scholarly journals Regulation of Fumonisin Biosynthesis in Fusarium verticillioides by a Zinc Binuclear Cluster-Type Gene, ZFR1

2004 ◽  
Vol 70 (5) ◽  
pp. 2653-2659 ◽  
Author(s):  
Joseph E. Flaherty ◽  
Charles P. Woloshuk
Keyword(s):  
Expressed Sequence Tag ◽  
Sequence Similarity ◽  
Fusarium Verticillioides ◽  
Regulatory Gene ◽  
Normal Growth ◽  
Wild Type ◽  
Binuclear Cluster ◽  
Primary And Secondary Metabolism ◽  
Type Gene ◽  
Maize Kernels

ABSTRACT Fusarium verticillioides, a pathogen of maize, produces a class of mycotoxins called fumonisins in infected kernels. In this study, a candidate regulatory gene, ZFR1, was identified in an expressed sequence tag library enriched for transcripts expressed by F. verticillioides during fumonisin B1 (FB1) biosynthesis. ZFR1 deletion mutants exhibited normal growth and development on maize kernels, but fumonisin production was reduced to less than 10% of that of the wild-type strain. ZFR1 encodes a putative protein of 705 amino acids with sequence similarity to the Zn(II)2Cys6 binuclear cluster family that are regulators of both primary and secondary metabolism in fungi. Expression of ZFR1 in colonized germ and degermed kernel tissues correlated with FB1 levels. Overexpression of ZFR1 in zfr1 mutants restored FB1 production to wild-type levels; however, FB1 was not restored in an fcc1 (Fusarium C-type cyclin) mutant by overexpression of ZFR1. The results of this study indicate that ZFR1 is a positive regulator of FB1 biosynthesis in F. verticillioides and suggest that FCC1 is required for ZFR1 function.

scholarly journals PAC1, a pH-Regulatory Gene from Fusarium verticillioides

2003 ◽  
Vol 69 (9) ◽  
pp. 5222-5227 ◽  
Author(s):  
Joseph E. Flaherty ◽  
Anna Maria Pirttilä ◽  
Burton H. Bluhm ◽  
Charles P. Woloshuk
Keyword(s):  
Fusarium Verticillioides ◽  
Regulatory Gene ◽  
Synthetic Medium ◽  
Transcriptional Regulators ◽  
Acidic Ph ◽  
Alkaline Ph ◽  
Wild Type ◽  
Ph Responsive ◽  
Alkaline Conditions ◽  
Maize Kernels

ABSTRACT Fumonisins are a group of mycotoxins that contaminate maize and cause leukoencephalomalacia in equine, pulmonary edema in swine, and promote cancer in mice. Fumonisin biosynthesis in Fusarium verticillioides is repressed by nitrogen and alkaline pH. We cloned a PACC-like gene (PAC1) from F. verticillioides. PACC genes encode the major transcriptional regulators of several pH-responsive pathways in other filamentous fungi. In Northern blot analyses, a PAC1 probe hybridized to a 2.2-kb transcript present in F. verticillioides grown at alkaline pH. A mutant of F. verticillioides with a disrupted PAC1 gene had severely impaired growth at alkaline pH. The mutant produced more fumonisin than the wild type when grown on maize kernels and in a synthetic medium buffered at an acidic pH, 4.5. The mutant, but not the wild type, also produced fumonisin B1 when mycelia were resuspended in medium buffered at an alkaline pH, 8.4. Transcription of FUM1, a gene involved in fumonisin biosynthesis, was correlated with fumonisin production. We conclude that PAC1 is required for growth at alkaline pH and that Pac1 may have a role as a repressor of fumonisin biosynthesis under alkaline conditions.

scholarly journals Methylation of Histone H3 Lysine 36 Is Required for Normal Development in Neurospora crassa

2005 ◽  
Vol 4 (8) ◽  
pp. 1455-1464 ◽  
Author(s):  
Keyur K. Adhvaryu ◽  
Stephanie A. Morris ◽  
Brian D. Strahl ◽  
Eric U. Selker
Keyword(s):  
Neurospora Crassa ◽  
Growth And Development ◽  
Histone H3 ◽  
Normal Growth ◽  
Wild Type ◽  
Set Domain ◽  
Amino Terminal ◽  
Histone H3 Gene ◽  
Type Gene

