2/PARK9, have been linked to genetic forms of early onset parkinsonism. We previously described a strong genetic interaction between Ypk9 and another Parkinson's disease (PD) protein ?-synuclein in multiple model systems, and a role for Ypk9 in manganese detoxification in yeast. In humans, environmental exposure to toxic levels of manganese causes a syndrome similar to PD and is thus an environmental risk factor for the disease. How manganese contributes to neurodegeneration is poorly understood. Here we describe multiple genome-wide screens in yeast aimed at defining the cellular function of Ypk9 and the mechanisms by which it protects cells from manganese toxicity. In physiological conditions, we found that Ypk9 genetically interacts with essential genes involved in cellular trafficking and the cell cycle. Deletion of Ypk9 sensitizes yeast cells to exposure to excess manganese. Using a library of non-essential gene deletions, we screened for additional genes involved in tolerance to excess manganese exposure, discovering several novel pathways involved in manganese homeostasis. We defined the dependence of the deletion strain phenotypes in the presence of manganese on Ypk9, and found that Ypk9 deletion modifies the manganese tolerance of only a subset of strains. These results confirm a role for Ypk9 in manganese homeostasis and illuminates cellular pathways and biological processes in which Ypk9 likely functions.

Chesi, Alessandra; Kilaru, Austin; Fang, Xiaodong; Cooper, Antony A.; Gitler, Aaron D.

2012-01-01

223

trp, a Novel Mammalian Gene Family Essential for Agonist-Activated Capacitative Ca 2+ Entry  

Microsoft Academic Search

Capacitative calcium entry (CCE) describes Ca2+ influx into cells that replenishes Ca2+ stores emptied through the action of IP3 and other agents. It is an essential component of cellular responses to many hormones and growth factors. The molecular basis of this form of Ca2+ entry is complex and may involve more than one type of channel. Studies on visual signal

Xi Zhu; Meisheng Jiang; Michael Peyton; Guylain Boulay; Raymond Hurst; Enrico Stefani; Lutz Birnbaumer

1996-01-01

224

Ensuring the statistical soundness of competitive gene set approaches: gene filtering and genome-scale coverage are essential.  

PubMed

In this article, we focus on the analysis of competitive gene set methods for detecting the statistical significance of pathways from gene expression data. Our main result is to demonstrate that some of the most frequently used gene set methods, GSEA, GSEArot and GAGE, are severely influenced by the filtering of the data in a way that such an analysis is no longer reconcilable with the principles of statistical inference, rendering the obtained results in the worst case inexpressive. A possible consequence of this is that these methods can increase their power by the addition of unrelated data and noise. Our results are obtained within a bootstrapping framework that allows a rigorous assessment of the robustness of results and enables power estimates. Our results indicate that when using competitive gene set methods, it is imperative to apply a stringent gene filtering criterion. However, even when genes are filtered appropriately, for gene expression data from chips that do not provide a genome-scale coverage of the expression values of all mRNAs, this is not enough for GSEA, GSEArot and GAGE to ensure the statistical soundness of the applied procedure. For this reason, for biomedical and clinical studies, we strongly advice not to use GSEA, GSEArot and GAGE for such data sets. PMID:23389952

Tripathi, Shailesh; Glazko, Galina V; Emmert-Streib, Frank

2013-02-06

225

The lineage-specific gene ponzr1 is essential for zebrafish pronephric and pharyngeal arch development  

PubMed Central

The Homeobox (Hox) and Paired box (Pax) gene families are key determinants of animal body plans and organ structure. In particular, they function within regulatory networks that control organogenesis. How these conserved genes elicit differences in organ form and function in response to evolutionary pressures is incompletely understood. We molecularly and functionally characterized one member of an evolutionarily dynamic gene family, plac8 onzin related protein 1 (ponzr1), in the zebrafish. ponzr1 mRNA is expressed early in the developing kidney and pharyngeal arches. Using ponzr1-targeting morpholinos, we show that ponzr1 is required for formation of the glomerulus. Loss of ponzr1 results in a nonfunctional glomerulus but retention of a functional pronephros, an arrangement similar to the aglomerular kidneys found in a subset of marine fish. ponzr1 is integrated into the pax2a pathway, with ponzr1 expression requiring pax2a gene function, and proper pax2a expression requiring normal ponzr1 expression. In addition to pronephric function, ponzr1 is required for pharyngeal arch formation. We functionally demonstrate that ponzr1 can act as a transcription factor or co-factor, providing the first molecular mode of action for this newly described gene family. Together, this work provides experimental evidence of an additional mechanism that incorporates evolutionarily dynamic, lineage-specific gene families into conserved regulatory gene networks to create functional organ diversity.

Bedell, Victoria M.; Person, Anthony D.; Larson, Jon D.; McLoon, Anna; Balciunas, Darius; Clark, Karl J.; Neff, Kevin I.; Nelson, Katie E.; Bill, Brent R.; Schimmenti, Lisa A.; Beiraghi, Soraya; Ekker, Stephen C.

2012-01-01

226

Comparison of the Methyl Reductase Genes and Gene Products.  

National Technical Information Service (NTIS)

The DNA sequences encoding component C of methyl coenzyme M reductase (mcr genes) in Methanothermus fervidus, Methanobacterium thermoautotrophicum, ethanococcus vannielii, and Methanosarcina barkeri have been published. Comparisons of transcription initia...

C. F. Weil B. A. Sherf J. N. Reeve

1991-01-01

227

Combination of gamma radiation and essential oils from medicinal plants in managing Tribolium castaneum contamination of stored products.  

PubMed

Effectiveness of management of insect infestation of stored products with essential oils as viable alternatives to synthetic insecticides can be enhanced with gamma radiation. We studied effects of sublethal doses of essential oils from Rosmarinus officinalis (L.) and Perovskia atriplicifolia (Benth) (safe natural insecticides) in combination with gamma radiation on mortality of adults of Tribolium castaneum (Herbst). The insects were subjected to two radiation doses and two concentrations of the essential oils in the air. This combined treatment increased the mortality, which was also 3-6 times higher than could be expected from the sum of the effects of each of the treatments. The synergistic effect was more pronounced in the case of R. officinalis (L.) than in the case of P. atriplicifolia (Benth). The experiments have shown that the known insecticidal effectiveness of the essential oils can be enhanced by preliminary irradiation. Possible approaches to implementation of the combined treatment are discussed. PMID:23632647

Ahmadi, Mehrdad; Abd-alla, Adly Mohamed M; Moharramipour, Saeid

2013-03-28

228

Identification and isolation of genes essential for H sub 2 oxidation in Rhodobacter capsulatus. [Hydrogenase  

SciTech Connect

Mutants of Rhodobacter capsulatus unable to grow photoautotrophically with H{sub 2} and CO{sub 2} were isolated. Those lacking uptake hydrogenase activity as measured by H{sub 2}-dependent methylene blue reduction were analyzed genetically and used in complementation studies for the isolation of the wild-type genes. Results of further subcloning and transposon Tn5 mutagenesis suggest the involvement of a minimum of five genes. Hybridization to the 2.2-kilobase-pair SstI fragment that lies within the coding region for the large and small subunits of Bradyrhizobium japonicum uptake hydrogenase showed one region of strong homology among the R. capsulatus fragments isolated, which we interpret to mean that one or both structural genes were among the genes isolated.

Xu, H.W.; Love, J.; Borghese, R.; Wall, J.D. (Univ. of Missouri-Columbia (USA))

1989-02-01

229

The Major Immediate-Early Gene ie3 of Mouse Cytomegalovirus Is Essential for Viral Growth  

Microsoft Academic Search

The significance of the major immediate-early gene ie3 of mouse cytomegalovirus (MCMV) and that of the corresponding ie2 gene of human cytomegalovirus to viral replication are not known. To investigate the function of the MCMV IE3 regulatory protein, we generated two different MCMV recombinants that contained a large deletion in the IE3 open reading frame (ORF). The mutant genomes were

ANA ANGULO; PETER GHAZAL; MARTIN MESSERLE

2000-01-01

230

Rhizobitoxine production in Agrobacterium tumefaciens C58 by Bradyrhizobium elkanii rtxACDEFG genes.  

PubMed

We examined the genetic basis and transfer for production of rhizobitoxine, an inhibitor of ethylene biosynthesis in plants, directed by the rtx genes of Bradyrhizobium elkanii. Comparison with genome sequences of Bradyrhizobium japonicum and Xanthomonas oryzae suggests that the rtx genes extend from the previously identified rtxAC genes through four additional genes rtxDEFG. Reverse transcription-PCR analysis showed that the rtxACDEFG genes are expressed as an operon. Mutational analysis indicated that rtxDEG mutants reduced rhizobitoxine biosynthesis, while the rtxA gene is essential for its synthesis. Introduction of the rtxACDEFG into Agrobacterium tumefaciens resulted in strong expression of rtxACDEFG and production of RtxA protein, but no rhizobitoxine was detectable. Addition of O-acetylhomoserine, a precursor of rhizobitoxine, to the Agrobacterium derivative, however, fostered production of rhizobitoxine in culture. The diluted culture supernatant inhibited the activities of beta-cystathionase and 1-aminocyclopropane-1-carboxylate synthase, indicating that A. tumefaciens carrying rtxACDEFG genes excreted biologically active rhizobitoxine. PMID:17227467

Sugawara, Masayuki; Haramaki, Ryota; Nonaka, Satoko; Ezura, Hiroshi; Okazaki, Shin; Eda, Shima; Mitsui, Hisayuki; Minamisawa, Kiwamu

2007-01-15

231

A mutation in the Xanthomonas oryzae pv. oryzae wxoD gene affects xanthan production and chemotaxis.  

PubMed

Xanthomonas oryzae pv. oryzae causes bacterial blight in rice (Oryza sativa L.). The effect of a mutation in the wxoD gene, that encodes a putative O-antigen acetylase, on xanthan production as well as bacterial chemotaxis was investigated. The mutation increased xanthan production by 52 %. The mutant strain was non-motile on semi-solid agar swarm plates. In addition, several genes involved in chemotaxis, including the cheW, cheV, cheR, and cheD genes, were down-regulated by a mutation in the wxoD gene. Thus, the mutation in the wxoD gene affects xanthan production as well as bacterial chemotaxis. However, the wxoD gene is not essential for the virulence of X. oryzae. PMID:23881323

Nam, Jae-Young; Kim, Hong-Il; Lee, Chang-Soo; Park, Young-Jin

2013-07-24

232

Eukaryote to gut bacteria transfer of a glycoside hydrolase gene essential for starch breakdown in plants  

PubMed Central

Lateral gene transfer (LGT) between bacteria constitutes a strong force in prokaryote evolution, transforming the hierarchical tree of life into a network of relationships between species. In contrast, only a few cases of LGT from eukaryotes to prokaryotes have been reported so far. The distal animal intestine is predominantly a bacterial ecosystem, supplying the host with energy from dietary polysaccharides through carbohydrate-active enzymes absent from its genome. It has been suggested that LGT is particularly important for the human microbiota evolution. Here we show evidence for the first eukaryotic gene identified in multiple gut bacterial genomes. We found in the genome sequence of several gut bacteria, a typically eukaryotic glycoside-hydrolase necessary for starch breakdown in plants. The distribution of this gene is patchy in gut bacteria with presence otherwise detected only in a few environmental bacteria. We speculate that the transfer of this gene to gut bacteria occurred by a sequence of two key LGT events; first, an original eukaryotic gene was transferred probably from Archaeplastida to environmental bacteria specialized in plant polysaccharides degradation and second, the gene was transferred from the environmental bacteria to gut microbes.

Arias, Maria Cecilia; Danchin, Etienne G.J.; Coutinho, Pedro; Henrissat, Bernard; Ball, Steven

2012-01-01

233

Systemic Imbalance of Essential Metals and Cardiac Gene Expression in Rats Following Acute Pulmonary Zinc Exposure  

Microsoft Academic Search

It was recently demonstrated that particulate matter (PM) containing water-soluble zinc produces cardiac injury following pulmonary exposure. To investigate whether pulmonary zinc exposure produces systemic metal imbalance and direct cardiac effects, male Wistar Kyoto (WKY) rats (12–14 wk age) were intratracheally (IT) instilled with saline or 2 ?mol\\/kg zinc sulfate. Temporal analysis was performed for systemic levels of essential metals

Peter S. Gilmour; Mette C. Schladweiler; Abraham Nyska; John K. McGee; Ronald Thomas; Richard H. Jaskot; Judy Schmid; Urmila P. Kodavanti

2006-01-01

234

Cloning and characterization of rad21 an essential gene of Schizosaccharomyces pombe involved in DNA double-strand-break repair.  

PubMed Central

Analysis of the Schizosaccharomyces pombe chromosomes by pulsed field gel electrophoresis showed that the fission yeast has a very efficient DNA double-strand-break (dsb) repair system, which properly restores the three chromosomes after they are degraded by gamma-irradiation. The radiation-sensitive mutant rad21-45 is deficient in this repair pathway but is capable of cell-cycle arrest in G2 following DNA damage. We cloned the rad21 gene by complementing the radiation sensitivity of the rad21-45 mutant. The plasmid-borne gene completely reestablished the DNA dsb repair pathway. The rad21 gene was localized to chromosome III by hybridization. The transcript is 2.5 kb long and expressed at a moderate level. The 1884-bp open reading frame encodes a 628 amino acid, very acidic peptide with a calculated molecular mass of 67,854 D. The rad21 gene shows no significant homology to other known nucleotide or peptide sequences. The inability of the mutant to perform efficient DNA repair is caused by a single base substitution, which changes wild-type isoleucine67 into threonine in the mutant. Deletion of the genomic rad21 gene showed that it is essential for mitotic growth of S.pombe. Images

Birkenbihl, R P; Subramani, S

1992-01-01

235

Beyond the enhanceosome: cluster of novel ?B sites downstream of the human IFN-? gene is essential for lipopolysaccharide-induced gene activation.  

PubMed

The expression of interferon-? (IFN-?) in virus-infected HeLa cells established a paradigm of multifactorial gene regulation, in which cooperative assembly of transcription factors (TFs) at the composite DNA element (enhanceosome), is central for amplification of weak activating signals provided by individual TFs. However, whether the same TFs and the same DNA element are essential for IFN-? induction in response to bacterial stimuli are less well understood. Here we report that rapid and transient transcription of IFN-? in response to TLR4 stimulation with bacterial lipopolysaccharide (LPS) follows nuclear factor-?B (NF-?B) RelA activation and recruitment to the IFN-? genomic locus at multiple spatially separated regulatory regions. We demonstrate that the IFN-? enhanceosome region is not sufficient for maximal gene induction in response to LPS and identify an essential cluster of homotypic ?B sites in the 3' downstream of the gene. The cluster is characterized by elevated levels of histone 3 lysine 4 mono-methylation, a chromatin signature of enhancers, and efficiently binds RelA-containing NF-?B complexes in vitro and in vivo. These findings demonstrate that IFN-? gene activation via multifactorial enhanceosome assembly is potentiated in LPS-stimulated cells by NF-?B interactions with all functional ?B sites in the locus. PMID:20855868

Goh, Fui G; Thomson, Scott J P; Krausgruber, Thomas; Lanfrancotti, Alessandra; Copley, Richard R; Udalova, Irina A

2010-09-20

236

Fratricide is essential for efficient gene transfer between pneumococci in biofilms.  

PubMed

Streptococcus pneumoniae and a number of commensal streptococcal species are competent for natural genetic transformation. The natural habitat of these bacteria is multispecies biofilms in the human oral cavity and nasopharynx. Studies investigating lateral transfer of virulence and antibiotic resistance determinants among streptococci have shown that interspecies as well as intraspecies gene exchange takes place in these environments. We have previously shown that the action of a competence-specific murein hydrolase termed CbpD strongly increases the rate of gene transfer between pneumococci grown in liquid cultures. CbpD is the key component of a bacteriolytic mechanism termed the fratricide mechanism. It is secreted by competent pneumococci and mediates the release of donor DNA from sensitive streptococci present in the same environment. However, in nature, gene exchange between streptococci takes place in biofilms and not in liquid cultures. In the present study, we therefore investigated whether CbpD affects the rate of gene transfer in laboratory-grown biofilms. Our results show that the fratricide mechanism has a strong positive impact on intrabiofilm gene exchange, indicating that it is important for active acquisition of homologous donor DNA under natural conditions. Furthermore, we found that competent biofilm cells of S. pneumoniae acquire a Nov(r) marker much more efficiently from neighboring cells than from the growth medium. Efficient lysis of target cells requires that CbpD act in conjunction with the murein hydrolase LytC. In contrast, the major autolysin LytA does not seem to be important for fratricide-mediated gene exchange in a biofilm environment. PMID:22706053

Wei, Hua; Håvarstein, Leiv Sigve

2012-06-15

237

The Carney complex gene PRKAR1A plays an essential role in cardiac development and myxomagenesis.  

PubMed

Cardiac myxomas are the most common primary tumors of the heart, although little is known about their etiology. Mutations of the protein kinase A regulatory subunit gene PRKAR1A cause inherited myxomas in the setting of the Carney complex tumor syndrome, providing a possible window for understanding their pathogenesis. We recently reported that cardiac-specific knockout of this gene causes myxomatous changes in the heart, although the mice die during gestation from cardiac failure. In this review, we discuss these findings and place them in the larger understanding of how protein kinase A dysregulation might affect cardiac function and cause myxomagenesis. PMID:19577711

Yin, Zhirong; Kirschner, Lawrence S

2009-02-01

238

An essential role for the circadian-regulated gene Nocturnin in osteogenesis: the importance of local timekeeping in skeletal homeostasis  

PubMed Central

The role of circadian proteins in regulating whole body metabolism and bone turnover has been studied in detail and has led to the discovery of an elemental system for timekeeping involving the core genes Clock, Bmal1, Per, and Cry. Nocturnin, a peripheral circadian-regulated gene has been shown to play a very important role in regulating adipogenesis by deadenylation of key mRNAs and intra-cytoplasmic transport of PPAR?. The role that it plays in osteogenesis has previously not been studied in detail. In this report we examined in vitro and in vivo osteogenesis in the presence and absence of Nocturnin and show that loss of Nocturnin enhances bone formation and can rescue Rosiglitazone induced bone loss in mice. The circadian rhythm of Nocturnin is likely to be an essential element of marrow stromal cell fate.

Guntur, Anyonya R.; Kawai, Masanobu; Le, Phuong; Bouxsein, Mary L.; Bornstein, Sheila; Green, Carla B.; Rosen, Clifford J.

2012-01-01

239

An essential role for the circadian-regulated gene nocturnin in osteogenesis: the importance of local timekeeping in skeletal homeostasis.  

PubMed

The role of circadian proteins in regulating whole-body metabolism and bone turnover has been studied in detail and has led to the discovery of an elemental system for timekeeping involving the core genes Clock, Bmal1, Per, and Cry. Nocturnin?(Noc; Ccrn4l), a peripheral circadian-regulated gene has been shown to play a very important role in regulating adipogenesis by deadenylation of key mRNAs and intracytoplasmic transport of PPAR?. The role that it plays in osteogenesis has previously not been studied in detail. In this report we examined in vitro and in vivo osteogenesis in the presence and absence of Noc and show that loss of Noc enhances bone formation and can rescue rosiglitazone-induced bone loss in mice. The circadian rhythm of Noc is likely to be an essential element of marrow stromal cell fate. PMID:22082366

Guntur, Anyonya R; Kawai, Masanobu; Le, Phuong; Bouxsein, Mary L; Bornstein, Sheila; Green, Carla B; Rosen, Clifford J

2011-11-01

240

Fumigant toxicity and oviposition deterrency of the essential oil from cardamom, Elettaria cardamomum, against three stored–product insects.  

PubMed

Use of insecticides can have disruptive effects on the environment. Replacing the chemical compounds in these insecticides with plant materials, however, can be a safe method with low environmental risk. In the current study, chemical composition and insecticidal activities of the essential oil from cardamom, Elettaria cardamomum L. (Maton) (Zingiberales: Zingiberaceae) on the adults of three stored product pests was investigated. Results indicated that essential oil of E. cardamomum toxic to the bruchid beetle, Callosobruchus maculatus Fabricius (Coleoptera: Bruchidae), the red flour beetle, Tribolium castaneum Herbst (Coleoptera: Tenebrionidae), and the flour moth, Ephestia kuehniella Zeller (Lepidoptera: Pyralidae). Adults of E. kuehniella were more sensitive than the Coleoptera. Also, the highest mortality of these insects was seen after 12 hours. Results of the LT?? tests showed that the lethal time of mortality occurred between 10-20 hours in various test concentrations. Essential oil of E. cardamomum had a good efficacy on oviposition deterrence of C. maculatus females, too. The chemical constituents of the essential oils were analyzed by gas chromatography-mass spectrometry. The major constituents of cardamom were identified as 1,8-cineol, ?-terpinyl acetate, terpinene and fenchyl alcohol. These results suggest that essential oil of E. cardamomum is a good choice for control of stored product pests. PMID:22242564

Abbasipour, Habib; Mahmoudvand, Mohammad; Rastegar, Fahimeh; Hosseinpour, Mohammad Hossein

2011-01-01

241

Fumigant Toxicity and Oviposition Deterrency of the Essential Oil from Cardamom, Elettaria cardamomum, Against Three Stored--product Insects  

PubMed Central

Use of insecticides can have disruptive effects on the environment. Replacing the chemical compounds in these insecticides with plant materials, however, can be a safe method with low environmental risk. In the current study, chemical composition and insecticidal activities of the essential oil from cardamom, Elettaria cardamomum L. (Maton) (Zingiberales: Zingiberaceae) on the adults of three stored product pests was investigated. Results indicated that essential oil of E. cardamomum toxic to the bruchid beetle, Callosobruchus maculatus Fabricius (Coleoptera: Bruchidae), the red flour beetle, Tribolium castaneum Herbst (Coleoptera: Tenebrionidae), and the flour moth, Ephestia kuehniella Zeller (Lepidoptera: Pyralidae). Adults of E. kuehniella were more sensitive than the Coleoptera. Also, the highest mortality of these insects was seen after 12 hours. Results of the LT50 tests showed that the lethal time of mortality occurred between 10–20 hours in various test concentrations. Essential oil of E. cardamomum had a good efficacy on oviposition deterrence of C. maculatus females, too. The chemical constituents of the essential oils were analyzed by gas chromatography—mass spectrometry. The major constituents of cardamom were identified as 1,8-cineol, ?-terpinyl acetate, terpinene and fenchyl alcohol. These results suggest that essential oil of E. cardamomum is a good choice for control of stored product pests.

Abbasipour, Habib; Mahmoudvand, Mohammad; Rastegar, Fahimeh; Hosseinpour, Mohammad Hossein

2011-01-01

242

Molecular Cloning, Expression, and Characterization of the Genes Encoding the Two Essential Protein Components of Micrococcus luteus B-P 26 Hexaprenyl Diphosphate Synthase  

PubMed Central

The structural genes encoding the two essential components A and B of hexaprenyl diphosphate synthase, which produce the precursor of the prenyl side chain of menaquinone-6, were cloned from Micrococcus luteus B-P 26.

Shimizu, Naoto; Koyama, Tanetoshi; Ogura, Kyozo

1998-01-01

243

EXPORTIN1 Genes are Essential for Development and Function of the Gametophytes in Arabidopsis thaliana  

Technology Transfer Automated Retrieval System (TEKTRAN)

Gametes are produced in plants through mitotic divisions in the haploid gametophytes. We investigated the role of EXPORTIN1 (XPO1) genes during the development of both female and male gametophytes of Arabidopsis. Exportins exclude target proteins from the nucleus and are also part of a complex recru...

244

The DEAH-box helicase RHAU is an essential gene and critical for mouse hematopoiesis.  

