scholarly journals A Polyketide Synthase Gene, ACRTS2, Is Responsible for Biosynthesis of Host-Selective ACR-Toxin in the Rough Lemon Pathotype of Alternaria alternata

2012 ◽  
Vol 25 (11) ◽  
pp. 1419-1429 ◽  
Author(s):  
Y. Izumi ◽  
K. Ohtani ◽  
Y. Miyamoto ◽  
A. Masunaka ◽  
T. Fukumoto ◽  
...  
Keyword(s):  
Alternaria Alternata ◽  
Polyketide Synthase ◽  
Attachment Site ◽  
Toxin Production ◽  
Leaf Spot Disease ◽  
Predicted Amino Acid Sequence ◽  
Type I ◽  
Methyl Transferase ◽  
Reading Frame ◽  
Rough Lemon

The rough lemon pathotype of Alternaria alternata produces host-selective ACR-toxin and causes Alternaria leaf spot disease of rough lemon (Citrus jambhiri). The structure of ACR-toxin I (MW = 496) consists of a polyketide with an α-dihydropyrone ring in a 19-carbon polyalcohol. Genes responsible for toxin production were localized to a 1.5-Mb chromosome in the genome of the rough lemon pathotype. Sequence analysis of this chromosome revealed an 8,338-bp open reading frame, ACRTS2, that was present only in the genomes of ACR-toxin-producing isolates. ACRTS2 is predicted to encode a putative polyketide synthase of 2,513 amino acids and belongs to the fungal reducing type I polyketide synthases. Typical polyketide functional domains were identified in the predicted amino acid sequence, including β-ketoacyl synthase, acyl transferase, methyl transferase, dehydratase, β-ketoreductase, and phosphopantetheine attachment site domains. Combined use of homologous recombination-mediated gene disruption and RNA silencing allowed examination of the functional role of multiple paralogs in ACR-toxin production. ACRTS2 was found to be essential for ACR-toxin production and pathogenicity of the rough lemon pathotype of A. alternata.

scholarly journals ACTTS3 Encoding a Polyketide Synthase Is Essential for the Biosynthesis of ACT-Toxin and Pathogenicity in the Tangerine Pathotype of Alternaria alternata

2010 ◽  
Vol 23 (4) ◽  
pp. 406-414 ◽  
Author(s):  
Y. Miyamoto ◽  
A. Masunaka ◽  
T. Tsuge ◽  
M. Yamamoto ◽  
K. Ohtani ◽  
...  
Keyword(s):  
Alternaria Alternata ◽  
Polyketide Synthase ◽  
Essential Gene ◽  
Attachment Site ◽  
Toxin Production ◽  
Open Reading Frames ◽  
Brown Spot ◽  
Bac Clone ◽  
Chain Reactions ◽  
Highly Correlated

The tangerine pathotype of Alternaria alternata produces host-selective ACT-toxin and causes Alternaria brown spot disease of tangerine and tangerine hybrids. Sequence analysis of a genomic BAC clone identified part of the ACT-toxin TOX (ACTT) gene cluster, and knockout experiments have implicated several open reading frames (ORF) contained within the cluster in the biosynthesis of ACT-toxin. One of the ORF, designated ACTTS3, encoding a putative polyketide synthase, was isolated by rapid amplification of cDNA ends and genomic/reverse transcription-polymerase chain reactions using the specific primers designed from the BAC sequences. The 7,374-bp ORF encodes a polyketide synthase with putative β-ketoacyl synthase, acyltransferase, methyltransferase, β-ketoacyl reductase, and phosphopantetheine attachment site domains. Genomic Southern blots demonstrated that ACTTS3 is present on the smallest chromosome in the tangerine pathotype of A. alternata, and the presence of ACTTS3 is highly correlated with ACT-toxin production and pathogenicity. Targeted gene disruption of two copies of ACTTS3 led to a complete loss of ACT-toxin production and pathogenicity. These results indicate that ACTTS3 is an essential gene for ACT-toxin biosynthesis in the tangerine pathotype of A. alternata and is required for pathogenicity of this fungus.

scholarly journals Role of the Pathotype-Specific ACRTS1 Gene Encoding a Hydroxylase Involved in the Biosynthesis of Host-Selective ACR-Toxin in the Rough Lemon Pathotype of Alternaria alternata

