Exploits Maize Kernels Using an "Orphan" Secondary Metabolite Cluster.

is a saprophytic cosmopolitan fungus, capable of infecting crops both pre- and post-harvest and exploiting different secondary metabolites, including aflatoxins. Aflatoxins are known carcinogens to animals and humans, but display no clear effect in host plants such as maize. In a previous study, we mined the genome of to identify secondary metabolite clusters putatively involving the pathogenesis process in maize.

Use of Dual RNA-seq for Systems Biology Analysis of and Interaction.

The interaction between and is complex, and the identification of plant genes and pathways conferring resistance to the fungus has been challenging. Therefore, the authors undertook a systems biology approach involving dual RNA-seq to determine the simultaneous response from the host and the pathogen. What was dramatically highlighted in the analysis is the uniformity in the development patterns of gene expression of the host and the pathogen during infection.

Carbon Dioxide Mediates the Response to Temperature and Water Activity Levels in Aspergillus flavus during Infection of Maize Kernels.

is a saprophytic fungus that may colonize several important crops, including cotton, maize, peanuts and tree nuts. Concomitant with colonization is its potential to secrete mycotoxins, of which the most prominent is aflatoxin. Temperature, water activity (a) and carbon dioxide (CO₂) are three environmental factors shown to influence the fungus-plant interaction, which are predicted to undergo significant changes in the next century.

Interactions between water activity and temperature on the Aspergillus flavus transcriptome and aflatoxin B production.

Effects of Aspergillus flavus colonization of maize kernels under different water activities (a; 0.99 and 0.91) and temperatures (30, 37°C) on (a) aflatoxin B (AFB) production and (b) the transcriptome using RNAseq were examined.

A Network Approach of Gene Co-expression in the / Pathosystem to Map Host/Pathogen Interaction Pathways.

A gene co-expression network (GEN) was generated using a dual RNA-seq study with the fungal pathogen and its plant host during the initial 3 days of infection. The analysis deciphered novel pathways and mapped genes of interest in both organisms during the infection. This network revealed a high degree of connectivity in many of the previously recognized pathways in such as jasmonic acid, ethylene, and reactive oxygen species (ROS).

Aspergillus flavus infection induces transcriptional and physical changes in developing maize kernels.

Maize kernels are susceptible to infection by the opportunistic pathogen Aspergillus flavus. Infection results in reduction of grain quality and contamination of kernels with the highly carcinogenic mycotoxin, aflatoxin. To understanding host response to infection by the fungus, transcription of approximately 9000 maize genes were monitored during the host-pathogen interaction with a custom designed Affymetrix GeneChip® DNA array.

Localization, morphology and transcriptional profile of Aspergillus flavus during seed colonization.

Aspergillus flavus is an opportunistic fungal pathogen that infects maize kernels pre-harvest, creating major human health concerns and causing substantial agricultural losses. Improved control strategies are needed, yet progress is hampered by the limited understanding of the mechanisms of infection. A series of studies were designed to investigate the localization, morphology and transcriptional profile of A.

Quantification of urinary aflatoxin B1 dialdehyde metabolites formed by aflatoxin aldehyde reductase using isotope dilution tandem mass spectrometry.

The aflatoxin B 1 aldehyde reductases (AFARs), inducible members of the aldo-keto reductase superfamily, convert aflatoxin B 1 dialdehyde derived from the exo- and endo-8,9-epoxides into a number of reduced alcohol products that might be less capable of forming covalent adducts with proteins. An isotope dilution tandem mass spectrometry method for quantification of the metabolites, C-8 monoalcohol, dialcohol, and C-6a monoalcohol, was developed to ascertain their possible role as urinary biomarkers for application to chemoprevention investigations. This method uses a novel (13)C 17-aflatoxin B 1 dialcohol internal standard, synthesized from (13)C 17-aflatoxin B 1 biologically produced by Aspergillus flavus.

Improved protocols for functional analysis in the pathogenic fungus Aspergillus flavus.

Background: An available whole genome sequence for Aspergillus flavus provides the opportunity to characterize factors involved in pathogenicity and to elucidate the regulatory networks involved in aflatoxin biosynthesis. Functional analysis of genes within the genome is greatly facilitated by the ability to disrupt or mis-express target genes and then evaluate their result on the phenotype of the fungus. Large-scale functional analysis requires an efficient genetic transformation system and the ability to readily select transformants with altered expression, and usually requires generation of double (or multi) gene deletion strains or the use of prototrophic strains.

Function and regulation of aflJ in the accumulation of aflatoxin early pathway intermediate in Aspergillus flavus.

aflJ resides within the aflatoxin biosynthetic gene cluster adjacent to the pathway regulatory gene aflR and is involved in aflatoxin production, but its function is unknown. Over-expression of aflJ in the aflatoxin-producing strain 86-10 resulted in increased aflatoxin. In an effort to study the function and regulation of aflJ, strain 649-1 lacking the entire biosynthetic cluster was transformed with either reporter constructs, expression constructs, or cosmid clones and analysed for gene expression or metabolite accumulation.