PubMed

The DEAH helicase RHAU (alias DHX36, G4R1) is the only helicase shown to have G-quadruplex (G4)-RNA resolvase activity and the major source of G4-DNA resolvase activity. Previous report showed RHAU mRNA expression to be elevated in human lymphoid and CD34(+) BM cells, suggesting a potential role in hematopoiesis. Here, we generated a conditional knockout of the RHAU gene in mice. Germ line deletion of RHAU led to embryonic lethality. We then targeted the RHAU gene specifically in the hematopoiesis system, using a Cre-inducible system in which an optimized variant of Cre recombinase was expressed under the control of the Vav1 promoter. RHAU deletion in hematopoietic system caused hemolytic anemia and differentiation defect at the proerythroblast stage. The partial differentiation block of proerythroblasts was because of a proliferation defect. Transcriptome analysis of RHAU knockout proerythroblasts showed that a statistically significant portion of the deregulated genes contain G4 motifs in their promoters. This suggests that RHAU may play a role in the regulation of gene expression that relies on its G4 resolvase activity. PMID:22422825

Lai, Janice Ching; Ponti, Svetlana; Pan, Dejing; Kohler, Hubertus; Skoda, Radek C; Matthias, Patrick; Nagamine, Yoshikuni

2012-03-15

245

Twist is an essential regulator of the skeletogenic gene regulatory network in the sea urchin embryo  

Microsoft Academic Search

Recent work on the sea urchin endomesoderm gene regulatory network (GRN) offers many opportunities to study the specification and differentiation of each cell type during early development at a mechanistic level. The mesoderm lineages consist of two cell populations, primary and secondary mesenchyme cells (PMCs and SMCs). The micromere-PMC GRN governs the development of the larval skeleton, which is the

Shu-Yu Wu; Yu-Ping Yang; David R. McClay

2008-01-01

246

Genes for the peptidoglycan synthesis pathway are essential for chloroplast division in moss  

Microsoft Academic Search

The general consensus is that a cyanobacterium phagocytosed by a host cell evolved into the plastids of red and green algae, land plants, and glaucophytes. In contrast to the plastids of glaucophytes, which retain a cyanobacterial-type peptidoglycan layer, no wall-like structures have been detected in plastids from other sources. Although the genome of Arabidopsis thaliana contains five genes that are

Mariko Machida; Katsuaki Takechi; Hiroshi Sato; Sung Jin Chung; Haruko Kuroiwa; Susumu Takio; Motoaki Seki; Kazuo Shinozaki; Tomomichi Fujita; Mitsuyasu Hasebe; Hiroyoshi Takano

2006-01-01

247

Systematic Identification of Selective Essential Genes in Helicobacter pylori by Genome Prioritization and Allelic Replacement Mutagenesis  

Microsoft Academic Search

A comparative genomic approach was used to identify Helicobacter pylori 26695 open reading frames (ORFs) which are conserved in H. pylori J99 but highly diverged in other eubacteria. A survey of selected pathways of central intermediary metabolism was also carried out, and genes with a potentially selective role in H. pylori were identified. Forty-five ORFs identified in these two analyses

ALISON F. CHALKER; HEATHER W. MINEHART; NICKY J. HUGHES; KRISTIN K. KORETKE; MICHAEL A. LONETTO; KERRY K. BRINKMAN; PATRICK V. WARREN; ANDREI LUPAS; MICHAEL J. STANHOPE; JAMES R. BROWN; PAUL S. HOFFMAN

2001-01-01

248

Identification of Actinobacillus suis Genes Essential for the Colonization of the Upper Respiratory Tract of Swine  

PubMed Central

Actinobacillus suis has emerged as an important opportunistic pathogen of high-health-status swine. A colonization challenge method was developed, and using PCR-based signature-tagged transposon mutagenesis, 13 genes belonging to 9 different functional classes were identified that were necessary for A. suis colonization of the upper respiratory tract of swine.

Ojha, Shivani; Sirois, Marc; MacInnes, Janet I.

2005-01-01

249

Islands of non-essential genes, including a DNA translocation operon, in the genome of bacteriophage 0305?8-36.  

PubMed

We investigate genes of lytic, Bacillus thuringiensis bacteriophage 0305?8-36 that are non-essential for laboratory propagation, but might have a function in the wild. We isolate deletion mutants to identify these genes. The non-permutation of the genome (218.948 Kb, with a 6.479 Kb terminal repeat and 247 identified orfs) simplifies isolation of deletion mutants. We find two islands of non-essential genes. The first island (3.01% of the genomic DNA) has an informatically identified DNA translocation operon. Deletion causes no detectable growth defect during propagation in a dilute agarose overlay. Identification of the DNA translocation operon begins with a DNA relaxase and continues with a translocase and membrane-binding anchor proteins. The relaxase is in a family, first identified here, with homologs in other bacteriophages. The second deleted island (3.71% of the genome) has genes for two metallo-protein chaperonins and two tRNAs. Deletion causes a significant growth defect. In addition, (1) we find by "in situ" (in-plaque) single-particle fluorescence microscopy that adsorption to the host occurs at the tip of the 486 nm long tail, (2) we develop a procedure of 0305?8-36 purification that does not cause tail contraction, and (3) we then find by electron microscopy that 0305?8-36 undergoes tail tip-tail tip dimerization that potentially blocks adsorption to host cells, presumably with effectiveness that increases as the bacteriophage particle concentration increases. These observations provide an explanation of the previous observation that 0305?8-36 does not lyse liquid cultures, even though 0305?8-36 is genomically lytic. PMID:22666654

Pathria, Saurav; Rolando, Mandy; Lieman, Karen; Hayes, Shirley; Hardies, Stephen; Serwer, Philip

2012-01-01

250

appR gene product activates transcription of microcin C7 plasmid genes.  

PubMed Central

Microcin C7 (MccC7) is encoded by Escherichia coli plasmid pMccC7. However, some strains of E. coli K-12 carrying this plasmid do not produce this antibiotic. Here we show that these strains differ in the gene locus appR. This chromosomal gene product controls MccC7 production by activating the transcription of some, but not all, MccC7 plasmid genes.

Diaz-Guerra, L; Moreno, F; San Millan, J L

1989-01-01

251

Functions of the gene products of Escherichia coli.  

PubMed Central

A list of currently identified gene products of Escherichia coli is given, together with a bibliography that provides pointers to the literature on each gene product. A scheme to categorize cellular functions is used to classify the gene products of E. coli so far identified. A count shows that the numbers of genes concerned with small-molecule metabolism are on the same order as the numbers concerned with macromolecule biosynthesis and degradation. One large category is the category of tRNAs and their synthetases. Another is the category of transport elements. The categories of cell structure and cellular processes other than metabolism are smaller. Other subjects discussed are the occurrence in the E. coli genome of redundant pairs and groups of genes of identical or closely similar function, as well as variation in the degree of density of genetic information in different parts of the genome.

Riley, M

1993-01-01

252

Inhibitory effect of the essential oil of Curcuma longa L. and curcumin on aflatoxin production by Aspergillus flavus Link.  

PubMed

Aflatoxins are highly toxic, mutagenic, teratogenic and carcinogenic mycotoxins. Consumption of aflatoxin-contaminated food and commodities poses serious hazards to the health of humans and animals. Turmeric, Curcuma longa L., is a native plant of Southeast Asia and has antimicrobial, antioxidant and antifungal properties. This paper reports the antiaflatoxigenic activities of the essential oil of C. longa and curcumin. The medium tests were prepared with the oil of C. longa, and the curcumin standard at concentrations varied from 0.01% to 5.0%. All doses of the essential oil of the plant and the curcumin standard interfered with mycotoxin production. Both the essential oil and curcumin significantly inhibited the production of aflatoxins; the 0.5% level had a greater than 96% inhibitory effect. The levels of aflatoxin B(1) (AFB(1)) production were 1.0 and 42.7 ?g/mL, respectively, for the samples treated with the essential oil of C. longa L. and curcumin at a concentration of 0.5%. PMID:23122128

Ferreira, Flavio Dias; Kemmelmeier, Carlos; Arrotéia, Carla Cristina; da Costa, Christiane Luciana; Mallmann, Carlos Augusto; Janeiro, Vanderly; Ferreira, Francine Maery Dias; Mossini, Simone Aparecida Galerani; Silva, Expedito Leite; Machinski, Miguel

2012-08-10

253

A conditional mouse model for measuring the frequency of homologous recombination events in vivo in the absence of essential genes.  

PubMed

The ability to detect and repair DNA damage is crucial to the prevention of various diseases. Loss of function of genes involved in these processes is known to result in significant developmental defects and/or predisposition to cancer. One such DNA repair mechanism, homologous recombination, has the capacity to repair a wide variety of lesions. Knockout mouse models of genes thought to be involved in DNA repair processes are frequently lethal, making in vivo studies very difficult, if not impossible. Therefore, we set out to develop an in vivo conditional mouse model system to facilitate investigations into the involvement of essential genes in homologous recombination. To test our model, we measured the frequency of spontaneous homologous recombination using the pink-eyed unstable mouse model, in which we conditionally excised either Blm or full-length Brca1 (breast cancer 1, early onset). These two genes are hypothesized to have opposing roles in homologous recombination. In summary, our in vivo data supports in vitro studies suggesting that BLM suppresses homologous recombination, while full-length BRCA1 promotes this process. PMID:21709021

Brown, Adam D; Claybon, Alison B; Bishop, Alexander J R

2011-06-27

254

The MET13 Methylenetetrahydrofolate Reductase Gene Is Essential for Infection-Related Morphogenesis in the Rice Blast Fungus Magnaporthe oryzae  

PubMed Central

Methylenetetrahydrofolate reductases (MTHFRs) play a key role in the biosynthesis of methionine in both prokaryotic and eukaryotic organisms. In this study, we report the identification of a novel T-DNA-tagged mutant WH672 in the rice blast fungus Magnaporthe oryzae, which was defective in vegetative growth, conidiation and pathogenicity. Analysis of the mutation confirmed a single T-DNA insertion upstream of MET13, which encodes a 626-amino-acid protein encoding a MTHFR. Targeted gene deletion of MET13 resulted in mutants that were non-pathogenic and significantly impaired in aerial growth and melanin pigmentation. All phenotypes associated with ?met13 mutants could be overcome by addition of exogenous methionine. The M. oryzae genome contains a second predicted MTHFR-encoding gene, MET12. The deduced amino acid sequences of Met13 and Met12 share 32% identity. Interestingly, ?met12 mutants produced significantly less conidia compared with the isogenic wild-type strain and grew very poorly in the absence of methionine, but were fully pathogenic. Deletion of both genes resulted in ?met13?met12 mutants that showed similar phenotypes to single ?met13 mutants. Taken together, we conclude that the MTHFR gene, MET13, is essential for infection-related morphogenesis by the rice blast fungus M. oryzae.

Wang, Hong; Wang, Congcong; Li, Ya; Yue, Xiaofeng; Ma, Zhonghua; Talbot, Nicholas J.; Wang, Zhengyi

2013-01-01

255

A nuclear-encoded mitochondrial gene AtCIB22 is essential for plant development in Arabidopsis.  

PubMed

Complex I (the NADH:ubiquinone oxidoreductase) of the mitochondrial respiratory chain is a complicated, multi-subunit, membrane-bound assembly and contains more than 40 different proteins in higher plants. In this paper, we characterize the Arabidopsis homologue (designated as AtCIB22) of the B22 subunit of eukaryotic mitochondrial Complex I. AtCIB22 is a single-copy gene and is highly conserved throughout eukaryotes. AtCIB22 protein is located in mitochondria and the AtCIB22 gene is widely expressed in different tissues. Mutant Arabidopsis plants with a disrupted AtCIB22 gene display pleiotropic phenotypes including shorter roots, smaller plants and delayed flowering. Stress analysis indicates that the AtCIB22 mutants' seed germination and early seedling growth are severely inhibited by sucrose deprivation stress but more tolerant to ethanol stress. Molecular analysis reveals that in moderate knockdown AtCIB22 mutants, genes including cell redox proteins and stress related proteins are significantly up-regulated, and that in severe knockdown AtCIB22 mutants, the alternative respiratory pathways including NDA1, NDB2, AOX1a and AtPUMP1 are remarkably elevated. These data demonstrate that AtCIB22 is essential for plant development and mitochondrial electron transport chains in Arabidopsis. Our findings also enhance our understanding about the physiological role of Complex I in plants. PMID:21035093

Han, Lihua; Qin, Genji; Kang, Dingming; Chen, Zhangliang; Gu, Hongya; Qu, Li-Jia

2010-10-01

256

The MET13 Methylenetetrahydrofolate Reductase Gene Is Essential for Infection-Related Morphogenesis in the Rice Blast Fungus Magnaporthe oryzae.  

PubMed

Methylenetetrahydrofolate reductases (MTHFRs) play a key role in the biosynthesis of methionine in both prokaryotic and eukaryotic organisms. In this study, we report the identification of a novel T-DNA-tagged mutant WH672 in the rice blast fungus Magnaporthe oryzae, which was defective in vegetative growth, conidiation and pathogenicity. Analysis of the mutation confirmed a single T-DNA insertion upstream of MET13, which encodes a 626-amino-acid protein encoding a MTHFR. Targeted gene deletion of MET13 resulted in mutants that were non-pathogenic and significantly impaired in aerial growth and melanin pigmentation. All phenotypes associated with ?met13 mutants could be overcome by addition of exogenous methionine. The M. oryzae genome contains a second predicted MTHFR-encoding gene, MET12. The deduced amino acid sequences of Met13 and Met12 share 32% identity. Interestingly, ?met12 mutants produced significantly less conidia compared with the isogenic wild-type strain and grew very poorly in the absence of methionine, but were fully pathogenic. Deletion of both genes resulted in ?met13?met12 mutants that showed similar phenotypes to single ?met13 mutants. Taken together, we conclude that the MTHFR gene, MET13, is essential for infection-related morphogenesis by the rice blast fungus M. oryzae. PMID:24116181

Yan, Xia; Que, Yawei; Wang, Hong; Wang, Congcong; Li, Ya; Yue, Xiaofeng; Ma, Zhonghua; Talbot, Nicholas J; Wang, Zhengyi

2013-10-07

257

Transcriptional profiling of endogenous germ layer precursor cells identifies dusp4 as an essential gene in zebrafish endoderm specification.  

PubMed

A major goal for developmental biologists is to define the behaviors and molecular contents of differentiating cells. We have devised a strategy for isolating cells from diverse embryonic regions and stages in the zebrafish, using computer-guided laser photoconversion of injected Kaede protein and flow cytometry. This strategy enabled us to perform a genome-wide transcriptome comparison of germ layer precursor cells. Mesendoderm and ectoderm precursors cells isolated by this method differentiated appropriately in transplantation assays. Microarray analysis of these cells reidentified known genes at least as efficiently as previously reported strategies that relied on artificial mesendoderm activation or inhibition. We also identified a large set of uncharacterized mesendoderm-enriched genes as well as ectoderm-enriched genes. Loss-of-function studies revealed that one of these genes, the MAP kinase inhibitor dusp4, is essential for early development. Embryos injected with antisense morpholino oligonucleotides that targeted Dusp4 displayed necrosis of head tissues. Marker analysis during late gastrulation revealed a specific loss of sox17, but not of other endoderm markers, and analysis at later stages revealed a loss of foregut and pancreatic endoderm. This specific loss of sox17 establishes a new class of endoderm specification defect. PMID:18719100

Brown, Jamie L; Snir, Mirit; Noushmehr, Houtan; Kirby, Martha; Hong, Sung-Kook; Elkahloun, Abdel G; Feldman, Benjamin

2008-08-21

258

Analysis of the allelic diversity of a (CA)n repeat polymorphism among ?1-antitrypsin gene products from northern Portugal  

Microsoft Academic Search

The level of molecular heterogeneity associated with ?1-antitrypsin gene products was assessed in the population of northern\\u000a Portugal using three restriction fragment length polymorphisms (RFLPs) corresponding to specific amino acid substitutions\\u000a and a highly variable (CA)n repeat polymorphism located at the 5? end of the PI gene. The allelic affinities inferred from\\u000a the analysis of the DNA polymorphisms essentially agree

Jorge Rocha; Dalila Pinto; M. Teresa Santos; António Amorim; Jorge Amil-Dias; Fernando Cardoso-Rodrigues; Álvaro Aguiar

1997-01-01

259

The CDC8 gene product is required for transformation with episomal and integrative plasmids in Saccharomyces cerevisiae  

Microsoft Academic Search

The product of the yeast CDC8 gene (thymidylate kinase), which is required for chromosomal, mitochondrial and 2 µ plasmid replication, also participates in plasmid transformation processes in S. cerevisiae. The thermosensitive cdc8-1 mutant strain was transformed with episomal pDQ9 and integrative pDQ9-1 plasmids both of which carry the CDC8 gene. The results suggest that thymidylate kinase is essential for the

Beata Lecka-Czernik; Jerzy Zuk

1991-01-01

260

Staphylococcus aureus TargetArray: Comprehensive Differential Essential Gene Expression as a Mechanistic Tool To Profile Antibacterials? ¶  

PubMed Central

The widespread emergence of antibiotic-resistant bacteria and a lack of new pharmaceutical development have catalyzed a need for new and innovative approaches for antibiotic drug discovery. One bottleneck in antibiotic discovery is the lack of a rapid and comprehensive method to identify compound mode of action (MOA). Since a hallmark of antibiotic action is as an inhibitor of essential cellular targets and processes, we identify a set of 308 essential genes in the clinically important pathogen Staphylococcus aureus. A total of 446 strains differentially expressing these genes were constructed in a comprehensive platform of sensitized and resistant strains. A subset of strains allows either target underexpression or target overexpression by heterologous promoter replacements with a suite of tetracycline-regulatable promoters. A further subset of 236 antisense RNA-expressing clones allows knockdown expression of cognate targets. Knockdown expression confers selective antibiotic hypersensitivity, while target overexpression confers resistance. The antisense strains were configured into a TargetArray in which pools of sensitized strains were challenged in fitness tests. A rapid detection method measures strain responses toward antibiotics. The TargetArray antibiotic fitness test results show mechanistically informative biological fingerprints that allow MOA elucidation.

Xu, H. Howard; Trawick, John D.; Haselbeck, Robert J.; Forsyth, R. Allyn; Yamamoto, Robert T.; Archer, Rich; Patterson, Joe; Allen, Molly; Froelich, Jamie M.; Taylor, Ian; Nakaji, Danny; Maile, Randy; G. C., Kedar; Pilcher, Marshall; Brown-Driver, Vickie; McCarthy, Melissa; Files, Amy; Robbins, David; King, Paula; Sillaots, Susan; Malone, Cheryl; Zamudio, Carlos S.; Roemer, Terry; Wang, Liangsu; Youngman, Philip J.; Wall, Daniel

2010-01-01

261

TIF1-GAMMA PLAYS AN ESSENTIAL ROLE IN MURINE HEMATOPOIESIS AND REGULATES TRANSCRIPTIONAL ELONGATION OF ERYTHROID GENES  

PubMed Central

Transcriptional regulators play critical roles in the regulation of cell fate during hematopoiesis. Previous studies in zebrafish have identified an essential role for the transcriptional intermediary factor TIF1? in erythropoiesis through regulating the transcription elongation of erythroid genes. To study if TIF1? plays a similar role in murine erythropoiesis and to assess its function in other blood lineages, we generated mouse models with hematopoietic deletion of TIF1?. Our results showed a block in erythroid maturation in the bone marrow following tif1? deletion that was compensated with enhanced spleen erythropoiesis. Further analyses revealed a defect in transcription elongation of erythroid genes in the bone marrow. In addition, loss of TIF1? resulted in defects in other blood compartments, including a profound loss of B cells, a dramatic expansion of granulocytes and decreased HSC function. TIF1? exerts its functions in a cell-autonomous manner as revealed by competitive transplantation experiments. Our study therefore demonstrates that TIF1? plays essential roles in multiple murine blood lineages and that its function in transcription elongation is evolutionally conserved.

Bai, Xiaoying; Trowbridge, Jennifer J.; Riley, Elizabeth; Lee, Joseph A.; DiBiase, Anthony; Kaartinen, Vesa M.; Orkin, Stuart H.; Zon, Leonard I.

2012-01-01

262

Interacting contribution of the five polymorphisms in three genes of Hsp70 family to essential hypertension in Uygur ethnicity  

PubMed Central

Experimental evidence suggesting that heat shock protein 70 (Hsp70) gene or associated genes are responsible for the pathophysiology of hypertension is accumulating. In this study, we focused on five polymorphisms in three genes (HSPA1A, HSPA1B, and HSPA1L) of Hsp70 family to explore the genetic contribution, alone and in combination, of these polymorphisms to essential hypertension risk in a Uygur population. Genotyping was performed using PCR-RFLP and direct sequencing techniques. Data were analyzed using haplotype and multifactor dimensionality reduction (MDR) methods. Genotype distributions of all the polymorphisms satisfied the Hardy–Weinberg proportions in cases and controls. Statistical significance was only observed in the genotype (P?=?0.0028) and (P?=?0.0146) allele distributions of ?110A/C polymorphism, with the ?110C allele conferring a 1.45- and 2.83-fold of relative risk, assuming the additive and recessive models, respectively, and in 1267A/G genotype distribution (P?=?0.0106) with the 1267G allele conferring a 44% reduced risk. The interaction information analysis indicated that polymorphisms ?110A/C and 1267A/G had a strong synergistic effect, while polymorphisms 2074G/C and 2437T/C had a moderate synergistic effect. Haplotype analyses further strengthened the interaction information. Using the haplotype H1 as a reference, haplotype H4 had a 40% reduced risk, while haplotypes H5 and H8 had a significantly 5.00- and 3.75-fold increased risk for essential hypertension, respectively. Taken together, our results supported strong genetic interaction of the studied polymorphisms with the risk of having essential hypertension in Uygur ethnicity. Functional studies are warranted to confirm or refute these findings. This is the first study to evaluate the genetic interaction information of the Hsp70 in Uygur ethnicity, which represents one of the major nationalities in China with high homogeneity and unique lifestyles. Moreover, we employed the haplotype and MDR methods to explore the potential interaction of Hsp70 genetic polymorphisms in the pathogenesis of essential hypertension in Uygur.

Li, Jin-Xin; Tang, Bao-Peng; Sun, Hui-Ping; Feng, Min

2008-01-01

263

Identification of a Gene Essential for Sheathed Structure Formation in Sphaerotilus natans, a Filamentous Sheathed Bacterium  

Microsoft Academic Search

Sphaerotilus natans, a filamentous bacterium that causes bulking in activated sludge processes, can assume two distinct morphologies, depending on the substrate concentration for growth; in substrate-rich media it grows as single rod-shaped cells, whereas in substrate-limited media it grows as filaments. To identify genes responsible for sheath formation, we carried out transposon Tn5 mutagenesis. Of the approximately 20,000 mutants obtained,

Toshihiko Suzuki; Takahiro Kanagawa; Yoichi Kamagata

2002-01-01

264

The Zebrafish moonshine Gene Encodes Transcriptional Intermediary Factor 1?, an Essential Regulator of Hematopoiesis  

Microsoft Academic Search

Hematopoiesis is precisely orchestrated by lineage-specific DNA-binding proteins that regulate transcription in concert with coactivators and corepressors. Mutations in the zebrafish moonshine (mon) gene specifically disrupt both embryonic and adult hematopoiesis, resulting in severe red blood cell aplasia. We report that mon encodes the zebrafish ortholog of mammalian transcriptional intermediary factor 1? (TIF1?) (or TRIM33), a member of the TIF1

David G Ransom; Nathan Bahary; Knut Niss; David Traver; Caroline Burns; Nikolaus S Trede; Noelle Paffett-Lugassy; Walter J Saganic; C. Anthoney Lim; Candace Hersey; Yi Zhou; Bruce A Barut; Shuo Lin; Paul D Kingsley; James Palis; Stuart H Orkin; Leonard I Zon

2004-01-01

265

Identification of viral genes essential for replication of murine -herpesvirus 68 using signature-tagged mutagenesis  

Microsoft Academic Search

-Herpesviruses, Epstein-Barr virus, and Kaposi's sarcoma-associated herpesvirus are important human pathogens, because they are involved in tumor development. Murine -herpesvirus-68 (MHV-68 or HV-68) has emerged as a small animal model system for the study of -herpesvirus pathogenesis and host-virus interactions. To identify the genes required for viral replication in vitro and in vivo, we generated 1,152 mutants using signature-tagged transposon

Moon Jung Song; Seungmin Hwang; Wendy H. Wong; Ting-Ting Wu; Sangmi Lee; Hsiang-I. Liao; Ren Sun

2005-01-01

266

Identification of a Gene Essential for Sheathed Structure Formation in Sphaerotilus natans, a Filamentous Sheathed Bacterium  

PubMed Central

Sphaerotilus natans, a filamentous bacterium that causes bulking in activated sludge processes, can assume two distinct morphologies, depending on the substrate concentration for growth; in substrate-rich media it grows as single rod-shaped cells, whereas in substrate-limited media it grows as filaments. To identify genes responsible for sheath formation, we carried out transposon Tn5 mutagenesis. Of the approximately 20,000 mutants obtained, 7 did not form sheathed structures. Sequencing of the Tn5-flanking regions showed that five of the seven Tn5 insertions converged at the same open reading frame, designated sthA. The deduced amino acids encoded by sthA were found to be homologous to glycosyltransferase, which is known to be involved in linking sugars to lipid carriers during bacterial exopolysaccharide biosynthesis. Disruption of the gene of the wild-type strain by inserting a kanamycin resistance gene cassette also resulted in sheathless growth under either type of nutrient condition. These findings indicate that sthA is a crucial component responsible for sheath formation.