2012 ◽  
Vol 102 (8) ◽  
pp. 741-748 ◽  
Author(s):  
Yuriko Izumi ◽  
Eri Kamei ◽  
Yoko Miyamoto ◽  
Kouhei Ohtani ◽  
Akira Masunaka ◽  
...  
Keyword(s):  
Alternaria Alternata ◽  
Essential Gene ◽  
Toxin Production ◽  
Open Reading Frames ◽  
Leaf Spot Disease ◽  
Bac Clone ◽  
Biosynthesis Gene Cluster ◽  
Rough Lemon ◽  
Reading Frames

The rough lemon pathotype of Alternaria alternata produces host-selective ACR-toxin and causes Alternaria leaf spot disease of the rootstock species rough lemon (Citrus jambhiri) and Rangpur lime (C. limonia). Genes controlling toxin production were localized to a 1.5-Mb chromosome carrying the ACR-toxin biosynthesis gene cluster (ACRT) in the genome of the rough lemon pathotype. A genomic BAC clone containing a portion of the ACRT cluster was sequenced which allowed identification of three open reading frames present only in the genomes of ACR-toxin producing isolates. We studied the functional role of one of these open reading frames, ACRTS1 encoding a putative hydroxylase, in ACR-toxin production by homologous recombination-mediated gene disruption. There are at least three copies of ACRTS1 gene in the genome and disruption of two copies of this gene significantly reduced ACR-toxin production as well as pathogenicity; however, transcription of ACRTS1 and production of ACR-toxin were not completely eliminated due to remaining functional copies of the gene. RNA-silencing was used to knock down the remaining ACRTS1 transcripts to levels undetectable by reverse transcription-polymerase chain reaction. The silenced transformants did not produce detectable ACR-toxin and were not pathogenic. These results indicate that ACRTS1 is an essential gene in ACR-toxin biosynthesis in the rough lemon pathotype of A. alternata and is required for full virulence of this fungus.

scholarly journals Distribution and Characterization of AKT Homologs in the Tangerine Pathotype of Alternaria alternata

2000 ◽  
Vol 90 (7) ◽  
pp. 762-768 ◽  
Author(s):  
A. Masunaka ◽  
A. Tanaka ◽  
T. Tsuge ◽  
T. L. Peever ◽  
L. W. Timmer ◽  
...  
Keyword(s):  
Alternaria Alternata ◽  
Sequence Similarity ◽  
Structural Similarity ◽  
Toxin Production ◽  
Brown Spot ◽  
Japanese Pear ◽  
Black Rot ◽  
Acid Moiety ◽  
High Sequence Similarity ◽  
Rough Lemon

The tangerine pathotype of Alternaria alternata produces a host-selective toxin (HST), known as ACT-toxin, and causes Alternaria brown spot disease of citrus. The structure of ACT-toxin is closely related to AK- and AF-toxins, which are HSTs produced by the Japanese pear and strawberry pathotypes of A. alternata, respectively. AC-, AK-, and AF-toxins are chemically similar and share a 9,10-epoxy-8-hydroxy-9-methyl-decatrienoic acid moiety. Two genes controlling AK-toxin biosynthesis (AKT1 and AKT2) were recently cloned from the Japanese pear pathotype of A. alternata. Portions of these genes were used as heterologous probes in Southern blots, that detected homologs in 13 isolates of A. alternata tangerine pathotype from Minneola tangelo in Florida. Partial sequencing of the homologs in one of these isolates demonstrated high sequence similarity to AKT1 (89.8%) and to AKT2 (90.7%). AKT homologs were not detected in nine isolates of A. alternata from rough lemon, six isolates of nonpathogenic A. alternata, and one isolate of A. citri that causes citrus black rot. The presence of homologs in the Minneola isolates and not in the rough lemon isolates, nonpathogens or black rot isolates, correlates perfectly to pathogenicity on Iyo tangerine and ACT-toxin production. Functionality of the homologs was demonstrated by detection of transcripts using reverse transcription-polymerase chain reaction (RT-PCR) in total RNA of the tangerine pathotype of A. alternata. The high sequence similarity of AKT and AKT homologs in the tangerine patho-type, combined with the structural similarity of AK-toxin and ACT-toxin, may indicate that these homologs are involved in the biosynthesis of the decatrienoic acid moiety of ACT-toxin.

scholarly journals Putative Polyketide Synthase and Laccase Genes for Biosynthesis of Aurofusarin in Gibberella zeae

2005 ◽  
Vol 71 (4) ◽  
pp. 1701-1708 ◽  
Author(s):  
Jung-Eun Kim ◽  
Kap-Hoon Han ◽  
Jianming Jin ◽  
Hun Kim ◽  
Jin-Cheol Kim ◽  
...  
Keyword(s):  
Polyketide Synthase ◽  
Insertion Site ◽  
Amino Acid Identity ◽  
Gibberella Zeae ◽  
Type I ◽  
Hindiii Site ◽  
Genome Database ◽  
Reading Frame ◽  
Targeted Gene Deletion ◽  
Laccase Genes

