Isolation and characterization of two novel A20-like proteins

2001 ◽  
Vol 357 (3) ◽  
pp. 617-623 ◽  
Author(s):  
Paul C. EVANS ◽  
Ellen R. TAYLOR ◽  
John COADWELL ◽  
Karen HEYNINCK ◽  
Rudi BEYAERT ◽  
...  
Keyword(s):  
Zinc Finger ◽  
Sequence Similarity ◽  
Nuclear Factor Κb ◽  
Negative Regulator ◽  
Tumour Necrosis Factor Receptor ◽  
Isolation And Characterization ◽  
Important Negative Regulator ◽  
Inflammatory Processes ◽  
Cellular Zinc

The transcription factor nuclear factor κB (NF-κB) plays a pivotal role in inflammatory processes through induction of adhesion molecules and chemokines. The zinc finger molecule A20 is an important negative regulator of NF-κB. The mechanism utilized by A20 is not fully understood, but A20 has been shown to bind to tumour-necrosis-factor-receptor-associated factor (TRAF) molecules, which are necessary for pro-inflammatory cytokine signalling. We report two novel genes, Cezanne (cellular zinc finger anti-NF-kB) and TRABID (TRAF-bnding domain), with sequence similarity to A20. Co-immunoprecipitation studies indicated that TRAF6 was able to interact with both Cezanne and TRABID. In contrast, reporter gene experiments revealed a specific ability of Cezanne to down-regulate NF-κB. It is likely, therefore, that Cezanne participates in the regulation of inflammatory processes.

scholarly journals Single amino acid exchanges in separate domains of the Drosophila serendipity delta zinc finger protein cause embryonic and sex biased lethality.

Genetics ◽  
1992 ◽  
Vol 131 (4) ◽  
pp. 905-916 ◽  
Author(s):  
M Crozatier ◽  
K Kongsuwan ◽  
P Ferrer ◽  
J R Merriam ◽  
J A Lengyel ◽  
...  
Keyword(s):  
Amino Acid ◽  
Dna Binding ◽  
Zinc Finger ◽  
Zinc Finger Protein ◽  
Essential Gene ◽  
Larval Instar ◽  
Single Amino Acid ◽  
Isolation And Characterization ◽  
Finger Protein

Abstract The Drosophila serendipity (sry) delta (delta) zinc finger protein is a sequence-specific DNA binding protein, maternally inherited by the embryo and present in nuclei of transcriptionally active cells throughout fly development. We report here the isolation and characterization of four ethyl methanesulfate-induced zygotic lethal mutations of different strengths in the sry delta gene. For the stronger allele, all of the lethality occurs during late embryogenesis or the first larval instar. In the cases of the three weaker alleles, most of the lethality occurs during pupation; moreover, those adult escapers that emerge are sterile males lacking partially or completely in spermatozoa bundles. Genetic analysis of sry delta thus indicates that it is an essential gene, whose continued expression throughout the life cycle, notably during embryogenesis and pupal stage, is required for viability. Phenotypic analysis of sry delta hemizygote escaper males further suggests that sry delta may be involved in regulation of two different sets of genes: genes required for viability and genes involved in gonadal development. All four sry delta alleles are fully rescued by a wild-type copy of sry delta, but not by an additional copy of the sry beta gene, reinforcing the view that, although structurally related, these two genes exert distinct functions. Molecular characterization of the four sry delta mutations revealed that these mutations correspond to single amino acid replacements in the sry delta protein. Three of these replacements map to the same (third out of seven) zinc finger in the carboxy-terminal DNA binding domain; interestingly, none affects the zinc finger consensus residues. The fourth mutation is located in the NH2-proximal part of the protein, in a domain proposed to be involved in specific protein-protein interactions.

scholarly journals Isolation and characterization of PEP3, a gene required for vacuolar biogenesis in Saccharomyces cerevisiae

1991 ◽  
Vol 11 (12) ◽  
pp. 5801-5812
Author(s):  
R A Preston ◽  
M F Manolson ◽  
K Becherer ◽  
E Weidenhammer ◽  
D Kirkpatrick ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Product ◽  
Zinc Finger ◽  
Genomic Library ◽  
Sequence Similarity ◽  
Carboxypeptidase Y ◽  
Lag Phase ◽  
Significant Similarity ◽  
Reading Frame ◽  
Isolation And Characterization