ABSTRACT The SET domain is an evolutionarily conserved domain found predominantly in histone methyltransferases (HMTs). The Neurospora crassa genome includes nine SET domain genes (set-1 through set-9) in addition to dim-5, which encodes a histone H3 lysine 9 HMT required for DNA methylation. We demonstrate that Neurospora set-2 encodes a histone H3 lysine 36 (K36) methyltransferase and that it is essential for normal growth and development. We used repeat induced point mutation to make a set-2 mutant (set-2 RIP1 ) with multiple nonsense mutations. Western analyses revealed that the mutant lacks SET-2 protein and K36 methylation. An amino-terminal fragment that includes the AWS, SET, and post-SET domains of SET-2 proved sufficient for K36 HMT activity in vitro. Nucleosomes were better substrates than free histones. The set-2 RIP1 mutant grows slowly, conidiates poorly, and is female sterile. Introducing the wild-type gene into the mutant complemented the defects, confirming that they resulted from loss of set-2 function. We replaced the wild-type histone H3 gene (hH3) with an allele producing a Lys to Leu substitution at position 36 and found that this hH3 K36L mutant phenocopied the set-2 RIP1 mutant, confirming that the observed defects in growth and development result from inability to methylate K36 of H3. Finally, we used chromatin immunoprecipitation to demonstrate that actively transcribed genes in Neurospora crassa are enriched for H3 methylated at lysines 4 and 36. Taken together, our results suggest that methylation of K36 in Neurospora crassa is essential for normal growth and development.

In planta reduction of maize seedling stalk lesions by the bacterial endophyteBacillus mojavensis

2011 ◽  
Vol 57 (6) ◽  
pp. 485-492 ◽  
Author(s):  
Charles W. Bacon ◽  
Dorothy M. Hinton
Keyword(s):  
Fusarium Verticillioides ◽  
Fusaric Acid ◽  
Maize Seedling ◽  
Wild Type ◽  
In Planta ◽  
Lesion Development ◽  
Bacillus Mojavensis ◽  
Acid Tolerant ◽  
Bacteria And Fungi ◽  
Maize Kernels

Maize ( Zea mays L.) is susceptible to infection by Fusarium verticillioides through autoinfection and alloinfection, resulting in diseases and contamination of maize kernels with the fumonisin mycotoxins. Attempts at controlling this fungus are currently being done with biocontrol agents such as bacteria, and this includes bacterial endophytes, such as Bacillus mojavensis . In addition to producing fumonisins, which are phytotoxic and mycotoxic, F. verticillioides also produces fusaric acid, which acts both as a phytotoxin and as an antibiotic. The question now is Can B. mojavensis reduce lesion development in maize during the alloinfection process, simulated by internode injection of the fungus? Mutant strains of B. mojavensis that tolerate fusaric acid were used in a growth room study to determine the development of stalk lesions, indicative of maize seedling blight, by co-inoculations with a wild-type strain of F. verticillioides and with non-fusaric acid producing mutants of F. verticillioides. Lesions were measured on 14-day-old maize stalks consisting of treatment groups inoculated with and without mutants and wild-type strains of bacteria and fungi. The results indicate that the fusaric-acid-tolerant B. mojavensis mutant reduced stalk lesions, suggesting an in planta role for this substance as an antibiotic. Further, lesion development occurred in maize infected with F. verticillioides mutants that do not produce fusaric acid, indicating a role for other phytotoxins, such as the fumonisins. Thus, additional pathological components should be examined before strains of B. mojavensis can be identified as being effective as a biocontrol agent, particularly for the control of seedling disease of maize.

scholarly journals Amylopectin Induces Fumonisin B1 Production by Fusarium verticillioides During Colonization of Maize Kernels

2005 ◽  
Vol 18 (12) ◽  
pp. 1333-1339 ◽  
Author(s):  
B. H. Bluhm ◽  
C. P. Woloshuk
Keyword(s):  
Quantitative Polymerase Chain Reaction ◽  
Fusarium Verticillioides ◽  
Animal Health ◽  
Fumonisin B1 ◽  
Polymerase Chain Reaction Analysis ◽  
Defined Medium ◽  
Wild Type ◽  
Fungal Genes ◽  
High Amylose ◽  
Maize Kernels