Suzuki, Toshihiko; Kanagawa, Takahiro; Kamagata, Yoichi

2002-01-01

267

ideR, an Essential Gene in Mycobacterium tuberculosis: Role of IdeR in Iron-Dependent Gene Expression, Iron Metabolism, and Oxidative Stress Response†  

PubMed Central

The mycobacterial IdeR protein is a metal-dependent regulator of the DtxR (diphtheria toxin repressor) family. In the presence of iron, it binds to a specific DNA sequence in the promoter regions of the genes that it regulates, thus controlling their transcription. In this study, we provide evidence that ideR is an essential gene in Mycobacterium tuberculosis. ideR cannot normally be disrupted in this mycobacterium in the absence of a second functional copy of the gene. However, a rare ideR mutant was obtained in which the lethal effects of ideR inactivation were alleviated by a second-site suppressor mutation and which exhibited restricted iron assimilation capacity. Studies of this strain and a derivative in which IdeR expression was restored allowed us to identify phenotypic effects resulting from ideR inactivation. Using DNA microarrays, the iron-dependent transcriptional profiles of the wild-type, ideR mutant, and ideR-complemented mutant strains were analyzed, and the genes regulated by iron and IdeR were identified. These genes encode a variety of proteins, including putative transporters, proteins involved in siderophore synthesis and iron storage, members of the PE/PPE family, a membrane protein involved in virulence, transcriptional regulators, and enzymes involved in lipid metabolism.

Rodriguez, G. Marcela; Voskuil, Martin I.; Gold, Benjamin; Schoolnik, Gary K.; Smith, Issar

2002-01-01

268

Cytomegalovirus-mediated induction of antisense mRNA expression to UL44 inhibits virus replication in an astrocytoma cell line: identification of an essential gene.  

PubMed Central

We have used an antisense RNA approach in the analysis of gene function in human cytomegalovirus (HCMV). An astrocytoma cell line (U373-MG) that is permissive for virus replication was permanently transfected with a construct bearing sequence from HCMV UL44 (coding for the major late DNA-binding protein, ppUL44, also known as pp52 or ICP36) in an antisense orientation and under the control of the immediate-early enhancer-promoter element. Upon HCMV infection at a high multiplicity, we found a marked reduction in UL44 protein products (the ICP36 family of proteins) in established cell transfectants and a strong inhibition of virus yield in infected-cell supernatants at two weeks postinfection, while herpes simplex virus replication was not affected. In infected cells, viral DNA replication was strongly inhibited. While gene products such as pUS22 and pUL32 were also inhibited, pUL123 and pUL82 accumulated in the infected cells over time. Our data suggest an essential role for the UL44 family of proteins in HCMV replication and represent a model of virus inhibition by virus-induced antisense RNA synthesis in genetically modified cells.

Ripalti, A; Boccuni, M C; Campanini, F; Landini, M P

1995-01-01

269

Functional characterisation of the non-essential protein kinases and phosphatases regulating Aspergillus nidulans hydrolytic enzyme production  

PubMed Central

Background Despite recent advances in the understanding of lignocellulolytic enzyme regulation, less is known about how different carbon sources are sensed and the signaling cascades that result in the adaptation of cellular metabolism and hydrolase secretion. Therefore, the role played by non-essential protein kinases (NPK) and phosphatases (NPP) in the sensing of carbon and/or energetic status was investigated in the model filamentous fungus Aspergillus nidulans. Results Eleven NPKs and seven NPPs were identified as being involved in cellulase, and in some cases also hemicellulase, production in A. nidulans. The regulation of CreA-mediated carbon catabolite repression (CCR) in the parental strain was determined by fluorescence microscopy, utilising a CreA::GFP fusion protein. The sensing of phosphorylated glucose, via the RAS signalling pathway induced CreA repression, while carbon starvation resulted in derepression. Growth on cellulose represented carbon starvation and derepressing conditions. The involvement of the identified NPKs in the regulation of cellulose-induced responses and CreA derepression was assessed by genome-wide transcriptomics (GEO accession 47810). CreA::GFP localisation and the restoration of endocellulase activity via the introduction of the ?creA mutation, was assessed in the NPK-deficient backgrounds. The absence of either the schA or snfA kinase dramatically reduced cellulose-induced transcriptional responses, including the expression of hydrolytic enzymes and transporters. The mechanism by which these two NPKs controlled gene transcription was identified, as the NPK-deficient mutants were not able to unlock CreA-mediated carbon catabolite repression under derepressing conditions, such as carbon starvation or growth on cellulose. Conclusions Collectively, this study identified multiple kinases and phosphatases involved in the sensing of carbon and/or energetic status, while demonstrating the overlapping, synergistic roles of schA and snfA in the regulation of CreA derepression and hydrolytic enzyme production in A. nidulans. The importance of a carbon starvation-induced signal for CreA derepression, permitting transcriptional activator binding, appeared paramount for hydrolase secretion.

2013-01-01

270

Associations between leptin Gene polymorphisms and production  

Microsoft Academic Search

Leptin is a 16-kDa protein synthesized by adipose tissue and is involved in regulation of feed intake, energy balance, fertility, and immune functions. Since evidence of a genetic correlation between start of luteal activity and energy balance, milk yield, and live weights is present, we investigated the association of genetic differences in the bovine leptin gene with these traits. Between

S. C. Liefers; Pas te M. F. W; R. F. Veerkamp; Lende van der T

2002-01-01

271

Post transcriptional regulation of chloroplast gene expression by nuclear encoded gene products  

SciTech Connect

Many individual chloroplast genes require the products of a collection of nuclear genes for their successful expression. These nuclear gene products apparently work with great specificity, each committed to the expression of a single chloroplast gene. We have chosen as a model nuclear mutants of Chlamydomonas affected in different stages in the expression of the chloroplast encoded Photosystem II polypeptide, D2. We have made the progress in understanding how nuclear gene products affect the translation of the D2 encoding MRNA. Two nuclear genes are required for this process which have been mapped genetically. In contrast to other examples of nuclear control of translation in the chloroplast, these nuclear gene products appear to be required either for specific stages in translation elongation or for the post-translational stabilization of the nascent D2 protein. Pseudoreversion analysis has led us to a locus which may be directly involved in D2 expression. We have made considerable progress in pursuing the molecular basis of psbd MRNA stabilization. psbD 5' UTR specific transcripts have been synthesized in vitro and used in gel mobility shift assays. UV-crosslinking studies are underway to identify the transacting factors which bind to these sequences. The continued examination of these mutants will help us to understand how nuclear gene products work in this specific case of chloroplast gene expression, and will elucidate how two distinct genomes can interact generally.

Kuchka, M.R.

1992-01-01

272

RelA alone appears essential for (p)ppGpp production when Neisseria gonorrhoeae encounters nutritional stress  

Microsoft Academic Search

The bacterial stringent response is a pleiotrophic physiological response that is evoked when bacteria are subjected to nutrient stress and is mediated through the accumulation of hyperphosphorylated guanine nucleotides ((p)ppGpp) which are synthesized by the combined action of the relA and spoT gene products. The relA and spoT genes were cloned from Neisseria gonorrhoeae strain MS11 and various insertional and

Scott D. Fisher; Andrew D. Reger; Atalie Baum; Stuart A. Hill

2005-01-01

273

Association between Polymorphisms in the Renin-Angiotensin-Aldosterone System Genes and Essential Hypertension in the Han Chinese Population  

PubMed Central

Background Renin-angiotensin-aldosterone system (RAAS) is the most important endocrine blood pressure control mechanism in our body, genes encoding components of this system have been strong candidates for the investigation of the genetic basis of hypertension. However, previous studies mainly focused on limited polymorphisms, thus we carried out a case-control study in the Han Chinese population to systemically investigate the association between polymorphisms in the RAAS genes and essential hypertension. Methods 905 essential hypertensive cases and 905 normotensive controls were recruited based on stringent inclusion and exclusion criteria. All 41 tagSNPs within RAAS genes were retrieved from HapMap, and the genotyping was performed using the GenomeLab SNPstream Genotyping System. Logistic regression analysis, Multifactor dimensionality reduction (MDR), stratified analysis and crossover analysis were used to identify and characterize interactions among the SNPs and the non-genetic factors. Results Serum levels of total cholesterol (TC) and triglyceride (TG), and body mass index (BMI) were significantly higher in the hypertensive group than in the control group. Of 41 SNPs genotyped, rs3789678 and rs2493132 within AGT, rs4305 within ACE, rs275645 within AGTR1, rs3802230 and rs10086846 within CYP11B2 were shown to associate with hypertension. The MDR analysis demonstrated that the interaction between BMI and rs4305 increased the susceptibility to hypertension. Crossover analysis and stratified analysis further indicated that BMI has a major effect, and rs4305 has a minor effect. Conclusion These novel findings indicated that together with non-genetic factors, these genetic variants in the RAAS may play an important role in determining an individual’s susceptibility to hypertension in the Han Chinese.

Zhang, Lina; Fei, Lijuan; Wang, Lin; Su, Jia; Lazar, Lissy; Xu, Jin; Zhang, Yaping

2013-01-01

274

SENSORY EVALUATION OF CITRUS PEEL ESSENTIAL OILS AS FLAVOURING AGENTS IN VARIOUS FOOD PRODUCTS  

Microsoft Academic Search

A study was conducted at the Institute of Food Science and Technology, University of Agriculture, Faisalabad during 2002-2003. The objective was to evaluate the utility of peel essential oils of various Pakistani citrus varieties such as Kinnow (C. reticulata, var. mandarin), Fewtrell's Early (C. reticulata, var. tangenrine), Malta (C. sinensis var. malta), Mousami (C. sinensis var. mousami), grape fruit (C.

Muhammad Mushtaq Ahmad

275

Sperm-associated antigen-17 gene is essential for motile cilia function and neonatal survival.  

PubMed

Primary ciliary dyskinesia (PCD), resulting from defects in cilia assembly or motility, is caused by mutations in a number of genes encoding axonemal proteins. PCD phenotypes are variable, and include recurrent respiratory tract infections, bronchiectasis, hydrocephaly, situs inversus, and male infertility. We generated knockout mice for the sperm-associated antigen-17 (Spag17) gene, which encodes a central pair (CP) protein present in the axonemes of cells with "9 + 2" motile cilia or flagella. The targeting of Spag17 resulted in a severe phenotype characterized by immotile nasal and tracheal cilia, reduced clearance of nasal mucus, profound respiratory distress associated with lung fluid accumulation and disruption of the alveolar epithelium, cerebral ventricular expansion consistent with emerging hydrocephalus, failure to suckle, and neonatal demise within 12 hours of birth. Ultrastructural analysis revealed the loss of one CP microtubule in approximately one quarter of tracheal cilia axonemes, an absence of a C1 microtubule projection, and other less frequent CP structural abnormalities. SPAG6 and SPAG16 (CP proteins that interact with SPAG17) were increased in tracheal tissue from SPAG17-deficient mice. We conclude that Spag17 plays a critical role in the function and structure of motile cilia, and that neonatal lethality is likely explained by impaired airway mucociliary clearance. PMID:23418344

Teves, Maria Eugenia; Zhang, Zhibing; Costanzo, Richard M; Henderson, Scott C; Corwin, Frank D; Zweit, Jamal; Sundaresan, Gobalakrishnan; Subler, Mark; Salloum, Fadi N; Rubin, Bruce K; Strauss, Jerome F

2013-06-01

276

The Cebpd (C\\/EBP?) Gene Is Induced by Luteinizing Hormones in Ovarian Theca and Interstitial Cells But Is Not Essential for Mouse Ovary Function  

Microsoft Academic Search

The CCAAT\\/enhancer binding protein (CEBP) family of transcription factors includes five genes. In the ovary, both Cebpa and Cebpb are essential for granulosa cell function. In this study we have explored the role of the Cebpd gene in ovarian physiology by expression and functional studies. Here we report that Cebpd (C\\/EBP?) is expressed in the mouse ovary in a highly

A-Mei Huang; Martina Rudelius; Shikha Sharan; Jan M. McAllister; Mark Raffeld; Esta Sterneck

2007-01-01

277

Cellular Tissue and Gene Therapies: Product Development ...  

Center for Biologics Evaluation and Research (CBER)

Text Version... Ps); Animal models for porcine xenotransplantation products intended to treat Type 1 diabetes or acute liver failure; Clinical ... More results from www.fda.gov/downloads/biologicsbloodvaccines/newsevents

278

Estrogen-related receptor {alpha} is essential for the expression of antioxidant protection genes and mitochondrial function  

SciTech Connect

Estrogen-related receptor {alpha} (ERR{alpha}) is an important mediator of mitochondrial biogenesis and function. To investigate the transcriptional network controlling these phenomena, we investigated mitochondrial gene expression in embryonic fibroblasts isolated from ERR{alpha} null mice. Peroxisome proliferator-activated receptor {gamma} coactivator-1{alpha} (PGC-1{alpha}) stimulated mitochondrial gene expression program in control cells, but not in the ERR{alpha} null cells. Interestingly, the induction of levels of mitochondrial oxidative stress protection genes in response to increased PGC-1{alpha} levels was dependent on ERR{alpha}. Furthermore, we found that the PGC-1{alpha}-mediated induction of estrogen-related receptor {gamma} and nuclear respiratory factor 2 (NRF-2), was dependent on the presence of ERR{alpha}. Basal levels of NRF-2 were decreased in the absence of ERR{alpha}. The absence of ERR{alpha} resulted in a decrease in citrate synthase enzyme activity in response to PGC-1{alpha} overexpression. Our results indicate an essential role for ERR{alpha} as a key regulator of oxidative metabolism.

Rangwala, Shamina M. [Diabetes and Metabolism Disease Area, Novartis Institutes of BioMedical Research Institutes, 100 Technology Square, Cambridge, MA 02139 (United States)]. E-mail: shamina.rangwala@novartis.com; Li, Xiaoyan [Diabetes and Metabolism Disease Area, Novartis Institutes of BioMedical Research Institutes, 100 Technology Square, Cambridge, MA 02139 (United States); Lindsley, Loren [Diabetes and Metabolism Disease Area, Novartis Institutes of BioMedical Research Institutes, 100 Technology Square, Cambridge, MA 02139 (United States); Wang, Xiaomei [Diabetes and Metabolism Disease Area, Novartis Institutes of BioMedical Research Institutes, 100 Technology Square, Cambridge, MA 02139 (United States); Shaughnessy, Stacey [Diabetes and Metabolism Disease Area, Novartis Institutes of BioMedical Research Institutes, 100 Technology Square, Cambridge, MA 02139 (United States); Daniels, Thomas G. [Diabetes and Metabolism Disease Area, Novartis Institutes of BioMedical Research Institutes, 100 Technology Square, Cambridge, MA 02139 (United States); Szustakowski, Joseph [Genome and Proteome Sciences, Novartis Institutes of BioMedical Research Institutes, 500 Technology Square, Cambridge, MA 02139 (United States); Nirmala, N.R. [Genome and Proteome Sciences, Novartis Institutes of BioMedical Research Institutes, 500 Technology Square, Cambridge, MA 02139 (United States); Wu, Zhidan [Diabetes and Metabolism Disease Area, Novartis Institutes of BioMedical Research Institutes, 100 Technology Square, Cambridge, MA 02139 (United States); Stevenson, Susan C. [Diabetes and Metabolism Disease Area, Novartis Institutes of BioMedical Research Institutes, 100 Technology Square, Cambridge, MA 02139 (United States)

2007-05-25

279

Support for the minimal essential MHC hypothesis: a parrot with a single, highly polymorphic MHC class II B gene.  

PubMed

We characterized the MHC class II B gene in the green-rumped parrotlet, Forpus passerinus. Three approaches were used: polymerase chain reaction amplification using primers complementary to conserved regions of exon 2, sequencing clones from a genomic library, and amplification of exon 2 using species-specific primers. All three methods indicate that there is only a single class II B locus in this species and no pseudogenes. We suggest that this is the ancestral state for birds. The gene is highly polymorphic; 33 alleles were found in a sample of 25 individuals. Variation in exon 2 is concentrated in the peptide binding residues which show a significant excess of non-synonymous substitutions consistent with the operation of selection in maintaining this extraordinary polymorphism. Genomic clones show that major histocompatibility complex (MHC) gene organization is different from that of chickens; the class II A locus is close to II B. These data provide support for the hypothesis that the bird MHC constitutes a "minimal essential MHC" for responding to infectious disease. PMID:18431567

Hughes, Colin R; Miles, Shana; Walbroehl, Jaclyn M

2008-04-23

280

Identification of act2, an essential gene in the fission yeast Schizosaccharomyces pombe that encodes a protein related to actin.  

PubMed Central

Actins are a family of highly conserved proteins that are ubiquitously found among eukaryotic organisms. All actins that have previously been identified, including those of animals, plants, fungi, and protozoa, are 374-376 amino acids long and exhibit at least 70% amino acid sequence identity when compared with one another. We have cloned a gene from the fission yeast Schizosaccharomyces pombe that encodes a distantly related member of the actin protein family, herein referred to as act2. In contrast to all other actins, the derived amino acid sequence reveals that act2 is 427 residues long and exhibits only 35-40% identity to actins, including act1 from Sch. pombe. Comparison to the known x-ray crystallographic structure of rabbit skeletal muscle actin indicates that the ATP and divalent metal ion binding sites are largely conserved in act2, while regions involved in actin-actin and actin-myosin interactions are relatively divergent. Disruption of the act2 gene demonstrated that this gene encodes a function essential for germination of haploid spores. These findings indicate that while act2 and act1 are related proteins, they appear to have distinct functions. In addition, they demonstrate that the actin protein family is more diverse than was previously thought. Images

Lees-Miller, J P; Henry, G; Helfman, D M

1992-01-01

281

Gene expression profiling with principal component analysis depicts the biological continuum from essential thrombocythemia over polycythemia vera to myelofibrosis.  

PubMed

The recent discovery of the Janus activating kinase 2 V617F mutation in most patients with polycythemia vera (PV) and half of those with essential thrombocythemia (ET) and primary myelofibrosis (PMF) has favored the hypothesis of a biological continuum from ET over PV to PMF. We performed gene expression profiling of whole blood from control subjects (n = 21) and patients with ET (n = 19), PV (n = 41), and PMF (n = 9) using DNA microarrays. Applying an unsupervised method, principal component analysis, to search for patterns in the data, we demonstrated a separation of the four groups with biological relevant overlaps between the different entities. Moreover, the analysis separates Janus activating kinase 2-negative ET patients from Janus activating kinase 2-positive ET patients. Functional annotation analysis demonstrates that clusters of gene ontology terms related to inflammation, immune system, apoptosis, RNA metabolism, and secretory system were the most significantly deregulated terms in the three different disease groups. Our results yield further support for the hypothesis of a biological continuum originating from ET over PV to PMF. Functional analysis suggests an important implication of these gene ontology clusters in the pathogenesis of these neoplasms and in disease evolution from ET over PV to PMF. PMID:22659388

Skov, Vibe; Thomassen, Mads; Riley, Caroline H; Jensen, Morten K; Bjerrum, Ole Weis; Kruse, Torben A; Hasselbalch, Hans Carl; Larsen, Thomas Stauffer

2012-06-01

282

An essential gene, ESR1, is required for mitotic cell growth, DNA repair and meiotic recombination in Saccharomyces cerevisiae.  

PubMed Central

A new mutant, which was sensitive to both methyl-methanesulfonate (MMS) and ultra-violet light (UV) and defective in meiotic recombination, was isolated from Saccharomyces cerevisiae. The gene, ESR1, was cloned by complementation of the MMS sensitivity of the mutant and found to be essential for cell growth, as the deleted haploid strain was lethal. The ESR1 gene was adjacent to the CKS1 gene on chromosome II and encoded a putative 2368-amino acid protein with a molecular weight of 273 k. The ESR1 transcript was 8.0 kb long and was induced during meiosis. The predicted Esr1 protein had a mosaic structure composed of homologous regions and showed amino acid sequence similarities to Schizosaccharomyces pombe rad3+ protein, which monitors completion of DNA repair synthesis, and cut1+ protein, which is required for spindle pole body (SPB) duplication. The Esr1 protein was also similar to phosphatidylinositol (PI) 3-kinases, including Saccharomyces cerevisiae TOR2 (and DRR1), which are involved in G1 progression. These results suggest that ESR1 is multi-functional throughout mitosis and meiosis. Images

Kato, R; Ogawa, H

1994-01-01

283

Identification of viral genes essential for replication of murine gamma-herpesvirus 68 using signature-tagged mutagenesis.  

PubMed

Gamma-herpesviruses, Epstein-Barr virus, and Kaposi's sarcoma-associated herpesvirus are important human pathogens, because they are involved in tumor development. Murine gamma-herpesvirus-68 (MHV-68 or gammaHV-68) has emerged as a small animal model system for the study of gamma-herpesvirus pathogenesis and host-virus interactions. To identify the genes required for viral replication in vitro and in vivo, we generated 1,152 mutants using signature-tagged transposon mutagenesis on an infectious bacterial artificial chromosome of MHV-68. Almost every ORF was mutated by random insertion. For each ORF, a mutant with an insertion proximal to the N terminus of each ORF was examined for the ability to grow in fibroblasts. Our results indicate that 41 genes are essential for in vitro growth, whereas 26 are nonessential and 6 attenuated. Replication-competent mutants were pooled to infect mice, which led to the discovery of ORF 54 being important for MHV-68 to replicate in the lung. This genetic analysis of a tumor-associated herpesvirus at the whole genome level validates signature-tagged transposon mutagenesis screening as an effective genetic system to identify important virulent genes in vivo and define interactions with the host immune system. PMID:15738413

Song, Moon Jung; Hwang, Seungmin; Wong, Wendy H; Wu, Ting-Ting; Lee, Sangmi; Liao, Hsiang-I; Sun, Ren

2005-02-28

284

The Drosophila Clathrin Heavy Chain Gene: Clathrin Function Is Essential in a Multicellular Organism  

PubMed Central

The clathrin heavy chain (HC) is the major structural polypeptide of the cytoplasmic surface lattice of clathrin-coated pits and vesicles. As a genetic approach to understanding the role of clathrin in cellular morphogenesis and developmental signal transduction, a clathrin heavy chain (Chc) gene of Drosophila melanogaster has been identified by a combination of molecular and classical genetic approaches. Using degenerate primers based on mammalian and yeast clathrin HC sequences, a small fragment of the HC gene was amplified from genomic Drosophila DNA by the polymerase chain reaction. Genomic and cDNA clones from phage libraries were isolated and analyzed using this fragment as a probe. The amino acid sequence of the Drosophila clathrin HC deduced from cDNA sequences is 80%, 57% and 49% identical, respectively, with the mammalian, Dictyostelium and yeast HCs. Hybridization in situ to larval polytene chromosomes revealed a single Chc locus at position 13F2 on the X chromosome. A 13-kb genomic Drosophila fragment including the Chc transcription unit was reintroduced into the Drosophila genome via P element-mediated germline transformation. This DNA complemented a group of EMS-induced lethal mutations mapping to the same region of the X chromosome, thus identifying the Chc complementation group. Mutant individuals homozygous or hemizygous for the Chc(1), Chc(2) or Chc(3) alleles developed to a late stage of embryogenesis, but failed to hatch to the first larval stage. A fourth allele, Chc(4), exhibited polyphasic lethality, with a significant number of homozygous and hemizygous offspring surviving to adulthood. Germline clonal analysis of Chc mutant alleles indicated that the three tight lethal alleles were autonomous cell-lethal mutations in the female germline. In contrast, Chc(4) germline clones were viable at a rate comparable to wild type, giving rise to viable adult progeny. However, hemizygous Chc(4) males were invariably sterile. The sterility was efficiently rescued by an autosomal copy of the wild-type Chc gene reintroduced on a P element. These findings suggest a specialized role for clathrin in spermatogenesis.