The Saccharomyces cerevisiae PEP3 gene was cloned from a wild-type genomic library by complementation of the carboxypeptidase Y deficiency in a pep3-12 strain. Subclone complementation results localized the PEP3 gene to a 3.8-kb DNA fragment. The DNA sequence of the fragment was determined; a 2,754-bp open reading frame predicts that the PEP3 gene product is a hydrophilic, 107-kDa protein that has no significant similarity to any known protein. The PEP3 predicted protein has a zinc finger (CX2CX13CX2C) near its C terminus that has spacing and slight sequence similarity to the adenovirus E1a zinc finger. A radiolabeled PEP3 DNA probe hybridized to an RNA transcript of 3.1 kb in extracts of log-phase and diauxic lag-phase cells. Cells bearing pep3 deletion/disruption alleles were viable, had decreased levels of protease A, protease B, and carboxypeptidase Y antigens, had decreased repressible alkaline phosphatase activity, and contained very few normal vacuolelike organelles by fluorescence microscopy and electron microscopy but had an abundance of extremely small vesicles that stained with carboxyfluorescein diacetate, were severely inhibited for growth at 37 degrees C, and were incapable of sporulating (as homozygotes). Fractionation of cells expressing a bifunctional PEP3::SUC2 fusion protein indicated that the PEP3 gene product is present at low abundance in both log-phase and stationary cells and is a vacuolar peripheral membrane protein. Sequence identity established that PEP3 and VPS18 (J. S. Robinson, T. R. Graham, and S. D. Emr, Mol. Cell. Biol. 11:5813-5824, 1991) are the same gene.

scholarly journals Isolation and characterization of the SPT2 gene, a negative regulator of Ty-controlled yeast gene expression.

1985 ◽  
Vol 5 (7) ◽  
pp. 1543-1553 ◽  
Author(s):  
G S Roeder ◽  
C Beard ◽  
M Smith ◽  
S Keranen
Keyword(s):  
Gene Product ◽  
Regulatory Region ◽  
Negative Regulator ◽  
Transcriptional Level ◽  
Wild Type ◽  
Isolation And Characterization ◽  
Mutant Strains ◽  
Insertion Mutations ◽  
His4 Gene

The his4-917 mutation of Saccharomyces cerevisiae results from the insertion of the Ty element Ty917 into the regulatory region of the HIS4 gene and renders the cell His-. The hist4-912 delta mutant, which carries a solo delta in the 5'-noncoding region of HIS4, is His+ at 37 degrees C but His- at 23 degrees C. Both these mutations interfere with HIS4 expression at the transcriptional level. The His- phenotype of both insertion mutations is suppressed by mutations at the SPT2 locus. The product of the wild-type SPT2 gene apparently represses HIS4 transcription in these mutant strains; this repression is relieved when the SPT2 gene is destroyed by mutation. The repression of transcription by SPT2 presumably results from an interaction between the SPT2+ gene product and Ty or delta sequences. In this paper, we report the cloning and DNA sequence analysis of the wild-type SPT2 gene and show that the gene is capable of encoding a protein of 333 amino acids in length. In addition, we show that a dominant mutation of the SPT2 gene results from the generation of an ochre codon which is presumed to lead to a shortened SPT2 gene product.

scholarly journals Isolation and Characterization of the Gene Coding for the Amidinotransferase Involved in the Biosynthesis of Phaseolotoxin in Pseudomonas syringae pv. phaseolicola

2001 ◽  
Vol 14 (4) ◽  
pp. 545-554 ◽  
Author(s):  
Gustavo Hernández-Guzmán ◽  
Ariel Alvarez-Morales
Keyword(s):  
Aspartic Acid ◽  
Pseudomonas Syringae ◽  
Sequence Similarity ◽  
Insertion Mutant ◽  
Halo Blight ◽  
Streptomyces Spp ◽  
Isolation And Characterization ◽  
Gene Coding ◽  
Blight Disease

Pseudomonas syringae pv. phaseolicola is the causal agent of the “halo blight” disease of beans. A key component in the development of the disease is a nonhost-specific toxin, Nδ-(N'-sulphodiaminophosphinyl)-ornithyl-alanyl-homoarginine, known as phaseolotoxin. The homoarginine residue in this molecule has been suggested to be the product of Larginine:lysine amidinotransferase activity, previously detected in extracts of P. syringae pv. phaseolicola grown under conditions of phaseolotoxin production. We report the isolation and characterization of an amidinotransferase gene (amtA) from P. syringae pv. phaseolicola coding for a polypeptide of 362 residues (41.36 kDa) and showing approximately 40% sequence similarity to Larginine:inosamine-phosphate amidinotransferase from three species of Streptomyces spp. and 50.4% with an Larginine:glycine amidinotransferase from human mitochondria. The cysteine, histidine, and aspartic acid residues involved in substrate binding are conserved. Furthermore, expression of the amtA and argK genes and phaseolotoxin production occurs at 18°C but not at 28°C. An amidinotransferase insertion mutant was obtained that lost the capacity to synthesize homoarginine and phaseolotoxin. These results show that the amtA gene isolated is responsible for the amidinotransferase activity detected previously and that phaseolotoxin production depends upon the activity of this gene.