ABSTRACT Mycelia of Gibberella zeae (anamorph, Fusarium graminearum), an important pathogen of cereal crops, are yellow to tan with white to carmine red margins. We isolated genes encoding the following two proteins that are required for aurofusarin biosynthesis from G. zeae: a type I polyketide synthase (PKS) and a putative laccase. Screening of insertional mutants of G. zeae, which were generated by using a restriction enzyme-mediated integration procedure, resulted in the isolation of mutant S4B3076, which is a pigment mutant. In a sexual cross of the mutant with a strain with normal pigmentation, the pigment mutation was linked to the inserted vector. The vector insertion site in S4B3076 was a HindIII site 38 bp upstream from an open reading frame (ORF) on contig 1.116 in the F. graminearum genome database. The ORF, designated Gip1 (for Gibberella zeae pigment mutation 1), encodes a putative laccase. A 30-kb region surrounding the insertion site and Gip1 contains 10 additional ORFs, including a putative ORF identified as PKS12 whose product exhibits about 40% amino acid identity to the products of type I fungal PKS genes, which are involved in pigment biosynthesis. Targeted gene deletion and complementation analyses confirmed that both Gip1 and PKS12 are required for aurofusarin production in G. zeae. This information is the first information concerning the biosynthesis of these pigments by G. zeae and could help in studies of their toxicity in domesticated animals.

scholarly journals A Kaposi's Sarcoma-Associated Herpesvirus Protein That Forms Inhibitory Complexes with Type I Interferon Receptor Subunits, Jak and STAT Proteins, and Blocks Interferon-Mediated Signal Transduction

2009 ◽  
Vol 83 (10) ◽  
pp. 5056-5066 ◽  
Author(s):  
Sabine A. Bisson ◽  
Anne-Laure Page ◽  
Don Ganem
Keyword(s):  
Kaposi's Sarcoma ◽  
Human Tumor ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Type I Interferons ◽  
Type I ◽  
Type I Ifn ◽  
Reading Frame ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Type I interferons (IFNs) are important mediators of innate antiviral defense and function by activating a signaling pathway through their cognate type I receptor (IFNAR). Here we report that lytic replication of Kaposi's sarcoma-associated herpesvirus (KSHV) efficiently blocks type I IFN signaling and that an important effector of this blockade is the viral protein RIF, the product of open reading frame 10. RIF blocks IFN signaling by formation of inhibitory complexes that contain IFNAR subunits, the Janus kinases Jak1 and Tyk2, and the STAT2 transcription factor. Activation of both Tyk2 and Jak1 is inhibited, and abnormal recruitment of STAT2 to IFNAR1 occurs despite the decrement in Tyk2 activity. As a result of these actions, phosphorylation of both STAT2 and STAT1 is impaired, with subsequent failure of ISGF3 accumulation in the nucleus. The presence in the viral genome of potent inhibitors of type I IFN signaling, along with several viral genes that block IFN induction, highlights the importance of the IFN pathway in the control of this human tumor virus infection.

scholarly journals Biosynthesis of Dictyostelium discoideum differentiation-inducing factor by a hybrid type I fatty acid–type III polyketide synthase

2006 ◽  
Vol 2 (9) ◽  
pp. 494-502 ◽  
Author(s):  
Michael B Austin ◽  
Tamao Saito ◽  
Marianne E Bowman ◽  
Stephen Haydock ◽  
Atsushi Kato ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Dictyostelium Discoideum ◽  
Polyketide Synthase ◽  
Type I ◽  
Type Iii Polyketide Synthase ◽  
Type Iii ◽  
Hybrid Type ◽  
Acid Type ◽  
Fatty Acid Type

scholarly journals Human Cytomegalovirus Open Reading Frame TRL11/IRL11 Encodes an Immunoglobulin G Fc-Binding Protein

2001 ◽  
Vol 75 (22) ◽  
pp. 11218-11221 ◽  
Author(s):  
Brendan N. Lilley ◽  
Hidde L. Ploegh ◽  
Rebecca S. Tirabassi
Keyword(s):  
Human Cytomegalovirus ◽  
Immunoglobulin G ◽  
Binding Protein ◽  
Fc Receptors ◽  
Binding Activity ◽  
Open Reading Frame ◽  
Membrane Glycoprotein ◽  
Type I ◽  
Reading Frame

ABSTRACT Several herpesviruses encode Fc receptors that may play a role in preventing antibody-mediated clearance of the virus in vivo. Human cytomegalovirus (HCMV) induces an Fc-binding activity in cells upon infection, but the gene that encodes this Fc-binding protein has not been identified. Here, we demonstrate that the HCMV AD169 open reading frame TRL11 and its identical copy, IRL11, encode a type I membrane glycoprotein that possesses IgG Fc-binding capabilities.