Fusarium verticillioides, a fungal pathogen of maize, produces fumonisin mycotoxins that adversely affect human and animal health. Basic questions remain unanswered regarding the interactions between the host plant and the fungus that lead to the accumulation of fumonisins in maize kernels. In this study, we evaluated the role of kernel endosperm composition in regulating fumonisin B1 (FB1) biosynthesis. We found that kernels lacking starch due to physiological immaturity did not accumulate FB1. Quantitative polymerase chain reaction analysis indicated that kernel development also affected the expression of fungal genes involved in FB1 biosynthesis, starch metabolism, and nitrogen regulation. A mutant strain of F. verticillioides with a disrupted α-amylase gene was impaired in its ability to produce FB1 on starchy kernels, and both the wild-type and mutant strains produced significantly less FB1 on a high-amylose kernel mutant of maize. When grown on a defined medium with amylose as the sole carbon source, the wild-type strain produced only trace amounts of FB1, but it produced large amounts of FB1 when grown on amylopectin or dextrin, a product of amylopectin hydrolysis. We conclude that amylopectin induces FB1 production in F. verticillioides. This study provides new insight regarding the interaction between the fungus and maize kernel during pathogenesis and highlights important areas that need further study.

scholarly journals Pl-Bh, an anthocyanin regulatory gene of maize that leads to variegated pigmentation.

Genetics ◽  
1993 ◽  
Vol 135 (2) ◽  
pp. 575-588 ◽  
Author(s):  
S M Cocciolone ◽  
K C Cone
Keyword(s):  
Gene Family ◽  
Dna Sequences ◽  
Sequence Similarity ◽  
Regulatory Gene ◽  
Regulatory Genes ◽  
Anthocyanin Synthesis ◽  
Mrna Levels ◽  
Wild Type ◽  
Specific Expression ◽  
Organ Specific

Abstract Anthocyanins are purple pigments that can be produced in virtually all parts of the maize plant. The spatial distribution of anthocyanin synthesis is dictated by the organ-specific expression of a few regulatory genes that control the transcription of the structural genes. The regulatory genes are grouped into families based on functional identity and DNA sequence similarity. The C1/Pl gene family consists of C1, which controls pigmentation of the kernel, and Pl, which controls pigmentation of the vegetative and floral organs. We have determined the relationship of another gene, Blotched (Bh), to the C1 gene family. Bh was originally described as a gene that conditions blotches of pigmentation in kernels homozygous for recessive c1, suggesting that Bh could functionally replace C1 in the kernel. Our genetic and molecular analyses indicate that Bh is an allele of Pl, that we designate Pl-Bh. Pl-Bh differs from wild-type Pl alleles in two respects. In contrast to the uniform pigmentation observed in plants carrying Pl, the pattern of pigmentation in plants carrying Pl-Bh is variegated. Pl-Bh leads to variegated pigmentation in virtually all tissues of the plant, including the kernel, an organ not pigmented by other Pl alleles. To address the molecular basis for the unusual pattern of expression of Pl-Bh, we cloned and sequenced the gene. The nucleotide sequence of Pl-Bh showed only a single base-pair difference from that of Pl. However, genomic DNA sequences associated with Pl-Bh were found to be hypermethylated relative to the same sequences around the wild-type Pl allele. The methylation was inversely correlated with Pl mRNA levels in variegated plant tissues. Thus, we conclude that DNA methylation may play a role in regulating Pl-Bh expression.

scholarly journals A Mutation in a Purported Regulatory Gene Affects Control of Sterol Uptake in Saccharomyces cerevisiae

1998 ◽  
Vol 180 (16) ◽  
pp. 4177-4183 ◽  
Author(s):  
James H. Crowley ◽  
Frank W. Leak ◽  
Kevin V. Shianna ◽  
Shirley Tove ◽  
Leo W. Parks
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Regulatory Gene ◽  
Metabolic Inhibitors ◽  
Sterol Synthesis ◽  
Wild Type ◽  
Reading Frame ◽  
Binuclear Cluster ◽  
Sterol Uptake ◽  
Regulatory Defect ◽  
Type Strains

ABSTRACT Aerobically growing wild-type strains of Saccharomyces cerevisiae are unable to take exogenously supplied sterols from media. This aerobic sterol exclusion is vitiated under anaerobic conditions, in heme-deficient strains, and under some conditions of impaired sterol synthesis. Mutants which can take up sterols aerobically in heme-competent cells have been selected. One of these mutations, designated upc2-1, gives a pleiotropic phenotype in characteristics as diverse as aerobic accumulation of sterols, total lipid storage, sensitivity to metabolic inhibitors, response to altered sterol structures, and cation requirements. During experiments designed to ascertain the effects of various cations on yeast with sterol alterations, it was observed that upc2-1was hypersensitive to Ca2+. Using resistance to Ca2+ as a screening vehicle, we cloned UPC2 and showed that it is YDR213W, an open reading frame on chromosome IV. This belongs to a fungal regulatory family containing the Zn(II)2Cys6 binuclear cluster DNA binding domain. The single guanine-to-adenine transition in upc2-1 gives a predicted amino acid change from glycine to aspartic acid. The regulatory defect explains the semidominance and pleiotropic effects of upc2-1.

