Characterizing the regulatory mechanisms in Fusarium verticillioides secondary metabolism using functional genomics approaches /
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| Other Authors: | |
| Format: | Thesis eBook |
| Language: | English |
| Published: |
[College Station, Tex.] :
[Texas A&M University],
[2010]
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| Subjects: | |
| Online Access: | Link to OAK Trust copy |
| Abstract: | Fusarium verticillioides is one of the most important fungal pathogens of maize and has also received increasing attention due to its ability to produce various secondary metabolites, including fumonisin B₁ (FB₁) and bikaverin. However, little is known about the regulatory mechanisms associated with F. verticillioides secondary metabolism. In this study, I utilized functional genomics, forward and reverse genetics, proteomics, and high efficiency homologous recombination, to better understand the complex secondary metabolism regulations in F. verticillioides. First, using the reverse genetics approach, I characterized a putative protein phosphatase gene, CPP1 as a negative regulator of FB₁ biosynthesis. CPP1 gene deletion also affected multiple phenotypes such as radial growth, conidia germination rates, macroconidia formation, and hyphal swelling. Through gene complementation, I also demonstrated that the F. verticillioides CPP1 and Neurospora crassa wild-type ppe-1 gene are functionally conserved. Second, I used proteomics and quantitative real-time (qRT)-PCR, to advance our understanding of genes associated with fumonisin production. I analyzed the proteomic changes associated with the mutation in FCC1, a key positive regulator of fumonisins biosynthesis. I isolated proteins that were significantly up-regulated in either the wild-type or the fcc1 mutant, and transcriptional profiles of the genes corresponding to the selected proteins were analyzed via qRT-PCR. These genes showing expression patterns concomitant with fumonisin biosynthesis can be identified as primary targets for functional analysis. Next, I utilized REMI (Restriction Enzyme Mediated Integration) to isolate GAC1 gene, which encodes a GTPase activating protein, that serve as a negative regulator of bikaverin biosynthesis in F. verticillioides. AREA and PKS4 are downstream genes that are regulated positively and negatively by GAC1, respectively. Lastly, I generated a highly efficient homologous recombination strain of F. verticillioides. In eukaryotes, KU70 and KU80 play important roles in nonhomologous end-joining process, which leads to a high percentage of ectopic integration events during fungal transformation. By generating a KU70 gene deletion mutant (SF41), I have established a resource that will contribute to functional genomic research in F. verticillioides. |
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| Item Description: | "Major Subject: Plant Pathology" Title from author supplied metadata (automated record created 2010-03-12 12:08:51). Electronic resource. |
| Physical Description: | 1 online resource. |
| Bibliography: | Includes bibliographical references. |