Cloning and characterization of an electrogenic Na/HCO3− cotransporter from the squid giant fiber lobe

2007 ◽  
Vol 292 (6) ◽  
pp. C2032-C2045 ◽  
Author(s):  
Peter M. Piermarini ◽  
Inyeong Choi ◽  
Walter F. Boron
Keyword(s):  
Giant Axon ◽  
Predicted Amino Acid Sequence ◽  
Giant Fiber ◽  
Reading Frame ◽  
Excitable Membranes ◽  
Current Voltage ◽  
Partial Cdna ◽  
Partial Cdna Clone ◽  
Current Voltage Relationship

The squid giant axon is a classic model system for understanding both excitable membranes and ion transport. To date, a Na+-driven Cl-HCO3− exchanger, sqNDCBE—related to the SLC4 superfamily and cloned from giant fiber lobe cDNA—is the only HCO3−-transporting protein cloned and characterized from a squid. The goal of our study was to clone and characterize another SLC4-like cDNA. We used degenerate PCR to obtain a partial cDNA clone (squid fiber clone 3, SF3), which we extended in both the 5′ and 3′ directions to obtain the full-length open-reading frame. The predicted amino-acid sequence of SF3 is similar to sqNDCBE, and a phylogenetic analysis of the membrane domains indicates that SF3 clusters with electroneutral Na+-coupled SLC4 transporters. However, when we measure pHi and membrane potential—or use two-electrode voltage clamping to measure currents—on Xenopus oocytes expressing SF3, the oocytes exhibit the characteristics of an electrogenic Na/HCO3− cotransporter, NBCe. That is, exposure to extracellular CO2/HCO3− not only causes a fall in pHi, followed by a robust recovery, but also causes a rapid hyperpolarization. The current-voltage relationship is also characteristic of an electrogenic NBC. The pHi recovery and current require HCO3− and Na+, and are blocked by DIDS. Furthermore, neither K+ nor Li+ can fully replace Na+ in supporting the pHi recovery. Extracellular Cl− is not necessary for the transporter to operate. Therefore, SF3 is an NBCe, representing the first NBCe characterized from an invertebrate.

Molecular characterization of cell cycle gene CDC7 from Saccharomyces cerevisiae

1986 ◽  
Vol 6 (5) ◽  
pp. 1590-1598
Author(s):  
M Patterson ◽  
R A Sclafani ◽  
W L Fangman ◽  
J Rosamond
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Genomic Dna ◽  
Genetic Recombination ◽  
Multiple Roles ◽  
Predicted Amino Acid Sequence ◽  
Cell Cycle Gene ◽  
Temperature Sensitive ◽  
Multicopy Plasmid ◽  
Reading Frame ◽  
Genomic Insert

The product of the CDC7 gene of Saccharomyces cerevisiae appears to have multiple roles in cellular physiology. It is required for the initiation of mitotic DNA synthesis. While it is not required for the initiation of meiotic DNA replication, it is necessary for genetic recombination during meiosis and for the formation of ascospores. It has also been implicated in an error-prone DNA repair pathway. Plasmids capable of complementing temperature-sensitive cdc7 mutations were isolated from libraries of yeast genomic DNA in the multicopy plasmid vectors YRp7 and YEp24. The complementing activity was localized within a 3.0-kilobase genomic DNA fragment. Genetic studies that included integration of the genomic insert at or near the CDC7 locus and marker rescue of four cdc7 alleles proved that the cloned fragment contains the yeast chromosomal CDC7 gene. The RNA transcript of CDC7 is about 1,700 nucleotides. Analysis of the nucleotide sequence of a 2.1-kilobase region of the cloned fragment revealed the presence of an open reading frame of 1,521 nucleotides that is presumed to encode the CDC7 protein. Depending on which of two possible ATG codons initiates translation, the calculated size of the CDC7 protein is 58.2 or 56 kilodaltons. Comparison of the predicted amino acid sequence of the CDC7 gene product with other known protein sequences suggests that CDC7 encodes a protein kinase.

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