HXK1 regulates carbon catabolism, sporulation, fumonisin B1 production and pathogenesis in Fusarium verticillioides

Microbiology ◽  
2011 ◽  
Vol 157 (9) ◽  
pp. 2658-2669 ◽  
Author(s):  
Hun Kim ◽  
Jonathon E. Smith ◽  
John B. Ridenour ◽  
Charles P. Woloshuk ◽  
Burton H. Bluhm
Keyword(s):  
Carbon Source ◽  
Regulatory Networks ◽  
Sole Carbon Source ◽  
Metabolic Response ◽  
Fusarium Verticillioides ◽  
Fumonisin B1 ◽  
Genetic Regulatory Networks ◽  
Wild Type ◽  
Increased Sensitivity ◽  
Maize Kernels

In Fusarium verticillioides, a ubiquitous pathogen of maize, virulence and mycotoxigenesis are regulated in response to the types and amounts of carbohydrates present in maize kernels. In this study, we investigated the role of a putative hexokinase-encoding gene (HXK1) in growth, development and pathogenesis. A deletion mutant (Δhxk1) of HXK1 was not able to grow when supplied with fructose as the sole carbon source, and growth was impaired when glucose, sucrose or maltotriose was provided. Additionally, the Δhxk1 mutant produced unusual swollen hyphae when provided with fructose, but not glucose, as the sole carbon source. Moreover, the Δhxk1 mutant was impaired in fructose uptake, although glucose uptake was unaffected. On maize kernels, the Δhxk1 mutant was substantially less virulent than the wild-type, but virulence on maize stalks was not impaired, possibly indicating a metabolic response to tissue-specific differences in plant carbohydrate content. Finally, disruption of HXK1 had a pronounced effect on fungal metabolites produced during colonization of maize kernels; the Δhxk1 mutant produced approximately 50 % less trehalose and 80 % less fumonisin B1 (FB1) than the wild-type. The reduction in trehalose biosynthesis likely explains observations of increased sensitivity to osmotic stress in the Δhxk1 mutant. In summary, this study links early events in carbohydrate sensing and glycolysis to virulence and secondary metabolism in F. verticillioides, and thus provides a new foothold from which the genetic regulatory networks that underlie pathogenesis and mycotoxigenesis can be unravelled and defined.

Molecular and genetic characterization of natural variants of HIV-1 Nef gene from North India and its functional implication in down-regulation of MHC-I and CD-4

2020 ◽  
Vol 18 ◽  
Author(s):  
J. Singh ◽  
L. Ronsard ◽  
M. Pandey ◽  
R. Kapoor ◽  
V.G. Ramachandran ◽  
...  
Keyword(s):  
Biological Activities ◽  
Sequence Similarity ◽  
Eukaryotic Expression ◽  
North India ◽  
Cell Surface Expression ◽  
Functional Domains ◽  
Wild Type ◽  
Down Regulation ◽  
Subtype B ◽  
Hiv 1

Background: HIV-1 Nef is an important accessory protein with multiple effector functions. Genetic studies of HIV-1 Nef gene shows extensive genetic diversity and the functional studies have been carried out mostly with Nef derived from regions dominated by subtype B (North America & Europe). Objective: This study was carried out to characterize genetic variations of the Nef gene from HIV-1 infected individuals from North-India and to find out their functional implications. Methods: The unique representative variants were sub-cloned in eukaryotic expression vector and further characterized with respect to their ability to down regulate cell surface expression of CD4 and MHC-1molecules. Results: The phylogenetic analysis of Nef variants revealed sequence similarity with either consensus subtype B or B/C recombinants. Boot scan analysis of some of our variants showed homology to B/C recombinant and some to wild type Nef B. Extensive variations were observed in most of the variants. The dN/dS ratio revealed 80% purifying selection and 20% diversifying selection implying the importance of mutations in Nef variants. Intracellular stability of Nef variants differed greatly when compared with wild type Nef B and C. There were some variants that possessed mutations in the functional domains of Nef and responsible for its differential CD4 and MHC-1 down regulation activity. Conclusion: We observed enhanced biological activities in some of the variants, perhaps arising out of amino acid substitutions in their functional domains. The CD4 and MHC-1 down-regulation activity of Nef is likely to confer immense survival advantage allowing the most rare genotype in a population to become the most abundant after a single selection event.