Isolation and Characterization of Soil Myxobacteria from Nepal

2019 ◽  
Vol 24 (2) ◽  
pp. 7-16
Author(s):  
Nabin Rana ◽  
Saraswoti Khadka ◽  
Bishnu Prasad Marasini ◽  
Bishnu Joshi ◽  
Pramod Poudel ◽  
...  
Keyword(s):  
Sequence Similarity ◽  
Antimicrobial Properties ◽  
Antimicrobial Activities ◽  
Rrna Gene ◽  
Isolation And Characterization ◽  
The Family ◽  
Global Concern ◽  
Bacteria And Fungi ◽  
Antimicrobial Metabolites

 Realizing myxobacteria as a potential source of antimicrobial metabolites, we pursued research to isolate myxobacteria showing antimicrobial properties. We have successfully isolated three strains (NR-1, NR-2, NR-3) using the Escherichia coli baiting technique. These isolates showed typical myxobacterial growth characteristics. Phylogenetic analysis showed that all the strains (NR-1, NR-2, NR-3) belong to the family Archangiaceae, suborder Cystobacterineae, and order Myxococcales. Furthermore, 16S rRNA gene sequence similarity searched through BLAST revealed that strain NR-1 showed the closest similarity (91.8 %) to the type strain Vitiosangium cumulatum (NR-156939), NR-2 showed (98.8 %) to the type of Cystobacter badius (NR-043940), and NR-3 showed the closest similarity (83.5 %) to the type of strain Cystobacter fuscus (KP-306730). All isolates showed better growth in 0.5-1 % NaCl and pH around 7.0, whereas no growth was observed at pH 9.0 and below 5.0. All strains showed better growth at 32° C and hydrolyzed starch, whereas casein was efficiently hydrolyzed by NR-1 and NR-2. Besides, preliminary antimicrobial tests from crude extracts showed activities against Gram-positive, Gram-negative bacteria, and fungi. Our findings suggest that the arcane soil habitats of Nepal harbor myxobacteria with the capability to produce diverse antimicrobial activities that may be explored to overcome the rapidly rising global concern about antibiotic resistance.

scholarly journals Isolation and Characterization of Novel Psychrophilic, Neutrophilic, Fe-Oxidizing, Chemolithoautotrophic α- and γ-Proteobacteria from the Deep Sea

2003 ◽  
Vol 69 (5) ◽  
pp. 2906-2913 ◽  
Author(s):  
K. J. Edwards ◽  
D. R. Rogers ◽  
C. O. Wirsen ◽  
T. M. McCollom
Keyword(s):  
Deep Sea ◽  
Sequence Similarity ◽  
Heterotrophic Bacterium ◽  
Physiological Characterization ◽  
Metalliferous Sediments ◽  
Basalt Glass ◽  
Isolation And Characterization ◽  
Iron Oxidizing Bacteria ◽  
Microaerobic Conditions

ABSTRACT We report the isolation and physiological characterization of novel, psychrophilic, iron-oxidizing bacteria (FeOB) from low-temperature weathering habitats in the vicinity of the Juan de Fuca deep-sea hydrothermal area. The FeOB were cultured from the surfaces of weathered rock and metalliferous sediments. They are capable of growth on a variety of natural and synthetic solid rock and mineral substrates, such as pyrite (FeS2), basalt glass (∼10 wt% FeO), and siderite (FeCO3), as their sole energy source, as well as numerous aqueous Fe substrates. Growth temperature characteristics correspond to the in situ environmental conditions of sample origin; the FeOB grow optimally at 3 to 10°C and at generation times ranging from 57 to 74 h. They are obligate chemolithoautotrophs and grow optimally under microaerobic conditions in the presence of an oxygen gradient or anaerobically in the presence of nitrate. None of the strains are capable of using any organic or alternate inorganic substrates tested. The bacteria are phylogenetically diverse and have no close Fe-oxidizing or autotrophic relatives represented in pure culture. One group of isolates are γ-Proteobacteria most closely related to the heterotrophic bacterium Marinobacter aquaeolei (87 to 94% sequence similarity). A second group of isolates are α-Proteobacteria most closely related to the deep-sea heterotrophic bacterium Hyphomonas jannaschiana (81 to 89% sequence similarity). This study provides further evidence for the evolutionarily widespread capacity for Fe oxidation among bacteria and suggests that FeOB may play an unrecognized geomicrobiological role in rock weathering in the deep sea.