The effect of elevated temperature on gene transcription and aflatoxin biosynthesis.

The molecular regulation of aflatoxin biosynthesis is complex and influenced by several environmental conditions; one of these is temperature. Aflatoxins are produced optimally at 28-30 C, and production decreases as temperatures approach 37 C, the optimum temperature for fungal growth. To better characterize the influence of temperature on aflatoxin biosynthesis, we monitored the accumulation of aflatoxin and the expression of more than 5000 genes in Aspergillus flavus at 28 C and 37 C.

Elucidation of veA-dependent genes associated with aflatoxin and sclerotial production in Aspergillus flavus by functional genomics.

The aflatoxin-producing fungi, Aspergillus flavus and A. parasiticus, form structures called sclerotia that allow for survival under adverse conditions. Deletion of the veA gene in A.

Aflatoxin production in peanut lines selected to represent a range of linoleic acid concentrations.

To determine whether concentrations of linoleate in peanut (Arachis hypogaea L.) seed oil could be used to predict an ability to support aflatoxin production, seeds of genotypes representing a range of linoleate content were inoculated with Aspergillus flavus Link ex Fries and assayed for aflatoxin content. Seeds were blanched and quartered, inoculated with conidia of A.

Maize ribosome-inactivating proteins (RIPs) with distinct expression patterns have similar requirements for proenzyme activation.

Ribosome-inactivating proteins (RIPs, EC 3.2.2.

Comparison of Aflatoxin Production in Normal- and High-Oleic Backcross-Derived Peanut Lines.

The effect of the high-oleate trait of peanut on aflatoxin production was tested by comparing normal oleic lines with high-oleic backcross-derived lines. Seeds were blanched, quartered, and inoculated with Aspergillus flavus conidia, placed on moistened filter paper in petri dishes, and incubated for 8 days. In one experiment, dishes were stacked in plastic bags in a Latin square design with bags and positions in stacks as blocking variables.

Identification of genes differentially expressed during aflatoxin biosynthesis in Aspergillus flavus and Aspergillus parasiticus.

A complex regulatory network governs the biosynthesis of aflatoxin. While several genes involved in aflatoxin production are known, their action alone cannot account for its regulation. Arrays of clones from an Aspergillus flavus cDNA library and glass slide microarrays of ESTs were screened to identify additional genes.

Familial subepithelial corneal amyloidosis (gelatinous drop-like corneal dystrophy): exclusion of linkage to lactoferrin gene.

Purpose: Because corneal tissue with familial subepithelial corneal amyloidosis (FSCA; gelatinous drop-like dystrophy of the cornea) contains lactoferrin the possibility that the FSCA gene was the human lactoferrin (hLF) gene was investigated. Due to contradictory published information we also mapped the hLF gene.

Methods: We mapped the hLF gene using a genomic clone of the entire hLF gene as a probe by fluorescence in situ hybridization (FISH).

Characterization of aflJ, a gene required for conversion of pathway intermediates to aflatoxin.

The genes encoding the aflatoxin biosynthetic pathway enzymes have been localized as a cluster to a 75-kb DNA fragment. The enzymatic functions of the products of most of the genes in the cluster are known, but there are a few genes that have not yet been characterized. We report here the characterization of one of these genes, a gene designated aflJ.

Comparative analysis of BiP gene expression in maize endosperm.

Binding protein (BiP) is the endoplasmic reticulum member of the highly conserved HSP70 (heat shock protein 70) family of molecular chaperones. We have isolated and characterized two different BiP cDNA clones corresponding to genes expressed in immature kernels. These two cDNAs share extensive sequence similarity but map to unlinked loci in the maize genome.

Human estrogen receptor-like 1 (ESRL1) gene: genomic organization, chromosomal localization, and promoter characterization.

Estrogen receptor-like 1a (ESRL1a; same as estrogen receptor-related orphan receptors, ERR1) belongs to a subfamily of the nuclear receptor superfamily. We have previously shown that human ESRL1a modulates estrogen responsiveness of the lactoferrin gene promoter in transiently transfected endometrial carcinoma RL95-2 cells. In this study, we cloned and characterized the human ESRL1 gene.

A maize ribosome-inactivating protein is controlled by the transcriptional activator Opaque-2.

Although synthesis of the cytosolic maize albumin b-32 had been shown to be controlled by the Opaque-2 regulatory locus, its function was unknown. We show here that b-32 is a member of the large and widely distributed class of toxic plant proteins with ribosome-inactivating activity. These ribosome-inactivating proteins (RIPs) are RNA N-glycosidases that remove a single base from a conserved 28S rRNA loop required for elongation factor 1 alpha binding.

Characterization of an immunoglobulin binding protein homolog in the maize floury-2 endosperm mutant.

The maize b-70 protein is an endoplasmic reticulum protein overproduced in the floury-2 (fl2) endosperm mutant. The increase in b-70 levels in fl2 plants occurs during seed maturation and is endosperm specific. We have used amino acid sequence homology to identify b-70 as a homolog of mammalian immunoglobulin binding protein (BiP).