Bazinet, C.; Katzen, A. L.; Morgan, M.; Mahowald, A. P.; Lemmon, S. K.

1993-01-01

285

A FUSCA gene of Arabidopsis encodes a novel protein essential for plant development.  

PubMed Central

Arabidopsis fusca mutants display striking purple coloration due to anthocyanin accumulation in their cotyledons. We describe six recessive fusca mutants isolated from Agrobacterium-transformed Arabidopsis families. These mutants first become defective during embryogenesis and exhibit limited seedling development. Double mutant constructs revealed that developmental defects were not simply a consequence of anthocyanin accumulation. fusca seedlings showed altered responses to several environmental and endogenous factors. Allelism tests established that three fusca loci are represented by mutants previously described as defective in light-regulated responses. To study the molecular basis of the fusca phenotype, we cloned the FUS6 gene. FUS6 encodes a novel protein that is hydrophilic, alpha-helical, and contains potential protein kinase C phosphorylation sites. The FUSCA proteins appear to act in a network of signal transduction pathways critical for plant development.

Castle, L A; Meinke, D W

1994-01-01

286

Validation of Cold Chain Products - An essential need for Global Pharmaceutical Supply Chain  

Microsoft Academic Search

Cold chain products are the products which requires special temperature controlled storage.Cold chain storage system is used to store vaccines, certain injectable preparations. Maintaining the temperture is necessary not only during the manufacturing but the product has to be stored up to the destination (patient). Cold chains are common in the pharmaceutical and food industries.One common temperature range for a

Varsha M. Jadhav; Sachin B. Gholve; Vilasrao J. Kadam

287

The Dr-nanos gene is essential for germ cell specification in the planarian Dugesia ryukyuensis.  

PubMed

Homologs of nanos are required for the formation and maintenance of germline stem cell (GSC) systems and for gametogenesis in many metazoans. Planarians can change their reproductive mode seasonally, alternating between asexual and sexual reproduction; they develop and maintain their somatic stem cells (SSCs) and GCSs from pluripotent stem cells known as neoblasts. We isolated a nanos homolog, Dr-nanos, from the expressed sequence tags (ESTs) of the sexualized form of Dugesia ryukyuensis. We examined the expression of Dr-nanos in asexual and sexualized planarians by in situ hybridization and analyzed its function using RNA interference (RNAi) together with a planarian sexualization assay. A nanos homolog, Dr-nanos, was identified in the planarian D. ryukyuensis. Dr-nanos expression was observed in the ovarian primordial cells of the asexual worms. This expression increased in proportion to sexualization and was localized in the early germline cells of the ovaries and testes. In X-ray-irradiated worms, the expression of Dr-nanos decreased to a large extent, indicating that Dr-nanos is expressed in some subpopulations of stem cells, especially in GSCs. During the sexualization process, worms in which Dr-nanos was knocked down by RNAi exhibited decreased numbers of oogonia in the ovaries and failed to develop testes, whereas the somatic sexual organs were not affected. We conclude that Dr-nanos is essential for the development of germ cells in the ovaries and testes and may have a function in the early stages of germ cell specification, but not in the development of somatic sexual organs. PMID:22451004

Nakagawa, Haruka; Ishizu, Hirotsugu; Chinone, Ayako; Kobayashi, Kazuya; Matsumoto, Midori

2012-01-01

288

RPG1: an essential gene of Saccharomyces cerevisiae encoding a 110-kDa protein required for passage through the G1 phase  

Microsoft Academic Search

In Saccharomyces cerevisiae cells a number of genes are required for progression through, or else to pass beyond, the G1 phase. We characterized a novel gene, RPG1, which is also involved in this phase. RPG1 is an essential gene encoding a 110-kDa evolutionarily conserved protein. Elutriated or ?-factor-synchronized cells of the rpg1-1 temperature-sensitive mutant were arrested in the first cell

Pavel Kovarik; Ji?í Hašek; Leoš Valášek; Helmut Ruis

1998-01-01

289

Ci-Rga, a gene encoding an MtN3\\/saliva family transmembrane protein, is essential for tissue differentiation during embryogenesis of the ascidian Ciona intestinalis  

Microsoft Academic Search

A novel gene (Ci-Rga) essential for tissue differentiation during embryogenesis of the ascidian Ciona intestinalis is reported here. This gene was identified through functional screening of Ciona genes required for development by translational inhibition experiments with morpholino antisense oligonucleotides. The deduced protein of Ci-Rga contains two copies of a domain with unknown function called the MtN3\\/saliva domain. Phylogenetic analysis showed

Mayuko Hamada; Shuichi Wada; Kenji Kobayashi; Nori Satoh

2005-01-01

290

Biosynthetic pathway of sugar nucleotides essential for welan gum production in Alcaligenes sp. CGMCC2428  

Microsoft Academic Search

Welan gum is a microbial polysaccharide produced by Alcaligenes sp. CGMCC2428 that has d-glucose, d-glucuronic acid, d-glucose, and l-rhamnose as the main structural unit. The biosynthetic pathway of sugar nucleotides essential for producing welan gum in\\u000a this strain was established in the following ways: (1) the detection of the presence of several intermediates and key enzymes;\\u000a (2) the analysis of

Hui Li; Hong Xu; Hao Xu; Sha Li; Ping-Kai Ouyang

2010-01-01

291

A soxA Gene, Encoding a Diheme Cytochrome c, and a sox Locus, Essential for Sulfur Oxidation in a New Sulfur Lithotrophic Bacterium  

PubMed Central

A mobilizable suicide vector, pSUP5011, was used to introduce Tn5-mob in a new facultative sulfur lithotrophic bacterium, KCT001, to generate mutants defective in sulfur oxidation (Sox?). The Sox? mutants were unable to oxidize thiosulfate while grown mixotrophically in the presence of thiosulfate and succinate. The mutants were also impaired in oxidizing other reduced sulfur compounds and elemental sulfur as evident from the study of substrate oxidation by the whole cells. Sulfite oxidase activity was significantly diminished in the cell extracts of all the mutants. A soxA gene was identified from the transposon-adjacent genomic DNA of a Sox? mutant strain. The sequence analysis revealed that the soxA open reading frame (ORF) is preceded by a potential ribosome binding site and promoter region with ?10- and ?35-like sequences. The deduced nucleotide sequence of the soxA gene was predicted to code for a protein of 286 amino acids. It had a signal peptide of 26 N-terminal amino acids. The amino acid sequence showed similarity with a putative gene product of Aquifex aeolicus, soluble cytochrome c551 of Chlorobium limicola, and the available partial SoxA sequence of Paracoccus denitrificans. The soxA-encoded product seems to be a diheme cytochrome c for KCT001 and A. aeolicus, but the amino acid sequence of C. limicola cytochrome c551 revealed a single heme-binding region. Another transposon insertion mutation was mapped within the soxA ORF. Four other independent transposon insertion mutations were mapped in the 4.4-kb soxA contiguous genomic DNA region. The results thus suggest that a sox locus of KCT001, essential for sulfur oxidation, was affected by all these six independent insertion mutations.

Mukhopadhyaya, Pratap N.; Deb, Chirajyoti; Lahiri, Chandrajit; Roy, Pradosh

2000-01-01

292

Mouse gene targeting reveals an essential role of mTOR in hematopoietic stem cell engraftment and hematopoiesis  

PubMed Central

mTOR integrates signals from nutrients and growth factors to control protein synthesis, cell growth, and survival. Although mTOR has been established as a therapeutic target in hematologic malignancies, its physiological role in regulating hematopoiesis remains unclear. Here we show that conditional gene targeting of mTOR causes bone marrow failure and defects in multi-lineage hematopoiesis including myelopoiesis, erythropoiesis, thrombopoiesis, and lymphopoiesis. mTOR deficiency results in loss of quiescence of hematopoietic stem cells, leading to a transient increase but long-term exhaustion and defective engraftment of hematopoietic stem cells in lethally irradiated recipient mice. Furthermore, ablation of mTOR causes increased apoptosis in lineage-committed blood cells but not hematopoietic stem cells, indicating a differentiation stage-specific function. These results demonstrate that mTOR is essential for hematopoietic stem cell engraftment and multi-lineage hematopoiesis.

Guo, Fukun; Zhang, Shuangmin; Grogg, Matthew; Cancelas, Jose A.; Varney, Melinda E.; Starczynowski, Daniel T.; Du, Wei; Yang, Jun-Qi; Liu, Wei; Thomas, George; Kozma, Sara; Pang, Qishen; Zheng, Yi

2013-01-01

293

EXPORTIN1 Genes Are Essential for Development and Function of the Gametophytes in Arabidopsis thaliana  

PubMed Central

Gametes are produced in plants through mitotic divisions in the haploid gametophytes. We investigated the role of EXPORTIN1 (XPO1) genes during the development of both female and male gametophytes of Arabidopsis. Exportins exclude target proteins from the nucleus and are also part of a complex recruited at the kinetochores during mitosis. Here we show that double mutants in Arabidopsis XPO1A and XPO1B are gametophytic defective. In homozygous–heterozygous plants, 50% of the ovules were arrested at different stages according to the parental genotype. Double-mutant female gametophytes of xpo1a-3/+; xpo1b-1/xpo1b-1 plants failed to undergo all the mitotic divisions or failed to complete embryo sac maturation. Double-mutant female gametophytes of xpo1a-3/xpo1a-3; xpo1b-1/+ plants had normal mitotic divisions and fertilization occurred; in most of these embryo sacs the endosperm started to divide but an embryo failed to develop. Distortions in male transmission correlated with the occurrence of smaller pollen grains, poor pollen germination, and shorter pollen tubes. Our results show that mitotic divisions are possible without XPO1 during the haploid phase, but that XPO1 is crucial for the maternal-to-embryonic transition.

Blanvillain, Robert; Boavida, Leonor C.; McCormick, Sheila; Ow, David W.

2008-01-01

294

The Zebrafish moonshine Gene Encodes Transcriptional Intermediary Factor 1?, an Essential Regulator of Hematopoiesis  

PubMed Central

Hematopoiesis is precisely orchestrated by lineage-specific DNA-binding proteins that regulate transcription in concert with coactivators and corepressors. Mutations in the zebrafish moonshine (mon) gene specifically disrupt both embryonic and adult hematopoiesis, resulting in severe red blood cell aplasia. We report that mon encodes the zebrafish ortholog of mammalian transcriptional intermediary factor 1? (TIF1?) (or TRIM33), a member of the TIF1 family of coactivators and corepressors. During development, hematopoietic progenitor cells in mon mutants fail to express normal levels of hematopoietic transcription factors, including gata1, and undergo apoptosis. Three different mon mutant alleles each encode premature stop codons, and enforced expression of wild-type tif1? mRNA rescues embryonic hematopoiesis in homozygous mon mutants. Surprisingly, a high level of zygotic tif1? mRNA expression delineates ventral mesoderm during hematopoietic stem cell and progenitor formation prior to gata1 expression. Transplantation studies reveal that tif1? functions in a cell-autonomous manner during the differentiation of erythroid precursors. Studies in murine erythroid cell lines demonstrate that Tif1? protein is localized within novel nuclear foci, and expression decreases during erythroid cell maturation. Our results establish a major role for this transcriptional intermediary factor in the differentiation of hematopoietic cells in vertebrates.

Ransom, David G; Bahary, Nathan; Niss, Knut; Traver, David; Burns, Caroline; Trede, Nikolaus S; Paffett-Lugassy, Noelle; Saganic, Walter J; Lim, C. Anthoney; Hersey, Candace; Zhou, Yi; Barut, Bruce A; Lin, Shuo; Kingsley, Paul D; Palis, James; Orkin, Stuart H

2004-01-01

295

The zebrafish moonshine gene encodes transcriptional intermediary factor 1gamma, an essential regulator of hematopoiesis.  

PubMed

Hematopoiesis is precisely orchestrated by lineage-specific DNA-binding proteins that regulate transcription in concert with coactivators and corepressors. Mutations in the zebrafish moonshine (mon) gene specifically disrupt both embryonic and adult hematopoiesis, resulting in severe red blood cell aplasia. We report that mon encodes the zebrafish ortholog of mammalian transcriptional intermediary factor 1gamma (TIF1gamma) (or TRIM33), a member of the TIF1 family of coactivators and corepressors. During development, hematopoietic progenitor cells in mon mutants fail to express normal levels of hematopoietic transcription factors, including gata1, and undergo apoptosis. Three different mon mutant alleles each encode premature stop codons, and enforced expression of wild-type tif1gamma mRNA rescues embryonic hematopoiesis in homozygous mon mutants. Surprisingly, a high level of zygotic tif1gamma mRNA expression delineates ventral mesoderm during hematopoietic stem cell and progenitor formation prior to gata1 expression. Transplantation studies reveal that tif1gamma functions in a cell-autonomous manner during the differentiation of erythroid precursors. Studies in murine erythroid cell lines demonstrate that Tif1gamma protein is localized within novel nuclear foci, and expression decreases during erythroid cell maturation. Our results establish a major role for this transcriptional intermediary factor in the differentiation of hematopoietic cells in vertebrates. PMID:15314655

Ransom, David G; Bahary, Nathan; Niss, Knut; Traver, David; Burns, Caroline; Trede, Nikolaus S; Paffett-Lugassy, Noelle; Saganic, Walter J; Lim, C Anthoney; Hersey, Candace; Zhou, Yi; Barut, Bruce A; Lin, Shuo; Kingsley, Paul D; Palis, James; Orkin, Stuart H; Zon, Leonard I

2004-08-17

296

Drosophila liquid facets-Related encodes Golgi epsin and is an essential gene required for cell proliferation, growth, and patterning  

PubMed Central

Epsin and epsin-Related (epsinR) are multi-modular proteins that stimulate clathrin-coated vesicle formation. Epsin promotes endocytosis at the plasma membrane, and epsinR functions at the Golgi and early endosomes for trans-Golgi network/endosome vesicle trafficking. In Drosophila, endocytic epsin is known as Liquid facets, and it is essential specifically for Notch signaling. Here, by generating and analyzing loss-of-function mutants in the liquid facets-Related (lqfR) gene of Drosophila, we investigated the function of Golgi epsin in a multicellular context. We found that LqfR is indeed a Golgi protein, and that like liquid facets, lqfR is essential for Drosophila viability. In addition, primarily by analyzing mutant eye discs, we found that lqfR is required for cell proliferation, insulin-independent cell growth, and cell patterning, consistent with a role in one or several signaling pathways. Epsins in all organisms share an ENTH (epsin N-terminal homology) domain, which binds phosphoinositides enriched at the plasma membrane or the Golgi membrane. The epsinR ENTH domain is also the recognition element for particular cargos. By generating wild-type and mutant lqfR transgenes, we found that all apparent LqfR functions are independent of its ENTH domain. These results suggest that LqfR transports specific cargo critical to one or more signaling pathways, and lays the foundation for identifying those proteins.

Lee, Ji-Hoon; Overstreet, Erin; Fitch, Erin; Fleenor, Stephen; Fischer, Janice

2009-01-01

297

MatchMiner: a tool for batch navigation among gene and gene product identifiers  

PubMed Central

MatchMiner is a freely available program package for batch navigation among gene and gene product identifier types commonly encountered in microarray studies and other forms of 'omic' research. The user inputs a list of gene identifiers and then uses the Merge function to find the overlap with a second list of identifiers of either the same or a different type or uses the LookUp function to find corresponding identifiers.

Bussey, Kimberly J; Kane, David; Sunshine, Margot; Narasimhan, Sudar; Nishizuka, Satoshi; Reinhold, William C; Zeeberg, Barry; Ajay; Weinstein, John N

2003-01-01

298

The Overlapping angB and angG Genes Are Encoded within the trans-Acting Factor Region of the Virulence Plasmid in Vibrio anguillarum: Essential Role in Siderophore Biosynthesis  

PubMed Central

Products encoded in the trans-acting factor (TAF) region are necessary for the biosynthesis of anguibactin and for maximal expression of iron transport and biosynthesis genes in the plasmid-encoded iron-scavenging system of Vibrio anguillarum. Here we identify angB, a locus located in the TAF region, which encodes products essential for anguibactin biosynthesis. We demonstrate that a 287-amino-acid polypeptide, encoded by angB and designated AngB, has an isochorismate lyase activity necessary for the synthesis of 2,3-dihydroxybenzoic acid, an anguibactin biosynthesis intermediate. Complementation of various angB mutations provided evidence that an additional, overlapping gene exists at this locus. This second gene, designated angG, also has an essential biosynthetic function. The angG gene directs the expression of three polypeptides when overexpressed in Escherichia coli, all of which are translated in the same frame as AngB. The results of site-directed mutagenesis and in vivo phosphorylation experiments suggest that the carboxy-terminal end of AngB and the AngG polypeptide(s) function as aryl carrier proteins involved in the assembly of the anguibactin molecule. Our results also show that the regulatory functions of the TAF are encoded in a region, TAFr, which is distinct from and independent of the angB and angG genes.

Welch, Timothy J.; Chai, Sunghee; Crosa, Jorge H.

2000-01-01

299

The Arabidopsis AtRaptor genes are essential for post-embryonic plant growth  

PubMed Central

Background Flowering plant development is wholly reliant on growth from meristems, which contain totipotent cells that give rise to all post-embryonic organs in the plant. Plants are uniquely able to alter their development throughout their lifespan through the generation of new organs in response to external signals. To identify genes that regulate meristem-based growth, we considered homologues of Raptor proteins, which regulate cell growth in response to nutrients in yeast and metazoans as part of a signaling complex with the target of rapamycin (TOR) kinase. Results We identified AtRaptor1A and AtRaptor1B, two loci predicted to encode Raptor proteins in Arabidopsis. Disruption of AtRaptor1B yields plants with a wide range of developmental defects: roots are thick and grow slowly, leaf initiation and bolting are delayed and the shoot inflorescence shows reduced apical dominance. AtRaptor1A AtRaptor1B double mutants show normal embryonic development but are unable to maintain post-embryonic meristem-driven growth. AtRaptor transcripts accumulate in dividing and expanding cells and tissues. Conclusion The data implicate the TOR signaling pathway, a major regulator of cell growth in yeast and metazoans, in the maintenance of growth from the shoot apical meristem in plants. These results provide insights into the ways in which TOR/Raptor signaling has been adapted to regulate plant growth and development, and indicate that in plants, as in other eukaryotes, there is some Raptor-independent TOR activity.

Anderson, Garrett H; Veit, Bruce; Hanson, Maureen R

2005-01-01

300

Natural product biosynthetic gene diversity in geographically distinct soil microbiomes.  

PubMed

The number of bacterial species estimated to exist on Earth has increased dramatically in recent years. This newly recognized species diversity has raised the possibility that bacterial natural product biosynthetic diversity has also been significantly underestimated by previous culture-based studies. Here, we compare 454-pyrosequenced nonribosomal peptide adenylation domain, type I polyketide ketosynthase domain, and type II polyketide ketosynthase alpha gene fragments amplified from cosmid libraries constructed using DNA isolated from three different arid soils. While 16S rRNA gene sequence analysis indicates these cloned metagenomes contain DNA from similar distributions of major bacterial phyla, we found that they contain almost completely distinct collections of secondary metabolite biosynthetic gene sequences. When grouped at 85% identity, only 1.5% of the adenylation domain, 1.2% of the ketosynthase, and 9.3% of the ketosynthase alpha sequence clusters contained sequences from all three metagenomes. Although there is unlikely to be a simple correlation between biosynthetic gene sequence diversity and the diversity of metabolites encoded by the gene clusters in which these genes reside, our analysis further suggests that sequences in one soil metagenome are so distantly related to sequences in another metagenome that they are, in many cases, likely to arise from functionally distinct gene clusters. The marked differences observed among collections of biosynthetic genes found in even ecologically similar environments suggest that prokaryotic natural product biosynthesis diversity is, like bacterial species diversity, potentially much larger than appreciated from culture-based studies. PMID:22427492

Reddy, Boojala Vijay B; Kallifidas, Dimitris; Kim, Jeffrey H; Charlop-Powers, Zachary; Feng, Zhiyang; Brady, Sean F

2012-03-16

301

Effects of Cymbopogon citratus L. essential oil on the growth, morphogenesis and aflatoxin production of Aspergillus flavus ML2-strain.  

PubMed

The mycelial growth of Aspergillus flavus Link was completely inhibited using 1.5 (microl/ml or 2.0 (microl/ml of Cymbopogon citratus essential oil applied by fumigation or contact method in Czapek's liquid medium, respectively. This oil was found also to be fungicidal at the same concentrations. The sublethal doses 1.0 and 1.5 (microl/ml inhibited about 65% of fungal growth after five days of incubation and delayed conidiation as compared with the control. Microscopic observations using Light Microscope (LM), Scanning Electron Microscope (SEM) and Transmission Electron Microscope (TEM) were carried out to determine the ultra structural modifications of A. flavus hyphae after treatment with C. citratus essential oil. The hyphal diameter decreased and hyphal wall appeared as precipitates and disappeared in some regions. This oil also caused plasma membrane disruption and mitochondrial structure disorganization. Moreover, Ca(+2), K(+) and Mg(+2) leakages increased from the fumigated mycelium and its total lipid content decreased, while the saturated and unsaturated fatty acids increased. One of the most important results obtained during this study was the ability of C. citratus essential oil at its sublethal dose to completely inhibit aflatoxin B(1) production from A. flavus. These findings increase the possibility of exploiting C. citratus essential oil as an effective inhibitor of biodegradation and storage contaminating fungi and also in fruit juice preservation. PMID:17304618

Helal, G A; Sarhan, M M; Abu Shahla, A N K; Abou El-Khair, E K

2007-02-01

302

Functional Characterization of MAT1-1-Specific Mating-Type Genes in the Homothallic Ascomycete Sordaria macrospora Provides New Insights into Essential and Nonessential Sexual Regulators?†  

PubMed Central

Mating-type genes in fungi encode regulators of mating and sexual development. Heterothallic ascomycete species require different sets of mating-type genes to control nonself-recognition and mating of compatible partners of different mating types. Homothallic (self-fertile) species also carry mating-type genes in their genome that are essential for sexual development. To analyze the molecular basis of homothallism and the role of mating-type genes during fruiting-body development, we deleted each of the three genes, SmtA-1 (MAT1-1-1), SmtA-2 (MAT1-1-2), and SmtA-3 (MAT1-1-3), contained in the MAT1-1 part of the mating-type locus of the homothallic ascomycete species Sordaria macrospora. Phenotypic analysis of deletion mutants revealed that the PPF domain protein-encoding gene SmtA-2 is essential for sexual reproduction, whereas the ? domain protein-encoding genes SmtA-1 and SmtA-3 play no role in fruiting-body development. By means of cross-species microarray analysis using Neurospora crassa oligonucleotide microarrays hybridized with S. macrospora targets and quantitative real-time PCR, we identified genes expressed under the control of SmtA-1 and SmtA-2. Both genes are involved in the regulation of gene expression, including that of pheromone genes.