scholarly journals Two distinct signalling cascades target the NF-κB regulatory factor c-IAP1 for degradation

2009 ◽  
Vol 420 (1) ◽  
pp. 83-91 ◽  
Author(s):  
Rebecca A. Csomos ◽  
Casey W. Wright ◽  
Stefanie Galbán ◽  
Karolyn A. Oetjen ◽  
Colin S. Duckett
Keyword(s):  
Large Cell ◽  
Nuclear Factor Κb ◽  
Negative Regulator ◽  
Tumour Necrosis Factor Receptor ◽  
Physiological Mechanisms ◽  
Cellular Processes ◽  
Signalling Cascades ◽  
New Gene ◽  
Factor C ◽  
Necrosis Factor

c-IAP1 (cellular inhibitor of apoptosis 1) has recently emerged as a negative regulator of the non-canonical NF-κB (nuclear factor κB) signalling cascade. Whereas synthetic IAP inhibitors have been shown to trigger the autoubiquitination and degradation of c-IAP1, less is known about the physiological mechanisms by which c-IAP1 stability is regulated. In the present paper, we describe two distinct cellular processes that lead to the targeted loss of c-IAP1. Recruitment of a TRAF2 (tumour necrosis factor receptor-associated factor 2)–c-IAP1 complex to the cytoplasmic domain of the Hodgkin's/anaplastic large-cell lymphoma-associated receptor, CD30, leads to the targeting and degradation of the TRAF2–c-IAP1 heterodimer through a mechanism requiring the RING (really interesting new gene) domain of TRAF2, but not c-IAP1. In contrast, the induced autoubiquitination of c-IAP1 by IAP antagonists causes the selective loss of c-IAP1, but not TRAF2, thereby releasing TRAF2. Thus c-IAP1 can be targeted for degradation by two distinct processes, revealing the critical importance of this molecule as a regulator of numerous intracellular signalling cascades.

Isolation and characterization of the murine zinc finger coding gene, ZT2: expression in normal and transformed myogenic cells

Gene ◽  
1999 ◽  
Vol 230 (1) ◽  
pp. 81-90 ◽  
Author(s):  
Sabrina Giorgi ◽  
Mariarosa Polimeni ◽  
Maria I. Senni ◽  
Laura De Gregorio ◽  
Tommaso A. Dragani ◽  
...  
Keyword(s):  
Zinc Finger ◽  
Myogenic Cells ◽  
Isolation And Characterization

scholarly journals Isolation and characterization of a novel transcription factor that binds to and activates insulin control element-mediated expression.

1994 ◽  
Vol 14 (10) ◽  
pp. 6704-6714 ◽  
Author(s):  
G L Robinson ◽  
S R Cordle ◽  
E Henderson ◽  
P A Weil ◽  
G Teitelman ◽  
...  
Keyword(s):  
Beta Cell ◽  
Beta Cells ◽  
Insulin Gene ◽  
Negative Regulator ◽  
Bhlh Protein ◽  
Control Element ◽  
Sequence Element ◽  
Isolation And Characterization ◽  
Insulin Control

Pancreatic beta-cell-type-specific transcription of the insulin gene is principally regulated by a single cis-acting DNA sequence element, termed the insulin control element (ICE), which is found within the 5'-flanking region of the gene. The ICE activator is a heteromeric complex composed of an islet alpha/beta-cell-specific factor associated with the ubiquitously distributed E2A-encoded proteins (E12, E47, and E2-5). We describe the isolation and characterization of a cDNA for a protein present in alpha and beta cells, termed INSAF for insulin activator factor, which binds to and activates ICE-mediated expression. INSAF was isolated from a human insulinoma cDNA library. Transfection experiments demonstrated that INSAF activates ICE expression in insulin-expressing cells but not in non-insulin-expressing cells. Cotransfection experiments showed that activation by INSAF was inhibited by Id, a negative regulator of basic helix-loop-helix (bHLH) protein function. INSAF was also shown to associate in vitro with the bHLH protein E12. In addition, affinity-purified INSAF antiserum abolished the formation of the activator-specific ICE-binding complex. Immunohistochemical studies indicate that INSAF is restricted in terms of its expression pattern, in that INSAF appears to be detected only within the nuclei of islet pancreatic alpha and beta cells. All of these data are consistent with the proposal that INSAF is either part of the ICE activator or is antigenically related to the specific activator required for insulin gene transcription.

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