Intron architecture in Fusarium

Dissertation (MSc)--University of Pretoria, 2012.

The genus Fusarium constitutes fungi with diverse biological behaviours. This study focused on four plant pathogenic species. These were F. verticillioides which infects maize, F. oxysporum which infects tomato, F. graminearum, a pathogen of wheat and F. circinatum, which is pathogenic to pine. The genomes of F. verticillioides, F. oxysporum, F. graminearum and F. circinatum have been sequenced. These genomes were annotated using different gene prediction software. To study the architecture and distribution of Spliceosomal introns in these a set of Housekeeping (HK) genes common to all eukaryotes were used. These analyses revealed discrepancies in the annotations of these genomes, which most commonly included intron position incongruences, misidentified introns and sequencing errors. Spliceosomal introns have four cis-elements which include the 5ꞌ and 3ꞌ splice sites, the branch site and the polypyrimidine tract. Analysis of the first three elements of Spliceosomal introns in the four Fusarium species and comparisons to those in other fungi showed significant differences in the consensus sequences of these elements. Two additional branch site motifs were also found for Fusarium, while the polypyrimidine tract of these species was found to be very diverse. The results also indicated that the first introns of the HK genes of the Fusarium species significantly longer, which is consistent with what have been found for genes of other eukaryotic. Also consistent with what is known for other eukaryotes, the analysed Fusarium genes had much lower intron densities than those observed in higher eukaryotes. An average of 2.53 introns per gene was observed in Fusarium and most of these introns were located closer to the 5ꞌ end of the HK genes. This average is low compared A. thaliana and Homo sapiens which have averages of 4.3 and 8.82 introns per gene, respectively. With the aid of EST and genome data, F. circinatum was shown to harbour putative alternative introns at a frequency of approximately 0.3%. Homologues of a number of these genes from F. verticillioides, F. oxysporum and F. graminearum also harboured alternative splicing signals. Certain alternatively spliced transcripts harbored premature stop codons. These transcripts are targeted by non-sense mediated mRNA decay (NMD) system. The high rate at which these transcripts included premature stop codons suggested that such a quality control system is indeed needed for these fungi. Overall, however, it remains to be investigated whether these alternative transcripts are functional as is the case with some of them in humans, plants and insects. As more fungal genomes are being sequenced the need for accurate gene prediction methods is soaring. The incorporation of the findings of the architecture and distribution of Spliceosomal introns in Fusarium into gene prediction methods will thus increase the accuracy of such methods for Fusarium species, especially those related to F. circinatum. The identification of genes that are potentially regulated through alternative intron splicing also provide valuable targets for future studies on important biological processes such as pathogenicity and virulence. Note: The disc provided contains a spreadsheet with all the data included in this dissertation.

Microbiology and Plant Pathology

MSc

Unrestricted