Klix, V.; Nowrousian, M.; Ringelberg, C.; Loros, J. J.; Dunlap, J. C.; Poggeler, S.

2010-01-01

303

A review of plant-derived essential oils in ruminant nutrition and production  

Microsoft Academic Search

Public concern over use of antibiotics in livestock production has increased in recent years because of their possible contribution to emergence of antibiotic resistant bacteria, and their transmission from livestock to humans. Accordingly, ruminant microbiologists and nutritionists have been exploring alternative methods of favorably altering ruminal metabolism to improve feed efficiency and animal productivity. Plant extracts contain secondary metabolites, such

C. Benchaar; S. Calsamiglia; A. V. Chaves; G. R. Fraser; D. Colombatto; T. A. McAllister; K. A. Beauchemin

2008-01-01

304

Nitrilase from Rhodococcus rhodochrous J1. Sequencing and overexpression of the gene and identification of an essential cysteine residue.  

PubMed

The amino acid sequences of the NH2 terminus and internal peptide fragments of a Rhodococcus rhodochrous J1 nitrilase were determined to prepare synthetic oligonucleotides as primers for the polymerase chain reaction. A 750-base DNA fragment thus amplified was used as the probe to clone a 5.4-kilobase PstI fragment coding for the whole nitrilase. The nitrilase gene modified in the sequence upstream from the presumed ATG start codon was expressed to approximately 50% of the total soluble protein in Escherichia coli. The predicted amino acid sequence of the nitrilase gene showed similarity to that of the bromoxynil nitrilase from Klebsiella ozaenae. The 5,5'-dithiobis(2-nitrobenzoic acid) modification of the nitrilase from R. rhodochrous J1 resulted in inactivation with the loss of one sulfhydryl group/enzyme subunit. Of 4 cysteine residues in the Rhodococcus nitrilase, only Cys-165 is conserved in the Klebsiella nitrilase. Mutant enzymes containing Ala or Ser instead of Cys-165 did not exhibit nitrilase activity. These findings suggest that Cys-165 plays an essential role in the function of the active site. PMID:1400390

Kobayashi, M; Komeda, H; Yanaka, N; Nagasawa, T; Yamada, H

1992-10-15

305

Intrachromosomal tandem duplication and repeat expansion during attempts to inactivate the subtelomeric essential gene GSH1 in Leishmania  

PubMed Central

Gamma-glutamylcysteine synthetase encoded by GSH1 is the rate-limiting enzyme in the biosynthesis of glutathione and trypanothione in Leishmania. Attempts to generate GSH1 null mutants by gene disruption failed in Leishmania infantum. Removal of even a single allele invariably led to the generation of an extra copy of GSH1, maintaining two intact wild-type alleles. In the second and even third round of inactivation, the markers integrated at the homologous locus but always preserved two intact copies of GSH1. We probed into the mechanism of GSH1 duplication. GSH1 is subtelomeric on chromosome 18 and Southern blot analysis indicated that a 10-kb fragment flanked by 466-bp direct repeated sequences was duplicated in tandem on the same chromosomal allele each time GSH1 was targeted. Polymerase chain reaction analysis and sequencing confirmed the generation of novel junctions created at the level of the 466-bp repeats consequent to locus duplication. In loss of heterozygosity attempts, the same repeated sequences were utilized for generating extrachromosomal circular amplicons. Our results are consistent with break-induced replication as a mechanism for the generation of this regional polyploidy to compensate for the inactivation of an essential gene. This chromosomal repeat expansion through repeated sequences could be implicated in locus duplication in Leishmania.

Mukherjee, Angana; Langston, Lance D.; Ouellette, Marc

2011-01-01

306

The zebrafish udu gene encodes a novel nuclear factor and is essential for primitive erythroid cell development.  

PubMed

Hematopoiesis is a complex process which gives rise to all blood lineages in the course of an organism's lifespan. However, the underlying molecular mechanism governing this process is not fully understood. Here we report the isolation and detailed study of a newly identified zebrafish ugly duckling (Udu) mutant allele, Udu(sq1). We show that loss-of-function mutation in the udu gene disrupts primitive erythroid cell proliferation and differentiation in a cell-autonomous manner, resulting in red blood cell (RBC) hypoplasia. Positional cloning reveals that the Udu gene encodes a novel factor that contains 2 paired amphipathic alpha-helix-like (PAH-L) repeats and a putative SANT-L (SW13, ADA2, N-Cor, and TFIIIB-like) domain. We further show that the Udu protein is predominantly localized in the nucleus and deletion of the putative SANT-L domain abolishes its function. Our study indicates that the Udu protein is very likely to function as a transcription modulator essential for the proliferation and differentiation of erythroid lineage. PMID:17369489

Liu, Yanmei; Du, Linsen; Osato, Motomi; Teo, Eng Hui; Qian, Feng; Jin, Hao; Zhen, Fenghua; Xu, Jin; Guo, Lin; Huang, Honghui; Chen, Jun; Geisler, Robert; Jiang, Yun-Jin; Peng, Jinrong; Wen, Zilong

2007-03-16

307

Genes for a microaerobically induced oxidase complex in Bradyrhizobium japonicum are essential for a nitrogen-fixing endosymbiosis.  

PubMed Central

We report the discovery of a Bradyrhizobium japonicum gene cluster (fixNOQP) in which mutations resulted in defective soybean root-nodule bacteroid development and symbiotic nitrogen fixation. The predicted, DNA-derived protein sequences suggested that FixN is a heme b and copper-binding oxidase subunit, FixO a monoheme cytochrome c, FixQ a polypeptide of 54 amino acids, and FixP a diheme cytochrome c and that they are all membrane-bound. The isolation and analysis of membrane proteins from B. japonicum wild-type and mutant cells revealed two c-type cytochromes of 28 and 32 kDa as the likely products of the fixO and fixP genes and showed that both were synthesized only under oxygen-limited growth conditions. Furthermore, fixN insertion and fixNO deletion mutants grown microaerobically or anaerobically (with nitrate) exhibited a strong decrease in whole-cell oxidase activity as compared with the wild type. The data suggest that the fixNOQP gene products are induced at low oxygen concentrations and constitute a member of the bacterial heme/copper cytochrome oxidase superfamily. The described features are compatible with the postulate that this oxidase complex is specifically required to support bacterial respiration in endosymbiosis. Images Fig. 4 Fig. 5

Preisig, O; Anthamatten, D; Hennecke, H

1993-01-01

308

Use of Genomics To Identify Bacterial Undecaprenyl Pyrophosphate Synthetase: Cloning, Expression, and Characterization of the Essential uppS Gene  

PubMed Central

The prenyltransferase undecaprenyl pyrophosphate synthetase (di-trans,poly-cis-decaprenylcistransferase; EC 2.5.1.31) was purified from the soluble fraction of Escherichia coli by TSK-DEAE, ceramic hydroxyapatite, TSK-ether, Superdex 200, and heparin-Actigel chromatography. The protein was labeled with the photolabile analogue of the farnesyl pyrophosphate analogue (E,E)-[1-3H]-(2-diazo-3-trifluoropropionyloxy)geranyl diphos-phate and was detected on a sodium dodecyl sulfate-polyacrylamide gel as a protein with an apparent molecular mass of 29 kDa. This protein band was cut out from the gel, trypsin digested, and subjected to matrix-assisted laser desorption ionization mass spectrometric analysis. Comparison of the experimental data with computer-simulated trypsin digest data for all E. coli proteins yielded a single match with a protein of unassigned function (SWISS-PROT Q47675; YAES_ECOLI). Sequences with strong similarity indicative of homology to this protein were identified in 25 bacterial species, in Saccharomyces cerevisiae, and in Caenorhabditis elegans. The homologous genes (uppS) were cloned from E. coli, Haemophilus influenzae, and Streptococcus pneumoniae, expressed in E. coli as amino-terminal His-tagged fusion proteins, and purified over a Ni2+ affinity column. An untagged version of the E. coli uppS gene was also cloned and expressed, and the protein purified in two chromatographic steps. We were able to detect Upp synthetase activity for all purified enzymes. Further, biochemical characterization revealed no differences between the recombinant untagged E. coli Upp synthetase and the three His-tagged fusion proteins. All enzymes were absolutely Triton X-100 and MgCl2 dependent. With the use of a regulatable gene disruption system, we demonstrated that uppS is essential for growth in S. pneumoniae R6.

Apfel, Christian M.; Takacs, Bela; Fountoulakis, Michael; Stieger, Martin; Keck, Wolfgang

1999-01-01

309

liver-enriched gene 1a and 1b Encode Novel Secretory Proteins Essential for Normal Liver Development in Zebrafish  

PubMed Central

liver-enriched gene 1 (leg1) is a liver-enriched gene in zebrafish and encodes a novel protein. Our preliminary data suggested that Leg1 is probably involved in early liver development. However, no detailed characterization of Leg1 has been reported thus far. We undertook both bioinformatic and experimental approaches to study leg1 gene structure and its role in early liver development. We found that Leg1 identifies a new conserved protein superfamily featured by the presence of domain of unknown function 781 (DUF781). There are two copies of leg1 in zebrafish, namely leg1a and leg1b. Both leg1a and leg1b are expressed in the larvae and adult liver with leg1a being the predominant form. Knockdown of Leg1a or Leg1b by their respective morpholinos specifically targeting their 5?-UTR each resulted in a small liver phenotype, demonstrating that both Leg1a and Leg1b are important for early liver development. Meanwhile, we found that injection of leg1-ATGMO, a morpholino which can simultaneously block the translation of Leg1a and Leg1b, caused not only a small liver phenotype but hypoplastic exocrine pancreas and intestinal tube as well. Further examination of leg1-ATGMO morphants with early endoderm markers and early hepatic markers revealed that although depletion of total Leg1 does not alter the hepatic and pancreatic fate of the endoderm cells, it leads to cell cycle arrest that results in growth retardation of liver, exocrine pancreas and intestine. Finally, we proved that Leg1 is a secretory protein. This intrigued us to propose that Leg1 might act as a novel secreted regulator that is essential for liver and other digestive organ development in zebrafish.

Chang, Changqing; Hu, Minjie; Zhu, Zhihui; Lo, Li Jan; Chen, Jun; Peng, Jinrong

2011-01-01

310

Use of genomics to identify bacterial undecaprenyl pyrophosphate synthetase: cloning, expression, and characterization of the essential uppS gene.  

PubMed

The prenyltransferase undecaprenyl pyrophosphate synthetase (di-trans,poly-cis-decaprenylcistransferase; EC 2.5.1.31) was purified from the soluble fraction of Escherichia coli by TSK-DEAE, ceramic hydroxyapatite, TSK-ether, Superdex 200, and heparin-Actigel chromatography. The protein was labeled with the photolabile analogue of the farnesyl pyrophosphate analogue (E, E)-[1-3H]-(2-diazo-3-trifluoropropionyloxy)geranyl diphosphate and was detected on a sodium dodecyl sulfate-polyacrylamide gel as a protein with an apparent molecular mass of 29 kDa. This protein band was cut out from the gel, trypsin digested, and subjected to matrix-assisted laser desorption ionization mass spectrometric analysis. Comparison of the experimental data with computer-simulated trypsin digest data for all E. coli proteins yielded a single match with a protein of unassigned function (SWISS-PROT Q47675; YAES_ECOLI). Sequences with strong similarity indicative of homology to this protein were identified in 25 bacterial species, in Saccharomyces cerevisiae, and in Caenorhabditis elegans. The homologous genes (uppS) were cloned from E. coli, Haemophilus influenzae, and Streptococcus pneumoniae, expressed in E. coli as amino-terminal His-tagged fusion proteins, and purified over a Ni2+ affinity column. An untagged version of the E. coli uppS gene was also cloned and expressed, and the protein purified in two chromatographic steps. We were able to detect Upp synthetase activity for all purified enzymes. Further, biochemical characterization revealed no differences between the recombinant untagged E. coli Upp synthetase and the three His-tagged fusion proteins. All enzymes were absolutely Triton X-100 and MgCl2 dependent. With the use of a regulatable gene disruption system, we demonstrated that uppS is essential for growth in S. pneumoniae R6. PMID:9882662

Apfel, C M; Takács, B; Fountoulakis, M; Stieger, M; Keck, W

1999-01-01

311

Characterization of the cloned fip gene and its product.  

PubMed Central

A DNA fragment encoding the fip (filamentous phage production) gene from Escherichia coli, when cloned in a filamentous phage vector, restored to the phage ability to assemble progeny in fip mutant hosts. The fip gene was located just upstream of and transcribed in the same direction as the rho gene. Minicells containing fip+ phage or plasmids synthesized a 12,500-dalton protein that was missing or truncated when the Fip+ phenotype was inactivated by insertion of Tn5. The fip protein was cytoplasmic and was partially purified. Images

Russel, M; Model, P

1984-01-01

312

An association of BMI with A (-6) G, M235T and T174M polymorphisms in angiotensinogen gene in essential hypertension  

Microsoft Academic Search

The aim of the study was to assess the existence of possible associations among frequent polymorphisms in angiotensinogen genes and some of the risk factors for essential hypertension, especially body mass index (BMI) and smoking. A total of 192 control subjects (aged 45.87 ± 3.0 years) and 206 patients with the essential hypertension (aged 48.71 ± 8.42 years) were compared

A Vašk?; M Sou?ek; S Tschöplová; A Stejskalová

2002-01-01

313

Association between 894G>T Polymorphism in the Endothelial Nitric Oxide Synthase Gene and Circadian Variation of Blood Pressure in Patients with Essential Hypertension  

Microsoft Academic Search

Background: Human essential hypertension has a genetic basis, and hypertension is associated with altered endothelial nitric oxide (NO) release. We hypothesized that functional alterations of the endothelium-derived NO pathway in the presence of the 894G>T polymorphism of the endothelial NO synthase (eNOS) gene may be related to circadian variation of blood pressure in patients with essential hypertension. Methods: A total

An-Ning Feng; Wei-Hsian Yin; Mason Shing Young; Ming-Wei Lin

2005-01-01

314

Gene Discovery and Product Development for Grain Quality Traits  

NSDL National Science Digital Library

The composition of oils, proteins, and carbohydrates in seeds of corn, soybean, and other crops has been modified to produce grains with enhanced value. Both plant breeding and molecular technologies have been used to produce plants carrying the desired traits. Genomics-based strategies for gene discovery, coupled with high-throughput transformation processes and miniaturized, automated analytical and functionality assays, have accelerated the identification of product candidates. Molecular markerâÂÂbased breeding strategies have been used to accelerate the process of moving trait genes into high-yielding germplasm for commercialization. These products are being tested for applications in food, feed, and industrial markets.

Barbara Mazur (DuPont Agricultural Products Experimental Station;); Enno Krebers (DuPont Agricultural Products Experimental Station;); Scott Tingey (DuPont Agricultural Products Experimental Station;)

1999-07-16

315

Association between the M268T polymorphism in the angiotensinogen gene and essential hypertension in a South Indian population.  

PubMed

Essential hypertension is a complex multifactorial disease caused by interactions between genetic and environmental factors. It is an independent determinant of cardiovascular risk. The main aim of this study was to investigate the possible influence of angiotensinogen M268T polymorphisms on hypertension in two endogamous caste populations of South India. Systolic and diastolic blood pressure, anthropometric variables, and lipid profiles were assessed. Direct sequencing of PCR products was adopted for genotyping. This polymorphism was found to be in Hardy-Weinberg equilibrium in the patients and controls of both populations. Binary odds ratios showed significant association between the M268T polymorphism and hypertension in both populations. Multivariate analysis revealed significant differences in body mass index, chest girth, calf circumference, skinfold measurements, total cholesterol, and triglyceride levels between these genotypes in the Gavara and Vaishya populations. These data further support the hypothesis that hypertension is influenced by the AGT M268T polymorphism. PMID:21312059

Gopi Chand, M; Srinath, J; Rao, R S; Lakkakula, Bhaskar V K S; Kumar, Satish; Rao, V R

2011-02-11

316

Mutational analysis of the Saccharomyces cerevisiae ABD1 gene: cap methyltransferase activity is essential for cell growth.  

PubMed Central

RNA (guanine-7-)-methyltransferase is the enzyme responsible for methylating the 5' cap structure of eukaryotic mRNA. The Saccharomyces cerevisiae enzyme is a 436-amino-acid protein encoded by the essential ABD1 gene. In this study, deletion and point mutations in ABD1 were tested for the ability to support growth of an abd1 null strain. Elimination of 109 amino acids from the N terminus had no effect on cell viability, whereas a more extensive N-terminal deletion of 155 residues was lethal, as was a C-terminal deletion of 55 amino acids. Alanine substitution mutations were introduced at eight conserved residues within a 206-amino-acid region of similarity between ABD1 and the methyltransferase domain of the vaccinia virus capping enzyme. ABD1 alleles H253A (encoding a substitution of alanine for histidine at position 253), T282A, E287A, E361A, and Y362A were viable, whereas G174A, D178A, and Y254A were either lethal or severely defective for growth. Alanine-substituted and amino-truncated ABD1 proteins were expressed in bacteria, purified, and tested for cap methyltransferase activity in vitro. Mutations that were viable in yeast cells had either no effect or only a moderate effect on the specific methyltransferase activity of the mutated ABD1 protein, whereas mutations that were deleterious in vivo yielded proteins that were catalytically defective in vitro. These findings substantiate for the first time the long-held presumption that cap methylation is an essential function in eukaryotic cells.

Mao, X; Schwer, B; Shuman, S

1996-01-01

317

Effects of Zataria multiflora Boiss. essential oil on growth and gene expression of enterotoxins A, C and E in Staphylococcus aureus ATCC 29213.  

PubMed

Staphylococcal food poisoning results from the consumption of food in which enterotoxigenic staphylococci have grown and produced toxins. The present study was conducted with three principal aims: i) to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of Zataria multiflora Boiss. essential oil (EO) against Staphylococcus aureus ATCC 29213, ii) to evaluate the effect of subinhibitory concentrations (subMIC) of EO on the growth of bacteria over 72 h (at 25 and 35 °C), and iii) to investigate the expression of genes involved in the production of staphylococcal enterotoxins SEA, SEC and SEE over 72 h at 35 °C. The MIC and MBC of Z. multiflora Boiss. EO were 0.03 and 0.04%, respectively. Colony counting at 24, 48 and 72 h of 3 day cultures grown in the presence of 75%MIC of the EO showed that the growth rate was reduced 2.16, 2.78 and 2.91 log 10 cfu/ml at 25 °C, and 1.34, 2.35 and 2.57 log 10 cfu/ml at 35 °C, respectively, compared to control cultures. SubMIC levels of EO also significantly decreased the expression of staphylococcal enterotoxin (SE)-related genes and therefore the production of SEs in a dose dependent manner. For example, when cultured with 75% MIC, the transcriptional levels of sea, sec, see and agrA were decreased 11.7, 9.3, 10.45 and 10.3 fold after 18 h and 13.9, 11.21, 12.44 and 12.52 fold after 72 h in comparison to control, respectively. PMID:23558199

Azizkhani, Maryam; Misaghi, Ali; Basti, Afshin Akhondzadeh; Gandomi, Hassan; Hosseini, Hedayat

2013-03-06

318

A seven-gene operon essential for formate-dependent nitrite reduction to ammonia by enteric bacteria.  

PubMed

The DNA sequence of the regulatory region and the structural gene, nrfA, for cytochrome c552 of Escherichia coli K-12 have been reported. We have now established that nrfA is the first gene in a seven-gene operon, designated the nrf operon, at least five of which are essential for formate-dependent nitrite reduction to ammonia. This operon terminates just upstream of the previously sequenced gltP gene encoding a sodium-independent, glutamate and aspartate transporter. Expression of lac fused to nrfA, nrfE or nrfG is regulated by oxygen repression, FNR-dependent anaerobic induction, nitrite induction and nitrate repression during anaerobic growth, exactly as previously reported for the nrfA promoter. In contrast, expression of the gltP-lac fusion was FNR-independent. The open reading frame immediately downstream of nrfA encodes NrfB, a hydrophilic, penta-haem cytochrome c with an M(r) of 20,714. The structure of the N-terminal region is typical of a signal peptide for a periplasmic protein: cleavage at the putative signal peptide cleavage site, Ala-26, would result in a periplasmic cytochrome with a molecular mass of 18 kDa. The NrfC polypeptide, M(r) 24,567, contains 16 cysteine residues arranged in four clusters typical of the CooF super-family of non-haem iron-sulphur proteins. The NrfD sequence predicts a 318-residue hydrophobic protein with a distribution of acidic and basic amino acids which suggests that NrfD is an integral transmembrane protein with loops in both the periplasm and the cytoplasm. Proteins most similar to NrfD include the PsrC subunit of polysulphide reductase from Wolinella, but, as seven of the 10 most similar proteins are NADH-ubiquinone oxidoreductases, we propose that NrfD participates in the transfer of electrons from the quinone pool into the terminal components of the Nrf pathway. NrfE, M(r) 60,851, is predicted to be another hydrophobic, integral membrane protein homologous to the CdI1 protein of Rhodobacter capsulatus, which has been implicated in the assembly of periplasmic c-type cytochromes. The sequence of the 127 residue NrfF polypeptide, M(r) 14,522, is strikingly similar to the CcI2 protein of R. capsulatus, especially in the putative haem-binding motif, RCPQCQNQN.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:8057835

Hussain, H; Grove, J; Griffiths, L; Busby, S; Cole, J

1994-04-01

319

Growth, morphogenesis and essential oil production in Mentha spicata L. plantlets in vitro  

Technology Transfer Automated Retrieval System (TEKTRAN)

The influence of various physical environments were studied on the growth (fresh weight), morphogenesis (leaf, root and shoot numbers) and secondary metabolism [i.e., production of the monoterpene (-)-carvone] of Mentha spicata L. (spearmint) shoots cultured on Murashige and Skoog medium. Carvone a...

320

The haplotype of the growth-differentiation factor 15 gene is associated with left ventricular hypertrophy in human essential hypertension.  

PubMed

GDF15 (growth-differentiation factor 15) is a novel antihypertrophic factor which is induced in the heart in response to pressure overload and plays an important regulatory role in the process of hypertrophy. In the present study, we have investigated the relationship between GDF15 gene variants and left ventricular hypertrophy in human essential hypertension. A community-based hypertensive population sample of 1527 individuals (506 men and 1021 women) was genotyped for three GDF15 genetic variants, including one tag variant -3148C>G (rs4808793) and two exonic variants +157A>T (rs1059369) and +2438C>G (rs1058587). The effects of those variants on gene expression were studied by use of luciferase reporter assays and the determination of plasma GDF15 levels. Only the tag variant -3148G was significantly associated with a lower risk of left ventricular hypertrophy [odds ratio=0.75 (95% confidence interval, 0.63-0.89); P=0.0009]. Multiple regression analyses confirmed that -3148G predicted the decrease in left ventricular end-diastolic diameter (beta=-0.10, P=0.0001), end-systolic diameter (beta=-0.09, P=0.0007), mass (beta=-0.11, P<0.0001) and indexed mass (beta=-0.12, P<0.0001). These effects were independent of conventional factors, including gender, age, body surface area, blood pressure, diabetes, cigarette smoking and alcohol consumption. The transcription activity of the -3148G-containing construct was increased 1.45-fold (P=0.015) at baseline and 1.73-fold (P=0.008) after stimulation with phenylephrine when compared with the -3148C construct. The -3148G allele was also associated with a significant increase in the plasma GDF15 level in hypertensive subjects (P=0.04). In conclusion, the results show that a promoter haplotype containing the -3148G variant increases GDF15 transcription activity and is associated with favourable left ventricular remodelling in human essential hypertension. PMID:19505289

Wang, Xiaojian; Yang, Xu; Sun, Kai; Chen, Jingzhou; Song, Xiaodong; Wang, Hu; Liu, Zhe; Wang, Changxin; Zhang, Channa; Hui, Rutai

2009-10-19

321

Adrenotensin: An adrenomedullin gene product contracts pulmonary blood vessels  

Microsoft Academic Search

The purpose of the present study was to determine the effects of adrenotensin, a newly described product of the ADM gene, on cat pulmonary arterial (PA) rings. Under resting conditions, adrenotensin increased tension of PA rings in a concentration-dependent manner. Although addition of diphenhydramine, ONO-3708, phentclamine, methysergide, atropine, and meclofenamate did not alter the contractile response to adrenotensin, removal of

Bulent Gumusel; Jaw-Kang Chang; Qingzhong Hao; Albert Hyman; Howard Lippton

1996-01-01

322

Protein gene product 9.5 (PGP 9.5)  

Microsoft Academic Search

The guinea pig uterus is supplied by different populations of nerves which can be demonstrated by specific immunocytochemical and histochemical techniques. So far, there has been no single marker displaying entire peripheral innervation patterns. Recently, protein gene product (PGP) 9.5, a cytoplasmic protein in neurons and neuroendocrine cells, was found to visualize both different populations and subtypes of nerves. This

L.-M. Lundberg; P. Alm; J. Wharton; J. M. Polak

1988-01-01

323

Identification of Escherichia coli region III flagellar gene products and description of two new flagellar genes.  