The DNA Binding Protein Rfg1 Is a Repressor of Filamentation inCandida albicans

Genetics ◽  
2001 ◽  
Vol 157 (4) ◽  
pp. 1503-1512 ◽  
Author(s):  
Roy A Khalaf ◽  
Richard S Zitomer
Keyword(s):  
Dna Binding ◽  
Binding Protein ◽  
Hyphal Growth ◽  
Dna Binding Protein ◽  
Homozygous Deletion ◽  
Wild Type ◽  
Pathogenic Yeast ◽  
Repression Complex ◽  
Type Gene ◽  
Repression Domain

AbstractWe have identified a repressor of hyphal growth in the pathogenic yeast Candida albicans. The gene was originally cloned in an attempt to characterize the homologue of the Saccharomyces cerevisiae Rox1, a repressor of hypoxic genes. Rox1 is an HMG-domain, DNA binding protein with a repression domain that recruits the Tup1/Ssn6 general repression complex to achieve repression. The C. albicans clone also encoded an HMG protein that was capable of repression of a hypoxic gene in a S. cerevisiae rox1 deletion strain. Gel retardation experiments using the purified HMG domain of this protein demonstrated that it was capable of binding specifically to a S. cerevisiae hypoxic operator DNA sequence. These data seemed to indicate that this gene encoded a hypoxic repressor. However, surprisingly, when a homozygous deletion was generated in C. albicans, the cells became constitutive for hyphal growth. This phenotype was rescued by the reintroduction of the wild-type gene on a plasmid, proving that the hyphal growth phenotype was due to the deletion and not a secondary mutation. Furthermore, oxygen repression of the hypoxic HEM13 gene was not affected by the deletion nor was this putative ROX1 gene regulated positively by oxygen as is the case for the S. cerevisiae gene. All these data indicate that this gene, now designated RFG1 for Repressor of Filamentous Growth, is a repressor of genes required for hyphal growth and not a hypoxic repressor.

scholarly journals Overexpression of the pitaya phosphoethanolamine N-methyltransferase gene (HpPEAMT) enhanced simulated drought stress in tobacco

2021 ◽  
Author(s):  
Ai-Hua Wang ◽  
Lan Yang ◽  
Xin-Zhuan Yao ◽  
Xiao-Peng Wen
Keyword(s):  
Drought Stress ◽  
Stress Response ◽  
Transgenic Plants ◽  
Drought Tolerance ◽  
Transgenic Tobacco ◽  
Sequence Similarity ◽  
Amino Acid Sequence Similarity ◽  
Transgenic Tobacco Plants ◽  
Wild Type ◽  
Tobacco Plants

AbstractPhosphoethanolamine N-methyltransferase (PEAMTase) catalyzes the methylation of phosphoethanolamine to produce phosphocholine and plays an important role in the abiotic stress response. Although the PEAMT genes has been isolated from many species other than pitaya, its role in the drought stress response has not yet been fully elucidated. In the present study, we isolated a 1485 bp cDNA fragment of HpPEAMT from pitaya (Hylocereus polyrhizus). Phylogenetic analysis showed that, during its evolution, HpPEAMT has shown a high degree of amino acid sequence similarity with the orthologous genes in Chenopodiaceae species. To further investigate the function of HpPEAMT, we generated transgenic tobacco plants overexpressing HpPEAMT, and the transgenic plants accumulated significantly more glycine betaine (GB) than did the wild type (WT). Drought tolerance trials indicated that, compared with those of the wild-type (WT) plants, the roots of the transgenic plants showed higher drought tolerance ability and exhibited improved drought tolerance. Further analysis revealed that overexpression of HpPEAM in Nicotiana tabacum resulted in upregulation of transcript levels of GB biosynthesis-related genes (NiBADH, NiCMO and NiSDC) in the leaves. Furthermore, compared with the wild-type plants, the transgenic tobacco plants displayed a significantly lower malondialdehyde (MDA) accumulation and higher activities of the superoxide dismutase (SOD) and peroxidase (POD) antioxidant enzymes under drought stress. Taken together, our results suggested that HpPEAMT enhanced the drought tolerance of transgenic tobacco.

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