PubMed Central

Region III flagellar genes in Escherichia coli are involved with the assembly and rotation of the flagella, as well as taxis. We subcloned the flaB operon from a lambda fla transducing phage onto plasmid pMK2004. Two additional genes were found at the flaB locus, and we subdivided the flaB gene into flaB1, flaBII, and flaBIII. The cheY suppressor mutations which have previously been mapped to flaB were further localized to flaB11 (Parkinson et al., J. Bacteriol. 155:265-274, 1983). Until now, gene product identification has not been possible for these genes because of their low levels of gene expression. Overexpression of the flagellar genes was accomplished by placing the flaB operon under the control of the lacUV5 or tac promoters. Plasmid-encoded proteins were examined in a minicell expression system. By correlating various deletions and insertions in the flaB operon with the ability to complement specific flagellar mutants and code for polypeptides, we made the following gene product assignments: flaB 1, 60 kilodaltons; flaB 11, 38 kilodaltons; flaB111, 28 kilodaltons; flaC, 56 kilodaltons; fla0, 16 kilodaltons; and flaE, 54 kilodaltons. Images

Bartlett, D H; Matsumura, P

1984-01-01

324

Heterocyclic Compounds Against the Enzyme Tyrosinase Essential for Melanin Production: Biochemical Features of Inhibition  

Microsoft Academic Search

Tyrosinase is a copper-containing bifunctional metalloenzyme, widely distributed around the phylogeny.\\u000a This enzyme is involved in the production of melanin and some other pigments in humans, animals, etc. Abnormal\\u000a accumulation of melanin, which is due to the overexpression of the enzyme, is called hyperpigmentation and\\u000a underexpression is called vitiligo, which is a major skin problem around the world. The inhibitors\\u000a of

Mahmud Khan; Hassan Khan

325

AthPEX10, a nuclear gene essential for peroxisome and storage organelle formation during Arabidopsis embryogenesis.  

PubMed

In yeasts and mammals, PEX10 encodes an integral membrane protein with a C3HC4 RING finger motif in its C-terminal domain and is required for peroxisome biogenesis and matrix protein import. In humans, its dysfunction in peroxisome biogenesis leads to severe Zellweger Syndrome and infantile Refsum disease. Here we show that dysfunction of a homologous gene in Arabidopsis leads to lethality at the heart stage of embryogenesis, impairing the biogenesis of peroxisomes, lipid bodies, and protein bodies. In a T-DNA insertion mutant disrupting the fourth exon of the AthPEX10 gene, ultrastructural analyses fail to detect peroxisomes characteristic for wild-type embryogenesis. Storage triacyl glycerides are not assembled into lipid bodies (oil bodies; oleosomes) surrounded by the phospholipid-protein monolayer membrane. Instead, the dysfunctional monolayer membranes, which derive from the bilayer membrane of the endoplasmic reticulum, accumulate in the cytosol. Concomitantly the transfer of the storage proteins from their site of synthesis at the endoplasmic reticulum to the vacuoles is disturbed. The mutant can be rescued by transformation with wild-type AthPEX10 cDNA. Transformants of wild-type Hansenula polymorpha cells with the AthPEX10 cDNA did produce the encoded protein without targeting it to peroxisomes. Additionally, the cDNA could not complement a Hansenula pex10 mutant unable to form peroxisomes. The ultrastructural knockout phenotype of AthPEX10p suggests that this protein in Arabidopsis is essential for peroxisome, oleosome, and protein transport vesicle formation. PMID:12883010

Schumann, Uwe; Wanner, Gerhard; Veenhuis, Marten; Schmid, Markus; Gietl, Christine

2003-07-25

326

Association of Insertion\\/Deletion Polymorphism of Alpha-Adrenoceptor Gene in Essential Hypertension with or without Type 2 Diabetes Mellitus in Malaysian Subjects  

Microsoft Academic Search

An insertion\\/deletion (I\\/D) polymorphism of Alpha2B-Adrenoceptor (ADRA2B) gene located on chromosome 2 has been studied extensively in related to cardiovascular diseases. The main aim of the present study was to examine the potential association of D allele frequency of I\\/D polymorphism of ADRA2B gene in Malaysian essential hypertensive subjects with or without type 2 diabetes mellitus (T2DM). This study includes

R. Vasudevan; Patimah Ismail; Johnson Stanslas; Norashikin Shamsudin

327

Carotenoid-based phenotypic screen of the yeast deletion collection reveals new genes with roles in isoprenoid production.  

PubMed

Beside their essential cellular functions, isoprenoids have value as pharmaceuticals, nutriceuticals, pesticides, and fuel alternatives. Engineering microorganisms for production of isoprenoids is relatively easy, sustainable, and cost effective in comparison to chemical synthesis or extraction from natural producers. We introduced genes encoding carotenoid biosynthetic enzymes into the haploid yeast deletion collection to identify gene deletions that improved isoprenoid production. Deletions that showed significant improvement in carotenoid production were further screened for production of bisabolene, an isoprenoid alternative to petroleum-derived diesel. Combining those deletions with other mevalonate pathway modifications increased production of bisabolene from 40mg/L to 800mg/L in shake-flask cultures. In a fermentation process, this engineered strain produced 5.2g/L of bisabolene. PMID:22918085

Özayd?n, Bilge; Burd, Helcio; Lee, Taek Soon; Keasling, Jay D

2012-08-17

328

CCN2 (Connective Tissue Growth Factor) is essential for extracellular matrix production and integrin signaling in chondrocytes  

PubMed Central

The matricellular protein CCN2 (Connective Tissue Growth Factor; CTGF) is an essential mediator of ECM composition, as revealed through analysis of Ccn2 deficient mice. These die at birth due to complications arising from impaired endochondral ossification. However, the mechanism(s) by which CCN2 mediates its effects in cartilage are unclear. We investigated these mechanisms using Ccn2?/? chondrocytes. Expression of type II collagen and aggrecan were decreased in Ccn2?/? chondrocytes, confirming a defect in ECM production. Ccn2?/? chondrocytes also exhibited impaired DNA synthesis and reduced adhesion to fibronectin. This latter defect is associated with decreased expression of ?5 integrin. Moreover, CCN2 can bind to integrin ?5?1 in chondrocytes and can stimulate increased expression of integrin ?5. Consistent with an essential role for CCN2 as a ligand for integrins, immunofluorescence and Western blot analysis revealed that levels of focal adhesion kinase (FAK) and extracellular signal-regulated kinase (ERK)1/2 phosphorylation were reduced in Ccn2?/? chondrocytes. These findings argue that CCN2 exerts major effects in chondrocytes through its ability to (1) regulate ECM production and integrin ?5 expression, (2) engage integrins and (3) activate integrin-mediated signaling pathways.

Nishida, Takashi; Kawaki, Harumi; Baxter, Ruth M.; DeYoung, R. Andrea; Takigawa, Masaharu

2007-01-01

329

Gene targeting reveals the role of Oc90 as the essential organizer of the otoconial organic matrix  

PubMed Central

A critical part of the functional development of our peripheral balance system is the embryonic formation of otoconia, composite crystals that overlie and provide optimal stimulus input to the sensory epithelium of the gravity receptor in the inner ear. To date neither the functions of otoconial proteins nor the processes of crystal formation are clearly defined. Using gene targeting and protein analysis strategies, we demonstrate that the predominant mammalian otoconin, otoconin-90/95 (Oc90), is essential for formation of the organic matrix of otoconia by specifically recruiting other matrix components, which includes otolin, a novel mammalian otoconin that we identified to be in wildtype murine otoconia. We show that this matrix controls otoconia growth and morphology by embedding the crystallites during seeding and growth. During otoconia development, the organic matrix forms prior to CaCO3 deposition and provides optimal calcification efficiency. Histological and ultrastructural examinations show normal inner ear epithelial morphology but reduced acellular matrices, including otoconial, cupular and tectorial membranes, in Oc90 null mice, likely due to an absence of Oc90 and a profound reduction of otolin. Our data demonstrate the critical roles of otoconins in otoconia seeding, growth and anchoring and suggest mechanistic similarities and differences between otoconia and bone calcification.

Zhao, Xing; Yang, Hua; Yamoah, Ebenezer N; Lundberg, Yunxia Wang

2007-01-01

330

A glgC Gene Essential Only for the First of Two Spatially Distinct Phases of Glycogen Synthesis in Streptomyces coelicolor A3(2)  

Microsoft Academic Search

By using a PCR approach based on conserved regions of ADP-glucose pyrophosphorylases, a glgC gene was cloned from Streptomyces coelicolor A3(2). The deduced glgC gene product showed end-to-end relatedness to other bacterial ADP-glucose pyrophosphorylases. The glgC gene is about 1,000 kb from the leftmost chromo- some end and is not closely linked to either of the two glgB genes of

M. CRUZ MARTIN; DOMINIQUE SCHNEIDER; CELIA J. BRUTON; KEITH F. CHATER

1997-01-01

331

Association of a Variable Number of Tandem Repeats in the Endothelial Constitutive Nitric Oxide Synthase Gene With Essential Hypertension in Japanese  

Microsoft Academic Search

An impaired synthesis of nitric oxide (NO) by the vascular endothelium has been implicated in the pathogenesis of essential hypertension (EH). The possible association between a variable number of tandem repeats (VNTR) polymorphism in intron 4 of the endothelial constitutive NO synthase (ecNOS) gene and EH in Japanese subjects was investigated. A total of 123 individuals with EH and 120

Jiro Uwabo; Masayoshi Soma; Tomohiro Nakayama; Katsuo Kanmatsuse

1998-01-01

332

The Type IV Fimbrial Subunit Gene (fimA) of Dichelobacter nodosus Is Essential for Virulence, Protease Secretion, and Natural Competence  

Microsoft Academic Search

Dichelobacter nodosus is the essential causative agent of footrot in sheep. The major D. nodosus-encoded viru- lence factors that have been implicated in the disease are type IV fimbriae and extracellular proteases. To ex- amine the role of the fimbriae in virulence, allelic exchange was used to insertionally inactivate the fimA gene, which encodes the fimbrial subunit protein, from the

RUTH M. KENNAN; OM P. DHUNGYEL; RICHARD J. WHITTINGTON; JOHN R. EGERTON; JULIAN I. ROOD

2001-01-01

333

Comparison of Curcuma sp. in Yakushima with C. aeruginosa and C. zedoaria in Java by trn K gene sequence, RAPD pattern and essential oil component  

Microsoft Academic Search

As the result of a comparison of Curcuma sp. (Gajutsu in Japanese) in Yakushima Island, Japan, with Curcuma aeruginosa and C. zedoaria in Java, Indonesia, by trnK gene sequence, random amplified polymorphic DNA (RAPD) pattern and essential oil component, it is indicated that Curcuma sp. in Yakushima is more closely related to C. aeruginosa than to C. zedoaria.

Chinami Kitamura; Tetsuro Nagoe; Made Sri Prana; Andria Agusta; Kazuyoshi Ohashi; Hirotaka Shibuya

2007-01-01

334

scully, an Essential Gene of Drosophila, is Homologous to Mammalian Mitochondrial Type II L-3-hydroxyacyl-CoA Dehydrogenase\\/Amyloid-beta Peptide-binding Protein  

Microsoft Academic Search

The characterization of scully , an essential gene of Drosophila with phenocritical phases at embry- onic and pupal stages, shows its extensive homology with vertebrate type II l -3-hydroxyacyl-CoA dehydro- genase\\/ERAB. Genomic rescue demonstrates that four different lethal mutations are scu alleles, the molecular nature of which has been established. One of them, scu 3127 , generates a nonfunctional truncated

Laura Torroja; Daniel Ortuño-Sahagún; Alberto Ferrús; Barbara Hämmerle; Julio A. Barbas

1998-01-01

335

Natural Products Version 2.0: Connecting Genes to Molecules  

PubMed Central

Natural products have played a prominent role in the history of organic chemistry, and they continue to be important as drugs, biological probes, and targets of study for synthetic and analytical chemists. In this perspective, we explore how connecting Nature’s small molecules to the genes that encode them has sparked a renaissance in natural product research, focusing primarily on the biosynthesis of polyketides and nonribosomal peptides. We survey monomer biogenesis, coupling chemistries from templated and non-templated pathways, and the broad set of tailoring reactions and hybrid pathways that give rise to the diverse scaffolds and functionalization patterns of natural products. We conclude by considering two questions: What would it take to find all natural product scaffolds? What kind of scientists will be studying natural products in the future?

Walsh, Christopher T.; Fischbach, Michael A.

2009-01-01

336

The Role of Protein Interactions in Mediating Essentiality and Synthetic Lethality  

PubMed Central

Genes are characterized as essential if their knockout is associated with a lethal phenotype, and these “essential genes” play a central role in biological function. In addition, some genes are only essential when deleted in pairs, a phenomenon known as synthetic lethality. Here we consider genes displaying synthetic lethality as “essential pairs” of genes, and analyze the properties of yeast essential genes and synthetic lethal pairs together. As gene duplication initially produces an identical pair or sets of genes, it is often invoked as an explanation for synthetic lethality. However, we find that duplication explains only a minority of cases of synthetic lethality. Similarly, disruption of metabolic pathways leads to relatively few examples of synthetic lethality. By contrast, the vast majority of synthetic lethal gene pairs code for proteins with related functions that share interaction partners. We also find that essential genes and synthetic lethal pairs cluster in the protein-protein interaction network. These results suggest that synthetic lethality is strongly dependent on the formation of protein-protein interactions. Compensation by duplicates does not usually occur mainly because the genes involved are recent duplicates, but is more commonly due to functional similarity that permits preservation of essential protein complexes. This unified view, combining genes that are individually essential with those that form essential pairs, suggests that essentiality is a feature of physical interactions between proteins protein-protein interactions, rather than being inherent in gene and protein products themselves.

Talavera, David; Robertson, David L.; Lovell, Simon C.

2013-01-01

337

Chemical composition of fennel essential oil and its impact on Staphylococcus aureus exotoxin production.  

PubMed

In this study, fennel oil was isolated by hydrodistillation, and the chemical composition was determined by gas chromatography/mass spectral analysis. The antimicrobial activity of fennel oil against Staphylococcus aureus was evaluated by broth microdilution. A haemolysis assay, tumour necrosis factor (TNF) release assay, western blot, and real-time reverse transcription (RT)-PCR were applied to investigate the influence of fennel oil on the production of S. aureus virulence-related exoproteins. The data show that fennel oil, which contains a high level of trans-anethole, was active against S. aureus, with MICs ranging from 64 to 256 ?g/ml. Furthermore, fennel oil, when used at subinhibitory concentrations, could dose-dependently decrease the expression of S. aureus exotoxins, including ?-toxin, Staphylococcal enterotoxins (SEs) and toxic shock syndrome toxin 1 (TSST-1). PMID:22805920

Qiu, Jiazhang; Li, Hongen; Su, Hongwei; Dong, Jing; Luo, Mingjing; Wang, Jianfeng; Leng, Bingfeng; Deng, Yanhong; Liu, Juxiong; Deng, Xuming

2011-11-16

338

Deduced products of C4-dicarboxylate transport regulatory genes of Rhizobium leguminosarum are homologous to nitrogen regulatory gene products.  

PubMed

We have sequenced two genes dctB and dctD required for the activation of the C4-dicarboxylate transport structural gene dctA in free-living Rhizobium leguminosarum. The hydropathic profile of the dctB gene product (DctB) suggested that its N-terminal region may be located in the periplasm and its C-terminal region in the cytoplasm. The C-terminal region of DctB was strongly conserved with similar regions of the products of several regulatory genes that may act as environmental sensors, including ntrB, envZ, virA, phoR, cpxA, and phoM. The N-terminal domains of the products of several regulatory genes thought to be transcriptional activators, including ntrC, ompR, virG, phoB and sfrA. In addition, the central and C-terminal regions of DctD were strongly conserved with the products of ntrC and nifA, transcriptional activators that require the alternate sigma factor rpoN (ntrA) as co-activator. The central region of DctD also contained a potential ATP-binding domain. These results are consistent with recent results that show that rpoN product is required for dctA activation, and suggest that DctB plus DctD-mediated transcriptional activation of dctA may be mechanistically similar to NtrB plus NtrC-mediated activation of glnA in E. coli. PMID:3671068

Ronson, C W; Astwood, P M; Nixon, B T; Ausubel, F M

1987-10-12

339

SMC6 is an essential gene in mice, but a hypomorphic mutant in the ATPase domain has a mild phenotype with a range of subtle abnormalities.  

PubMed

Smc5-6 is a highly conserved protein complex related to cohesin and condensin involved in the structural maintenance of chromosomes. In yeasts the Smc5-6 complex is essential for proliferation and is involved in DNA repair and homologous recombination. siRNA depletion of genes involved in the Smc5-6 complex in cultured mammalian cells results in sensitivity to some DNA damaging agents. In order to gain further insight into its role in mammals we have generated mice mutated in the Smc6 gene. A complete knockout resulted in early embryonic lethality, demonstrating that this gene is essential in mammals. However, mutation of the highly conserved serine-994 to alanine in the ATP hydrolysis motif in the SMC6 C-terminal domain, resulted in mice with a surprisingly mild phenotype. With the neo gene selection marker in the intron following the mutation, resulting in reduced expression of the SMC6 gene, the mice were reduced in size, but fertile and had normal lifespans. When the neo gene was removed, the mice had normal size, but detailed phenotypic analysis revealed minor abnormalities in glucose tolerance, haematopoiesis, nociception and global gene expression patterns. Embryonic fibroblasts derived from the ser994 mutant mice were not sensitive to killing by a range of DNA damaging agents, but they were sensitive to the induction of sister chromatid exchanges induced by ultraviolet light or mitomycin C. They also accumulated more oxidative damage than wild-type cells. PMID:23518413

Ju, Limei; Wing, Jonathan; Taylor, Elaine; Brandt, Renata; Slijepcevic, Predrag; Horsch, Marion; Rathkolb, Birgit; Rácz, Ildikó; Becker, Lore; Hans, Wolfgang; Adler, Thure; Beckers, Johannes; Rozman, Jan; Klingenspor, Martin; Wolf, Eckhard; Zimmer, Andreas; Klopstock, Thomas; Busch, Dirk H; Gailus-Durner, Valérie; Fuchs, Helmut; de Angelis, Martin Hrab?; van der Horst, Gilbertus; Lehmann, Alan R

2013-03-18

340

AmrZ Beta-Sheet Residues Are Essential for DNA Binding and Transcriptional Control of Pseudomonas aeruginosa Virulence Genes ? †  

PubMed Central

AmrZ is a putative ribbon-helix-helix (RHH) transcriptional regulator. RHH proteins utilize residues within the ?-sheet for DNA binding, while the ?-helices promote oligomerization. AmrZ is of interest due to its dual roles as a transcriptional activator and as a repressor, regulating genes encoding virulence factors associated with both chronic and acute Pseudomonas aeruginosa infection. In this study, cross-linking revealed that AmrZ forms oligomers in solution but that the amino terminus, containing an unordered region and a ?-sheet, were not required for oligomerization. The first 12 unordered residues (extended amino terminus) contributed minimally to DNA binding. Mutagenesis of the AmrZ ?-sheet demonstrated that residues 18, 20, and 22 were essential for DNA binding at both activation and repressor sites, suggesting that AmrZ utilizes a similar mechanism for binding to these sites. Mice infected with amrZ mutants exhibited reduced bacterial burden, morbidity, and mortality. Direct in vivo competition assays showed a 5-fold competitive advantage for the wild type over an isogenic amrZ mutant. Finally, the reduced infection phenotype of the amrZ-null strain was similar to that of a strain expressing a DNA-binding-deficient AmrZ variant, indicating that DNA binding and transcriptional regulation by AmrZ is responsible for the in vivo virulence defect. These recent infection data, along with previously identified AmrZ-regulated virulence factors, suggest the necessity of AmrZ transcriptional regulation for optimal virulence during acute infection.

Waligora, Elizabeth A.; Ramsey, Deborah M.; Pryor, Edward E.; Lu, Haiping; Hollis, Thomas; Sloan, Gina P.; Deora, Rajendar; Wozniak, Daniel J.

2010-01-01

341

Immunocytochemical Localization of the Cystic Fibrosis Gene Product CFTR  

Microsoft Academic Search

Antisera against two peptides, corresponding to different domains of the cystic fibrosis gene product CFTR, have been raised and extensively characterized. Both antisera recognize CFTR as a 165-kDa polypeptide in Western analysis of cells transfected with CFTR cDNA as well as in epithelial cell lines. The cell and tissue distribution of CFTR has been studied by immunocytochemistry. CFTR is abundant

Isabelle Crawford; Peter C. Maloney; Pamela L. Zeitlin; William B. Guggino; Stephen C. Hyde; Helen Turley; Kevin C. Gatter; Ann Harris; Christopher F. Higgins

1991-01-01

342

Stabilization of emulsion and butter like products containing essential fatty acids using kalonji seeds extract and curcuminoids.  

PubMed

Owing to the tendency of essential fatty acids (EFAs) to undergo autoxidation, their storage becomes a key problem. Generally, they are stabilized by synthetic antioxidants like TBHQ that are toxic in nature. Recently many studies were reported where these EFAs are stabilized by natural antioxidants. In the present study, curcuminoids and kalonji seeds ethanol extract (KEE) were used to stabilize these EFAs in refined sunflower oil (RSFO), water-in-oil (w/o) emulsion and butter like products (BLPs). In RSFO, though curcuminoids alone exerted pro-oxidant effect, KEE and curcuminoids showed synergistic antioxidant activity that was comparable to TBHQ. KEE exhibited good antioxidant activity in emulsions and BLPs, providing fine physical properties like slipping point, dropping point and spreadability. EFAs increased the nutritional value of BLPs and antioxidants added for their stabilization provided their medicinal benefits. PMID:22188801

Rege, Sameera A; Momin, Shamim A; Bhowmick, Dipti N; Pratap, Amit A

2012-01-01

343

Among Developmental Regulators, StuA but Not BrlA Is Essential for Penicillin V Production in Penicillium chrysogenum? †  

PubMed Central

In filamentous fungi, secondary metabolism is often linked with developmental processes such as conidiation. In this study we analyzed the link between secondary metabolism and conidiation in the main industrial producer of the ?-lactam antibiotic penicillin, the ascomycete Penicillium chrysogenum. Therefore, we generated mutants defective in two central regulators of conidiation, the transcription factors BrlA and StuA. Inactivation of either brlA or stuA blocked conidiation and altered hyphal morphology during growth on solid media, as shown by light and scanning electron microscopy, but did not affect biomass production during liquid-submerged growth. Genome-wide transcriptional profiling identified a complex StuA- and BrlA-dependent regulatory network, including genes previously shown to be involved in development and secondary metabolism. Remarkably, inactivation of stuA, but not brlA, drastically downregulated expression of the penicillin biosynthetic gene cluster during solid and liquid-submerged growth. In agreement, penicillin V production was wild-type-like in brlA-deficient strains but 99% decreased in stuA-deficient strains during liquid-submerged growth, as shown by high-performance liquid chromatography (HPLC) analysis. Thus, among identified regulators of penicillin V production StuA has the most severe influence. Overexpression of stuA increased the transcript levels of brlA and abaA (another developmental regulator) and derepressed conidiation during liquid-submerged growth but did not affect penicillin V productivity. Taken together, these data demonstrate an intimate but not exclusive link between regulation of development and secondary metabolism in P. chrysogenum.

Sigl, Claudia; Haas, Hubertus; Specht, Thomas; Pfaller, Kristian; Kurnsteiner, Hubert; Zadra, Ivo

2011-01-01

344

RAM2, an essential gene of yeast, and RAM1 encode the two polypeptide components of the farnesyltransferase that prenylates a-factor and Ras proteins.  

PubMed Central

In the yeast Saccharomyces cerevisiae, mutations in either of two unlinked genes, RAM1 or RAM2, abolish the farnesyltransferase activity responsible for prenylation of Ras proteins and the a-factor mating pheromone. Here we report that the function of RAM1 and RAM2 genes is required for the membrane localization of Ras proteins and a-factor. The RAM2 gene was sequenced and can encode a 38-kDa protein. We examined the functional interaction of RAM2 and RAM1 by expressing the genes in Escherichia coli. Extracts derived from an E. coli strain that coexpressed RAM1 and RAM2 efficiently farnesylated a-factor peptide and Ras protein substrates. In contrast, extracts derived from E. coli strains that expressed either RAM gene alone were devoid of activity; however, when the latter extracts were mixed, protein farnesyltransferase activity was reconstituted. These results indicate that the yeast farnesyl-protein transferase is comprised of Ram1 and Ram2 polypeptides. Although Ram1 is a component of the enzyme, disruption of the RAM1 gene in yeast was not lethal, indicating that the Ram1-Ram2 farnesyltransferase is not essential for viability. In contrast, disruption of RAM2 was lethal, suggesting that Ram2 has an essential function in addition to its role with Ram1 in protein farnesylation. Images

He, B; Chen, P; Chen, S Y; Vancura, K L; Michaelis, S; Powers, S

1991-01-01

345

The nrfI gene is essential for the attachment of the active site haem group of Wolinella succinogenes cytochrome c nitrite reductase.  

PubMed

The cytochrome c nitrite reductase complex (NrfHA) is the terminal enzyme in the electron transport chain catalysing nitrite respiration of Wolinella succinogenes. The catalytic subunit NrfA is a pentahaem cytochrome c containing an active site haem group which is covalently bound via the cysteine residues of a unique CWTCK motif. The lysine residue serves as the axial ligand of the haem iron. The other four haem groups of NrfA are bound by conventional haem-binding motifs (CXXCH). The nrfHAIJ locus was restored on the genome of the W. succinogenes DeltanrfAIJ deletion mutant by integration of a plasmid, thus enabling the expression of modified alleles of nrfA and nrfI. A mutant (K134H) was constructed which contained a nrfA gene encoding a CWTCH motif instead of CWTCK. NrfA of strain K134H was found to be synthesized with five bound haem groups, as judged by matrix-assisted laser-desorption/ionization (MALDI) mass spectrometry. Its nitrite reduction activity with reduced benzyl viologen was 40% of the wild-type activity. Ammonia was formed as the only product of nitrite reduction. The mutant did not grow by nitrite respiration and its electron transport activity from formate to nitrite was 5% of that of the wild-type strain. The predicted nrfI gene product is similar to proteins involved in system II cytochrome c biogenesis. A mutant of W. succinogenes (stopI) was constructed that contained a nrfHAIJ gene cluster with the nrfI codons 47 and 48 altered to stop codons. The NrfA protein of this mutant did not catalyse nitrite reduction and lacked the active site haem group, whereas the other four haem groups were present. Mutant (K134H/stopI) which contained the K134H modification in NrfA in addition to the inactivated nrfI gene had essentially the same properties as strain K134H. NrfA from strain K134H/stopI contained five haem groups. It is concluded that NrfI is involved in haem attachment to the CWTCK motif in NrfA but not to any of the CXXCH motifs. The nrfI gene is obviously dispensable for maturation of a modified NrfA protein containing a CWTCH motif instead of CWTCK. Therefore, NrfI might function as a specific haem lyase that recognizes the active site lysine residue of NrfA. A similar role has been proposed for NrfE, F and G of Escherichia coli, although these proteins share no overall sequence similarity to NrfI and belong to system I cytochrome c biogenesis, which differs fundamentally from system II. PMID:11929530

Pisa, René; Stein, Torsten; Eichler, Robert; Gross, Roland; Simon, Jörg

2002-02-01

346

Search for genes essential for pneumococcal transformation: the RADA DNA repair protein plays a role in genomic recombination of donor DNA.  

PubMed

We applied a novel negative selection strategy called genomic array footprinting (GAF) to identify genes required for genetic transformation of the gram-positive bacterium Streptococcus pneumoniae. Genome-wide mariner transposon mutant libraries in S. pneumoniae strain R6 were challenged by transformation with an antibiotic resistance cassette and growth in the presence of the corresponding antibiotic. The GAF screen identified the enrichment of mutants in two genes, i.e., hexA and hexB, and the counterselection of mutants in 21 different genes during the challenge. Eight of the counterselected genes were known to be essential for pneumococcal transformation. Four other genes, i.e., radA, comGF, parB, and spr2011, have previously been linked to the competence regulon, and one, spr2014, was located adjacent to the essential competence gene comFA. Directed mutants of seven of the eight remaining genes, i.e., spr0459-spr0460, spr0777, spr0838, spr1259-spr1260, and spr1357, resulted in reduced, albeit modest, transformation rates. No connection to pneumococcal transformation could be made for the eighth gene, which encodes the response regulator RR03. We further demonstrated that the gene encoding the putative DNA repair protein RadA is required for efficient transformation with chromosomal markers, whereas transformation with replicating plasmid DNA was not significantly affected. The radA mutant also displayed an increased sensitivity to treatment with the DNA-damaging agent methyl methanesulfonate. Hence, RadA is considered to have a role in recombination of donor DNA and in DNA damage repair in S. pneumoniae. PMID:17631629

Burghout, Peter; Bootsma, Hester J; Kloosterman, Tomas G; Bijlsma, Jetta J E; de Jongh, Christa E; Kuipers, Oscar P; Hermans, Peter W M

2007-07-13

347

The Hansenula polymorpha PER1 Gene Is Essential for Peroxisome Biogenesis and Encodes a Peroxisomal Matrix Protein with Both Carboxy and Amino-terminal Targeting Signals  

Microsoft Academic Search

We describe the cloning of the Hansenula polymorpha PER1 gene and the characterization of the gene and its product, PER1p. The gene was cloned by functional complementation of a per1 mutant of H. polymorpha, which was impaired in the import of peroxisomal matrix proteins (Pim- phenotype). The DNA sequence of PER1 predicts that PER1p is a polypeptide of 650 amino

Hans R. Waterham; Vladimir I. Titorenko; Peter Haima; James M. Cregg; Wim Harder; Marten Veenhuis

1994-01-01

348

Inhibition of aflatoxin production and growth of Aspergillus parasiticus by Cuminum cyminum, Ziziphora clinopodioides, and Nigella sativa essential oils.  

PubMed

Aflatoxins are highly toxic and carcinogenic metabolites produced by Aspergillus parasiticus on food and agricultural commodities. Natural products may control the production of aflatoxins. The aims of this study were to evaluate the effects of the essential oils (EOs) of Cuminum cyminum, Ziziphora clinopodioides, and Nigella sativa on growth and aflatoxins production by A. parasiticus. Minimal inhibitory concentrations (MICs) and minimal fungicidal concentrations (MFCs) of the EOs were determined and compared with each other. Determination of aflatoxins (AFB(1), AFB(2), AFG(1), and AFG(2)) was performed by immunoaffinity column extraction using reverse phase-high performance liquid chromatography. The major oil components were ?-pinene (30%) in C. cyminum, pulegone (37%) in Z. clinopodioides, and trans-anthol (38.9%) in N. sativa oils. In broth microdilution method, C. cyminum oil exhibited the strongest activity (MIC(90): 1.6; MFC: 3.5?mg/mL), followed by Z. clinopodioides (MIC(90): 2.1; MFC: 5.5?mg/mL) and N. sativa (MIC(90): 2.75; MFC: 6.25?mg/mL) oils against A. parasiticus (p<0.05). Aflatoxin production was inhibited at 0.25?mg/mL of C. cyminum and Z. clinopodioides oils, of which that of C. cyminum was a stronger inhibitor. C. cyminum EO caused significant reductions in values of 94.2% for AFB(1), 100% for AFB(2), 98.9% for AFG(1), 100% for AFG(2), and 97.5% for total aflatoxin. It is concluded that the EOs of C. cyminum, Z. clinopodioides, and N. sativa could be used as natural inhibitors in foods at low concentrations to protect from fungal and toxin contaminations by A. parasiticus. PMID:21861703

Khosravi, Ali Reza; Shokri, Hojjatollah; Minooeianhaghighi, Mohammadhassan

2011-08-23

349

Post transcriptional regulation of chloroplast gene expression by nuclear encoded gene products  

SciTech Connect

The following is a review of research accomplished in the first two years of funding for the above mentioned project. The work performed is a molecular characterization of nuclear mutants of Chlamydomonas reinhardtii which are deficient in different stages in the post-transcriptional expression of a single chloroplast encoded polypeptide, the D2 protein of Photosystem II. Our long-term goals are to understand the molecular mechanisms by which nuclear gene products affect the expression of chloroplast genes. Specifically, we which to understand how specific nuclear gene products affect the turnover rate of the D2 encoding mRNA (psbD), how other nuclear encoded factors work to promote the translation of psbD mRNA and/or stabilize the D2 protein, and what the role of the D2 protein itself is in Photosystem II assembly and in the control of expression of other chloroplast genes. This progress report will be organized into four major sections concerning (I) The characterization of nuclear mutants affected in D2 translation/turnover, (II) The study of trans-acting factors which associate with the 5{prime} end of the psbD mRNA, (III) In vitro mutagenesis of the psbD gene, and (IV) Additional studies.

Kuchka, M.R.

1992-01-01

350

Identification and characterization of the herpes simplex virus type 2 gene encoding the essential capsid protein ICP32/VP19c.  

PubMed Central

We describe the characterization of the herpes simplex virus type 2 (HSV-2) gene encoding infected cell protein 32 (ICP32) and virion protein 19c (VP19c). We also demonstrate that the HSV-1 UL38/ORF.553 open reading frame (ORF), which has been shown to specify a viral protein essential for capsid formation (B. Pertuiset, M. Boccara, J. Cebrian, N. Berthelot, S. Chousterman, F. Puvian-Dutilleul, J. Sisman, and P. Sheldrick, J. Virol. 63: 2169-2179, 1989), must encode the cognate HSV type 1 (HSV-1) ICP32/VP19c protein. The region of the HSV-2 genome deduced to contain the gene specifying ICP32/VP19c was isolated and subcloned, and the nucleotide sequence of 2,158 base pairs of HSV-2 DNA mapping immediately upstream of the gene encoding the large subunit of the viral ribonucleotide reductase was determined. This region of the HSV-2 genome contains a large ORF capable of encoding two related 50,538- and 49,472-molecular-weight polypeptides. Direct evidence that this ORF encodes HSV-2 ICP32/VP19c was provided by immunoblotting experiments that utilized antisera directed against synthetic oligopeptides corresponding to internal portions of the predicted polypeptides encoded by the HSV-2 ORF or antisera directed against a TrpE/HSV-2 ORF fusion protein. The type-common immunoreactivity of the two antisera and comparison of the primary amino acid sequences of the predicted products of the HSV-2 ORF and the equivalent genomic region of HSV-1 provided evidence that the HSV-1 UL38 ORF encodes the HSV-1 ICP32/VP19c. Analysis of the expression of the HSV-1 and HSV-2 ICP32/VP19c cognate proteins indicated that there may be differences in their modes of synthesis. Comparison of the predicted structure of the HSV-2 ICP32/VP19c protein with the structures of related proteins encoded by other herpes viruses suggested that the internal capsid architecture of the herpes family of viruses varies substantially. Images

Yei, S P; Chowdhury, S I; Bhat, B M; Conley, A J; Wold, W S; Batterson, W

1990-01-01

351

Investigations of the expression of the cellular src gene product.  

PubMed

We have examined the expression of the c-src gene product in a variety of embryonic and adult tissues, in peripheral blood cells, and in cells transformed by other tumor viruses, in an attempt to identify the types of cells in which pp60c-src might provide a specific function. These studies have indicated that c-src gene expression is regulated at multiple levels in different cell types and have suggested that pp60c-src is not exclusively involved in the regulation of cell proliferation. The lowest levels of pp60c-src were found in fibroblasts, which have previously served as the standard cell type for comparisons between pp60c-src and pp60v-src. The highest levels of pp60c-src-specific kinase activity were detected in three types of cells: neurons, platelets, and polyoma virus transformed cells. In this report, we will compare the expression of pp60c-src in fibroblasts to that in platelets, neurons, and polyoma virus transformed fibroblasts. In each of the three latter cell types, the c-src gene product displays a unique pattern of expression which can be distinguished from that in fibroblasts (see diagram Fig. 1). PMID:3138228

Brugge, J S; Yonemoto, W; Lustig, A; Golden, A

1986-01-01

352

Identification of in vivo essential genes from Pseudomonas aeruginosa by PCR-based signature-tagged mutagenesis  

Microsoft Academic Search

We adapted PCR-based signature-tagged mutagenesis (STM) to Pseudomonas aeruginosa. A collection of 1056 mutants was screened in a chronic lung infection rat model. Thirteen mutants were confirmed to be attenuated. Analysis revealed that these STM mutants represented transposon insertions into eight genes previously described in databases, three genes encoding proteins sharing identity with hypothetical proteins and two genes that shared

Dario E. Lehoux; Roger C. Levesque

2002-01-01

353

Effect of salicylic acid, methyl jasmonate, ethephon and cantharidin on anthraquinone production by Rubia cordifolia callus cultures transformed with the rolB and rolC genes  

Microsoft Academic Search

It has been suggested that the rol genes of Agrobacterium rhizogenes could play an essential role in the activation of secondary metabolite production in plant transformed cultures. This study investigated whether the content of anthraquinone phytoalexins was changed in callus cultures of Rubia cordifolia transgenic for the 35S-rolB and 35S-rolC genes in comparison with a non-transformed callus culture. The anthraquinone

V. P. Bulgakov; G. K. Tchernoded; N. P. Mischenko; M. V. Khodakovskaya; V. P. Glazunov; S. V. Radchenko; E. V. Zvereva; S. A. Fedoreyev; Yu. N Zhuravlev

2002-01-01

354

Sequence and complementation analysis of recF genes from Escherichia coli, Salmonella typhimurium, Pseudomonas putida and Bacillus subtilis: evidence for an essential phosphate binding loop.  

PubMed Central

We have compared the recF genes from Escherichia coli K-12, Salmonella typhimurium, Pseudomonas putida, and Bacillus subtilis at the DNA and amino acid sequence levels. To do this we determined the complete nucleotide sequence of the recF gene from Salmonella typhimurium and we completed the nucleotide sequence of recF gene from Pseudomonas putida begun by Fujita et al. (1). We found that the RecF proteins encoded by these two genes contain respectively 92% and 38% amino acid identity with the E. coli RecF protein. Additionally, we have found that the S. typhimurium and P. putida recF genes will complement an E. coli recF mutant, but the recF gene from Bacillus subtilis [showing about 20% identity with E. coli (2)] will not. Amino acid sequence alignment of the four proteins identified four highly conserved regions. Two of these regions are part of a putative phosphate binding loop. In one region (position 36), we changed the lysine codon (which is essential for ATPase, GTPase and kinase activity in other proteins having this phosphate binding loop) to an arginine codon. We then tested this mutation (recF4101) on a multicopy plasmid for its ability to complement a recF chromosomal mutation and on the E. coli chromosome for its effect on sensitivity to UV irradiation. The strain with recF4101 on its chromosome is as sensitive as a null recF mutant strain. The strain with the plasmid-borne mutant allele is however more UV resistant than the null mutant strain. We conclude that lysine-36 and possibly a phosphate binding loop is essential for full recF activity. Lastly we made two chimeric recF genes by exchanging the amino terminal 48 amino acids of the S. typhimurium and E. coli recF genes. Both chimeras could complement E. coli chromosomal recF mutations.

Sandler, S J; Chackerian, B; Li, J T; Clark, A J

1992-01-01

355

Sequence and complementation analysis of recF genes from Escherichia coli, Salmonella typhimurium, Pseudomonas putida and Bacillus subtilis: evidence for an essential phosphate binding loop.  

PubMed

We have compared the recF genes from Escherichia coli K-12, Salmonella typhimurium, Pseudomonas putida, and Bacillus subtilis at the DNA and amino acid sequence levels. To do this we determined the complete nucleotide sequence of the recF gene from Salmonella typhimurium and we completed the nucleotide sequence of recF gene from Pseudomonas putida begun by Fujita et al. (1). We found that the RecF proteins encoded by these two genes contain respectively 92% and 38% amino acid identity with the E. coli RecF protein. Additionally, we have found that the S. typhimurium and P. putida recF genes will complement an E. coli recF mutant, but the recF gene from Bacillus subtilis [showing about 20% identity with E. coli (2)] will not. Amino acid sequence alignment of the four proteins identified four highly conserved regions. Two of these regions are part of a putative phosphate binding loop. In one region (position 36), we changed the lysine codon (which is essential for ATPase, GTPase and kinase activity in other proteins having this phosphate binding loop) to an arginine codon. We then tested this mutation (recF4101) on a multicopy plasmid for its ability to complement a recF chromosomal mutation and on the E. coli chromosome for its effect on sensitivity to UV irradiation. The strain with recF4101 on its chromosome is as sensitive as a null recF mutant strain. The strain with the plasmid-borne mutant allele is however more UV resistant than the null mutant strain. We conclude that lysine-36 and possibly a phosphate binding loop is essential for full recF activity. Lastly we made two chimeric recF genes by exchanging the amino terminal 48 amino acids of the S. typhimurium and E. coli recF genes. Both chimeras could complement E. coli chromosomal recF mutations. PMID:1542576

Sandler, S J; Chackerian, B; Li, J T; Clark, A J

1992-02-25

356

Identification of trophoblast-specific binding sites for GATA2 that are essential for rat placental lactogen-I gene expression  

Microsoft Academic Search

We identified a 3.4-kb 5?-flanking region of the rPL-I gene and examined its promoter activity using rat trophoblast Rcho-1\\u000a cells. A regulatory element between base pairs (bp) ?2,487 and ?2,310 in the 5?-flanking region was essential for maximum\\u000a promoter activity of the rPL-I gene. This regulatory element was further characterized between bp ?2,443 to ?2,415 and ?2,374\\u000a to ?2,345. Electrophoretic

Gon-Sup Kim; Yeoung-Gyu Ko; Oh-Sung Park; Hyoung Joon Park; Phil-Ok Koh; Kyu-Woan Cho; Kwan-Sik Min; Hwan-Hoo Seong; Chung-Kil Won; Jae-Hyeon Cho

2009-01-01

357

HIF-1? is essential for effective PMN bacterial killing, antimicrobial peptide production and apoptosis in Pseudomonas aeruginosa keratitis.  

PubMed

Hypoxia-inducible factor (HIF)-1?, is a transcription factor that controls energy metabolism and angiogenesis under hypoxic conditions, and a potent regulator of innate immunity. The studies described herein examined the role of HIF-1? in disease resolution in BALB/c (resistant, cornea heals) mice after ocular infection with Pseudomonas (P.) aeruginosa. Furthermore, the current studies focused on the neutrophil (PMN), the predominant cell infiltrate in keratitis. Using both siRNA and an antagonist (17-DMAG), the role of HIF-1? was assessed in P. aeruginosa-infected BALB/c mice. Clinical score and slit lamp photography indicated HIF-1? inhibition exacerbated disease and corneal destruction. Real time RT-PCR, immunohistochemistry, ELISA, Greiss and MPO assays, bacterial load, intracellular killing, phagocytosis and apoptosis assays further tested the regulatory role of HIF-1?. Despite increased pro-inflammatory cytokine expression and increased MPO levels after knocking down HIF-1? expression, in vivo studies revealed a decrease in NO production and higher bacterial load. In vitro studies using PMN provided evidence that although inhibition of HIF-1? did not affect phagocytosis, both bacterial killing and apoptosis were significantly affected, as was production of antimicrobial peptides. Overall, data provide evidence that inhibition of HIF-1? converts a normally resistant disease response to susceptible (corneal thinning and perforation) after induction of bacterial keratitis. Although this inhibition does not appear to affect PMN transmigration or phagocytosis, both in vivo and in vitro approaches indicate that the transcriptional factor is essential for effective bacterial killing, apoptosis and antimicrobial peptide production. PMID:23874197

Berger, Elizabeth A; McClellan, Sharon A; Vistisen, Kerry S; Hazlett, Linda D

2013-07-18

358

Replication of Orgyia pseudotsugata Baculovirus DNA: lef-2 and ie-1 Are Essential and ie-2, p34, and Op iap Are Stimulatory Genes  

Microsoft Academic Search

A transient DNA replication assay was used to identify genes located within m.u. 90.5-7.0 of the Orgyia pseudotsugata multinucleocapsid nuclear polyhedrosis virus (OpMNPV) genome that influenced replication of a reporter plasmid containing an OpMNPV origin of replication, when cotransfected into uninfected Lymantria dispar cells. The viral transactivator ie-1 and a 2.4-kb subclone were found to be essential for replication. The

Christian H. Ahrens; George F. Rohrmann

1995-01-01

359

CpG island methylation status and mutation analysis of the RB1 gene essential promoter region and protein-binding pocket domain in nervous system tumours  

Microsoft Academic Search

A series of 136 nervous system tumours were studied to determine the methylation status of the CpG island contained within the promoter region of the RB1 gene, as well as mutation analysis of the essential promoter region and exons 20–24 (and surrounding intronic regions) coding for the protein-binding pocket domain. Methylation of the RB1 CpG island was detected in 26

P Gonzalez-Gomez; M J Bello; M E Alonso; D Arjona; J Lomas; J M de Campos; A Isla; J A Rey

2003-01-01

360

The Relationship of Target Organ Damage and 24Hour Ambulatory Blood Pressure Monitoring with Vitamin D Receptor Gene Fok-I Polymorphism in Essential Hypertension  

Microsoft Academic Search

Background: The contribution of genetic factors in hypertension cannot be denied. Methods: In this study we evaluated the relationship between vitamin D receptor (VDR) gene polymorphisms (Bsm-I, Apa-I and Fok-I), and target organ damage in 74 patients (female\\/male 49\\/25, mean age 49.2 ± 8 years) with essential hypertension. The VDR genotypes were evaluated by polymerase chain reaction and digestion of

Eyup Kulah; Ahmet Dursun; Serefden Acikgoz; Murat Can; Sebnem Kargi; Sevil Ilikhan; Sevcan Bozdogan

2006-01-01

361

Gene and microRNA analysis of neutrophils from patients with polycythemia vera and essential thrombocytosis: down-regulation of micro RNA1 and -133a  

Microsoft Academic Search

BACKGROUND: Since the V617F mutation in JAK2 may not be the initiating event in myeloprofilerative disorders (MPDs) we compared molecular changes in neutrophils from patients with polycythemia vera (PV) and essential thrombocythosis (ET), to neutrophils stimulated by G-CSF administration and to normal unstimulated neutrophils METHODS: A gene expression oligonucleotide microarray with more than 35,000 probes and a microRNA (miR) expression

Stefanie Slezak; Ping Jin; Lorraine Caruccio; Jiaqiang Ren; Michael Bennett; Nausheen Zia; Sharon Adams; Ena Wang; Joao Ascensao; Geraldine Schechter; David Stroncek

2009-01-01

362

Involvement of distinct PKC gene products in T cell functions  

PubMed Central

It is well established that members of the protein kinase C (PKC) family seem to have important roles in T cells. Focusing on the physiological and non-redundant PKC functions established in primary mouse T cells via germline gene-targeting approaches, our current knowledge defines two particularly critical PKC gene products, PKC? and PKC?, as the “flavor of PKC” in T cells that appear to have a positive role in signaling pathways that are necessary for full antigen receptor-mediated T cell activation ex vivo and T cell-mediated immunity in vivo. Consistently, in spite of the current dogma that PKC? inhibition might be sufficient to achieve complete immunosuppressive effects, more recent results have indicated that the pharmacological inhibition of PKC?, and additionally, at least PKC?, appears to be needed to provide a successful approach for the prevention of allograft rejection and treatment of autoimmune diseases.

Pfeifhofer-Obermair, Christa; Thuille, Nikolaus; Baier, Gottfried

2012-01-01

363

Viral gene products in adenovirus type-2 transformed hamster cells.  

PubMed Central

I have analyzed viral gene products expressed in five adenovirus type 2 (Ad2)- cytoplasmic, viral RNA which was selected by hybridization to cloned restriction endonuclease fragments of Ad2 DNA. Proteins synthesized in vitro were analyzed by electrophoresis in sodium dodecyl sulfate-polyacrylamide gels and compared with those directed by RNAs prepared from productively infected cells. The early regions E1 and E4 of adenovirus type 2 (Ad2) were found to be expressed in all of five Ad2-transformed hamster embryo cells lines. RNA transcribed from early region E2, which codes for the 72,000-molecular-weight (72K) DNA-binding protein was detected in cell line HE1 only, and early region E3 was expressed exclusively in cell line HE4. RNA transcribed from the region between approximately 12 and 35 map units, coding for immediate early (13.5K, 52/53K) and immediate early proteins (13.6K, 16K, 17K, 87K), as well as RNA from late genes, was not found in any of the cell lines HE1 to HE5 had electrophoretic mobilities similar to those programmed by RNA from productively infected cells. Images

Esche, H

1982-01-01

364

Tissue Distibution of Murine Muc19/Smgc Gene Products  

PubMed Central

The recently identified gene Muc19/Smgc encodes two diverse splice variants, Smgc (submandibular gland protein C) and Muc19 (mucin 19). Muc19 is a member of the large gel-forming mucin family and is an exocrine product of sublingual mucous salivary glands in mice. SMGC is a transiently expressed secretion product of developing rodent submandibular and sublingual glands. Little is known about the expression of Muc19/Smgc gene products in other murine salivary and non-salivary tissues containing the mucous cell phenotype. Muc19 expression was therefore initially assessed by RT-PCR and immunohistochemistry. As a complementary approach, we developed a knockin mouse model, Muc19-EGFP, in which mice express a fusion protein containing the first 69 residues of Muc19 followed by enhanced green fluorescent protein (EGFP) as a marker of Muc19 expression. Results from both approaches are consistent, with preferential Muc19 expression in salivary major and minor mucous glands as well as submucosal glands of the tracheolarynx and bulbourethral glands. Evidence also indicates that individual mucous cells of minor salivary and bulbourethral glands produce another gel-forming mucin in addition to Muc19. We further find tissue expression of full-length Smgc transcripts, which encode for SMGC, and are restricted to neonatal tracheolarynx and all salivary tissues. (J Histochem Cytochem 58:141–156, 2010)

Das, Biswadip; Cash, Melanie N.; Hand, Arthur R.; Shivazad, Armin; Grieshaber, Scott S.; Robinson, Bently; Culp, David J.

2010-01-01

365

Isolation, sequencing, and disruption of the yeast CKA2 gene: casein kinase II is essential for viability in Saccharomyces cerevisiae.  

PubMed Central

Casein kinase II of Saccharomyces cerevisiae contains two distinct catalytic subunits, alpha and alpha', which are encoded by the CKA1 and CKA2 genes, respectively. Null mutations in the CKA1 gene do not confer a detectable phenotype (J. L.-P. Chen-Wu, R. Padmanabha, and C. V. C. Glover, Mol. Cell. Biol. 8:4981-4990, 1988), presumably because of the presence of the CKA2 gene. We report here the cloning, sequencing, and disruption of the CKA2 gene. The alpha' subunit encoded by the CKA2 gene is 60% identical to the CKA1-encoded alpha subunit and 55% identical to the Drosophila alpha subunit (A. Saxena, R. Padmanabha, and C. V. C. Glover, Mol. Cell. Biol. 7:3409-3417, 1987). Deletions of the CKA2 gene were constructed by gene replacement techniques. Haploid cells in which the CKA2 gene alone is disrupted show no detectable phenotype, but haploid cells carrying disruptions in both the CKA1 and CKA2 genes are inviable. Cells in which casein kinase II activity is depleted increase substantially in size prior to growth arrest, and a significant fraction of the arrested cells exhibit a pseudomycelial morphology. Disruption of the activity also results in flocculation. Yeast strains lacking both endogenous catalytic subunit genes can be rescued by expression of the alpha and beta subunits of Drosophila casein kinase II or by expression of the Drosophila alpha subunit alone, suggesting that casein kinase II function has been conserved through evolution. Images

Padmanabha, R; Chen-Wu, J L; Hanna, D E; Glover, C V

1990-01-01

366

Mutational analysis of essential IncP alpha plasmid transfer genes traF and traG and involvement of traF in phage sensitivity.  

PubMed Central

Although the broad-host-range IncP plasmids can vegetatively replicate in diverse gram-negative bacteria, the development of shuttle vector systems has established that the host range for IncP plasmid conjugative transfer is greater than the range of bacteria that sustain IncP replicons. Towards understanding IncP plasmid conjugation and the connection between IncP conjugation and Agrobacterium tumefaciens T-DNA transfer to plants, two sets of mutants were generated in the larger transfer region (Tra1) of the IncP alpha plasmid RK2. Mutagenesis strategies were chosen to minimize transcriptional polar effects. Mutant Tra1 clones were mapped, sequenced, and processed to reconstruct 49.5-kb Tra2-containing plasmid derivatives in order to assay for transfer activity and IncP plasmid-specific phage sensitivity. Focusing on the activities of the gene products of traF and traG in Escherichia coli, we found that mutations in traF abolished transfer activity and rendered the host cells phage resistant and mutations in traG abolished transfer activity but had no effect on phage sensitivity. Complementation of these mutant derivatives with corresponding trans-acting clones carrying traF or traG restored transfer activity and, in the case of the traF mutant, the phage sensitivity of the host cell. We conclude that in E. coli, both TraF and TraG are essential for IncP plasmid transfer and that TraF is necessary (but not sufficient) for donor-specific phage sensitivity, and sequencing data suggest that both TraF and TraG are membrane spanning. Images

Waters, V L; Strack, B; Pansegrau, W; Lanka, E; Guiney, D G

1992-01-01

367

The ABC-transporter hutCD genes of Photobacterium damselae subsp. piscicida are essential for haem utilization as iron source and are expressed during infection in fish.  

PubMed

The marine fish pathogen Photobacterium damselae subsp. piscicida utilizes haem compounds as the sole iron source. In a previous work, we characterized a gene cluster with ten potential haem uptake and utilization genes. Two of these genes, hutC and hutD, which are iron-regulated, conform a putative inner membrane haem ABC transporter. In this study, we constructed an insertional mutant, leading to the inactivation of hutCD genes. Reverse transcriptase-PCR analyses demonstrated that an insertion between the hutB and hutC genes abolished transcription of the downstream hutC and hutD genes. The hutCD mutant was unable to utilize haem as the sole iron source, demonstrating that the putative ABC-transporter proteins HutC and HutD are essential for haem utilization as an iron source in P. damselae subsp. piscicida. In addition, reverse transcriptase-PCR assays conducted with RNA samples isolated from experimentally infected fish revealed the presence of hutCD transcripts. The results demonstrate for the first time that haem uptake genes of a fish pathogen are expressed during the infective process in fish. PMID:20561140

Osorio, C R; Juiz-Río, S; Lemos, M L

2010-06-15

368

EDS1, an essential component of R gene-mediated disease resistance in Arabidopsis has homology to eukaryotic lipases  

Microsoft Academic Search

A major class of plant disease resistance (R) genes encodes leucine-rich-repeat proteins that possess a nucleotide binding site and amino-terminal similarity to the cytoplasmic domains of the Drosophila Toll and human IL-1 receptors. In Arabidopsis thaliana, EDS1 is indispensable for the function of these R genes. The EDS1 gene was cloned by targeted transposon tagging and found to encode a

ANDERS FALK; BART J. FEYS; LOUISE N. FROST; J ONATHAN; D. G. JONES; M ICHAEL J. DANIELS; JANE E. PARKER

1999-01-01

369

Immune-responsive gene 1 protein links metabolism to immunity by catalyzing itaconic acid production.  

PubMed

Immunoresponsive gene 1 (Irg1) is highly expressed in mammalian macrophages during inflammation, but its biological function has not yet been elucidated. Here, we identify Irg1 as the gene coding for an enzyme producing itaconic acid (also known as methylenesuccinic acid) through the decarboxylation of cis-aconitate, a tricarboxylic acid cycle intermediate. Using a gain-and-loss-of-function approach in both mouse and human immune cells, we found Irg1 expression levels correlating with the amounts of itaconic acid, a metabolite previously proposed to have an antimicrobial effect. We purified IRG1 protein and identified its cis-aconitate decarboxylating activity in an enzymatic assay. Itaconic acid is an organic compound that inhibits isocitrate lyase, the key enzyme of the glyoxylate shunt, a pathway essential for bacterial growth under specific conditions. Here we show that itaconic acid inhibits the growth of bacteria expressing isocitrate lyase, such as Salmonella enterica and Mycobacterium tuberculosis. Furthermore, Irg1 gene silencing in macrophages resulted in significantly decreased intracellular itaconic acid levels as well as significantly reduced antimicrobial activity during bacterial infections. Taken together, our results demonstrate that IRG1 links cellular metabolism with immune defense by catalyzing itaconic acid production. PMID:23610393

Michelucci, Alessandro; Cordes, Thekla; Ghelfi, Jenny; Pailot, Arnaud; Reiling, Norbert; Goldmann, Oliver; Binz, Tina; Wegner, André; Tallam, Aravind; Rausell, Antonio; Buttini, Manuel; Linster, Carole L; Medina, Eva; Balling, Rudi; Hiller, Karsten

2013-04-22

370

Effects of essential oils on methane production and fermentation by, and abundance and diversity of, rumen microbial populations.  

PubMed

Five essential oils (EOs), namely, clove oil (CLO), eucalyptus oil (EUO), garlic oil (GAO), origanum oil (ORO), and peppermint oil (PEO), were tested in vitro at 3 different doses (0.25, 0.50, and 1.0 g/liter) for their effect on methane production, fermentation, and select groups of ruminal microbes, including total bacteria, cellulolytic bacteria, archaea, and protozoa. All the EOs significantly reduced methane production with increasing doses, with reductions by 34.4%, 17.6%, 42.3%, 87%, and 25.7% for CLO, EUO, GAO, ORO, and PEO, respectively, at 1.0 g/liter compared with the control. However, apparent degradability of dry matter and neutral detergent fiber also decreased linearly with increasing doses by all EOs except GAO. The concentrations of total volatile fatty acids were not affected by GAO, EUO, or PEO but altered linearly and quadratically by CLO and ORO, respectively. All the EOs also differed in altering the molar proportions of acetate, propionate, and butyrate. As determined by quantitative real-time PCR, all the EOs decreased the abundance of archaea, protozoa, and major cellulolytic bacteria (i.e., Fibrobacter succinogenes, Ruminococcus flavefaciens, and R. albus) linearly with increasing EO doses. On the basis of denaturing gradient gel electrophoresis analysis, different EOs changed the composition of both archaeal and bacterial communities to different extents. The Shannon-Wiener diversity index (H') was reduced for archaea by all EOs in a dose-dependent manner but increased for bacteria at low and medium doses (0.25 and 0.50 g/liter) for all EOs except ORO. Due to the adverse effects on feed digestion and fermentation at high doses, a single EO may not effectively and practically mitigate methane emission from ruminants unless used at low doses in combinations with other antimethanogenic compounds. PMID:22492451

Patra, Amlan K; Yu, Zhongtang

2012-04-06

371

Initial catabolism of aromatic biogenic amines by Pseudomonas aeruginosa PAO: pathway description, mapping of mutations, and cloning of essential genes.  

PubMed Central

Pseudomonas aeruginosa PAO1 was able to utilize several aromatic biogenic amines as sole sources of carbon or nitrogen. These included the phenethylamines tyramine and dopamine and the phenethanolamines octopamine, synephrine, and norepinephrine. Initial catabolism of the phenethylamines was mediated by a membrane-bound tyramine dehydrogenase which produced 4-hydroxyphenylacetaldehyde (4HPAL) with tyramine as the substrate. The enzyme was induced by growth with both classes of amines. Initial catabolism of octopamine (except when present as the sole source of carbon and nitrogen) was mediated by a soluble enzyme with activity against the phenethanolamines but not against tyramine or dopamine. The product of the reaction with octopamine as substrate was also 4HPAL. Addition of NAD to reaction mixtures yielded 4-hydroxyphenylacetic acid and NADH. These activities, octopamine hydrolyase and 4-HPAL dehydrogenase (measured as a combined activity, OCAH-4HPALDH), were only induced by growth with phenethanolamines. However, the combined activities were not observed in extracts from cells grown with octopamine as the sole source of carbon and nitrogen, suggesting that an alternate pathway is used under this growth condition. Two independently isolated mutant strains were unable to utilize tyramine as a sole source of carbon or nitrogen. These mutants were also unable to utilize dopamine but grew at wild-type rates on the phenethanolamines. The mutations were mapped at about 70 min on the PAO1 chromosome with the chromosome-mobilizing plasmid R68.45, and both were linked to the catA1, mtu-9002, tyu-9009, and puuE mutations. DNA complementing both of the mutations was cloned on a single BamHI fragment approximately 13.8 kilobase pairs in length. Analysis of a subcloned fragment showed that the two mutations were in different genes.

Cuskey, S M; Peccoraro, V; Olsen, R H

1987-01-01

372

Gas-inducible product gene expression in bioreactors.  

PubMed

Inducible transgene expression technologies are of unmatched potential for biopharmaceutical manufacturing of unstable, growth-impairing and cytotoxic proteins as well as conditional metabolic engineering to improve desired cell phenotypes. Currently available transgene dosing modalities which rely on physical parameters or small-molecule drugs for transgene fine-tuning compromise downstream processing and/or are difficult to implement technologically. The recently designed gas-inducible acetaldehyde-inducible regulation (AIR) technology takes advantage of gaseous acetaldehyde to modulate product gene expression levels. At regulation effective concentrations gaseous acetaldehyde is physiologically inert and approved as food additive by the Federal Drug Administration (FDA). During standard bioreactor operation, gaseous acetaldehyde could simply be administered using standard/existing gas supply tubing and eventually eliminated by stripping with inducer-free air. We have determined key parameters controlling acetaldehyde transfer in three types of bioreactors and designed a mass balance-based model for optimal product gene expression fine-tuning using gaseous acetaldehyde. Operating a standard stirred-tank bioreactor set-up at 10 L scale we have validated AIR technology using CHO-K1-derived serum-free suspension cultures transgenic for gas-inducible production of human interferon-beta (IFN-beta). Gaseous acetaldehyde-inducible IFN-beta production management was fully reversible while maintaining cell viability at over 95% during the entire process. Compatible with standard bioreactor design and downstream processing procedures AIR-based technology will foster novel opportunities for pilot and large-scale manufacturing of difficult-to-produce protein pharmaceuticals. PMID:15885616

Weber, Wilfried; Rimann, Markus; de Glutz, François-Nicolas; Weber, Eric; Memmert, Klaus; Fussenegger, Martin

2005-05-01

373

Association of Insertion/Deletion Polymorphism of Alpha-Adrenoceptor Gene in Essential Hypertension with or without Type 2 Diabetes Mellitus in Malaysian Subjects  

PubMed Central

An insertion/deletion (I/D) polymorphism of Alpha2B-Adrenoceptor (ADRA2B) gene located on chromosome 2 has been studied extensively in related to cardiovascular diseases. The main aim of the present study was to examine the potential association of D allele frequency of I/D polymorphism of ADRA2B gene in Malaysian essential hypertensive subjects with or without type 2 diabetes mellitus (T2DM). This study includes 70 hypertensive subjects without T2DM, 65 hypertensive subjects with T2DM and 75 healthy volunteers as control subjects. Genotyping of I/D polymorphism was performed by conventional PCR method. There was significant difference found in age, body mass index, systolic/diastolic blood pressure and high density lipoprotein cholesterol level between the case and control subjects. DD genotypic frequency of I/D polymorphism was significantly higher in hypertensive subjects (42.84% vs. 29.33%; P­=0.029) and in hypertensive with T2DM subjects (46.15% vs. 29.33%; P=0.046) than control group. D allele frequency was higher in hypertensive group (67.41%) than control subjects (52.67%). However, no significant difference was found between the three genotypes of I/D polymorphism of ADRA2B gene and the clinical characteristics of the subjects. The result obtained in this study show D allele of ADRA2B gene was associated with essential hypertension with or without T2DM in Malaysian subjects.

Vasudevan, R.; Ismail, Patimah; Stanslas, Johnson; Shamsudin, Norashikin; Ali, Aisyah binti

2008-01-01

374

Homer1 gene products orchestrate Ca2+-permeable AMPA receptor distribution and LTP expression  

PubMed Central

We studied the role of Homer1 gene products on the presence of synaptic Ca2+-permeable AMPA receptors (AMPARs) and long-term potentiation (LTP) generation in hippocampal CA1 pyramidal neurons, using mice either lacking all Homer1 isoforms (Homer1 KO) or overexpressing the immediate early gene (IEG) product Homer1a (H1aTG). We found that Homer1 KO caused a significant redistribution of the AMPAR subunit GluA2 from the dendritic compartment to the soma. Furthermore, deletion of Homer1 enhanced the AMPAR-mediated component of glutamatergic currents at Schaffer collateral synapses as demonstrated by increased AMPA/NMDA current ratios. Meanwhile, LTP generation appeared to be unaffected. Conversely, sustained overexpression of Homer1a strongly reduced AMPA/NMDA current ratios and polyamine sensitivity of synaptic AMPAR, indicating that the proportion of synaptic GluA2-containing AMPAR increased relative to WT. LTP maintenance was abolished in H1aTG. Notably, overexpression of Homer1a in Homer1 KO or GluA2 KO mice did not affect LTP expression, suggesting activity-dependent interaction between Homer1a and long Homer1 isoforms with GluA2-containing AMPAR. Thus, Homer1a is essential for the activity-dependent regulation of excitatory synaptic transmission.

Rozov, Andrei; Zivkovic, Aleksandar R.; Schwarz, Martin K.

2012-01-01

375

Electron transfer to nitrogenase in Klebsiella pneumoniae. nifF gene cloned and the gene product, a flavodoxin, purified.  

PubMed Central

The nifF gene of Klebsiella pneumoniae was cloned into a multicopy plasmid in order to construct a strain that synthesizes and retains an elevated concentration of the gene product relative to the wild-type strain. Characterization of the isolated flavodoxin, which serves as an electron donor to nitrogenase, shows unambiguously that it is the product of the nifF gene. Images Fig. 3.

Deistung, J; Cannon, F C; Cannon, M C; Hill, S; Thorneley, R N

1985-01-01

376

Genome-wide screening of Escherichia coli genes involved in execution and promotion of cell-to-cell transfer of non-conjugative plasmids: rodZ (yfgA) is essential for plasmid acceptance in recipient cells.  

PubMed

Acquisition of new genetic traits by horizontal gene transfer is a bacterial strategy for adaptation to the environment. We previously showed that Escherichia coli can transmit non-conjugative plasmids laterally in a co-culture containing strains with and without the plasmid. In this study, using the Keio collection, a comprehensive library of E. coli knock-out mutants for non-essential genes, we screened for genes responsible for the execution and promotion of cell-to-cell plasmid transfer in recipient cells. By stepwise screening of 'transfer-down' mutants, two essential genes and six promoting genes were obtained. One of the essential genes was priA, which is involved in DNA replication. This priA mutant was also unable to be transformed by artificial transformation methods, probably due to the deficiency of the plasmid maintenance function. The other essential gene was rodZ (yfgA), a gene involved in the regulation of rod-shaped structure of E. coli cells. This rodZ mutant was transformable by all three methods of artificial transformation tested, suggesting that this gene is essential for cell-to-cell plasmid transfer but not for artificial transformation. These are the first data that suggest that rodZ plays an essential role in DNA acquisition. PMID:22497891

Kurono, Naomi; Matsuda, Ayako; Etchuya, Rika; Sobue, Rina; Sasaki, Yumi; Ito, Miki; Ando, Tsuyako; Maeda, Sumio

2012-04-03

377

Regulation of IFN-a\\/b genes: evidence for a dual function of the transcription factor complex ISGF3 in the production and action of IFN-a\\/b  

Microsoft Academic Search

Background: Efficient production of interferons (IFNs) in virally infected cells is an essential aspect of the host defence. The transcription factor complex ISGF3 (IFN-stimulated gene factor 3) was originally identified as a critical mediator of the IFN signal; it is formed upon IFN receptor (IFNR) stimulation and binds to ISREs (IFN- stimulated response elements) to activate IFN- inducible genes. It

Hisashi Harada; Masahito Matsumoto; Mitsuharu Sato; Yasuo Kashiwazaki; Tohru Kimura; Motoo Kitagawa; Taeko Yokochi; Rosemary Sok-Pin Tan; Tomohiro Takasugi; Yuzo Kadokawa; Chris Schindler; Robert D. Schreiber; Shigeru Noguchi; Tadatsugu Taniguchi

1996-01-01

378

Pseudomonas aeruginosa biofilms exposed to imipenem exhibit changes in global gene expression and beta-lactamase and alginate production.  

PubMed

The lungs of cystic fibrosis (CF) patients are commonly colonized with Pseudomonas aeruginosa biofilms. Chronic endobronchial P. aeruginosa infections are impossible to eradicate with antibiotics, but intensive suppressive antibiotic therapy is essential to maintain the lung function of CF patients. The treatment often includes beta-lactam antibiotics. How these antibiotics influence gene expression in the surviving biofilm population of P. aeruginosa is not clear. Thus, we used the microarray technology to study the effects of subinhibitory concentrations of a beta-lactam antibiotic, imipenem, on gene expression in biofilm populations. Many genes showed small but statistically significant differential expression in response to imipenem. We identified 34 genes that were induced or repressed in biofilms exposed to imipenem more than fivefold compared to the levels of induction or repression for the controls. As expected, the most strongly induced gene was ampC, which codes for chromosomal beta-lactamase. We also found that genes coding for alginate biosynthesis were induced by exposure to imipenem. Alginate production is correlated to the development of impaired lung function, and P. aeruginosa strains isolated from chronically colonized lungs of CF patients are nearly always mucoid due to the overproduction of alginate. Exposure to subinhibitory concentrations of imipenem caused structural changes in the biofilm, e.g., an increased biofilm volume. Increased levels of alginate production may be an unintended adverse consequence of imipenem treatment in CF patients. PMID:15047518

Bagge, Niels; Schuster, Martin; Hentzer, Morten; Ciofu, Oana; Givskov, Michael; Greenberg, Everett Peter; Høiby, Niels

2004-04-01

379

Two allelic genes responsible for vegetative incompatibility in the fungus Podospora anserina are not essential for cell viability  

Microsoft Academic Search

Vegetative incompatibility is a lethal reaction that destroys the heterokaryotic cells formed by the fusion of hyphae of non-isogenic strains in many fungi. That incompatibility is genetically determined is well known but the function of the genes triggering this rapid cell death is not. The two allelic incompatibility genes, s and S, of the fungus Podospora anserina were characterized. Both

B. Turcq; C. Deleu; M. Denayrolles; J. Bégueret