Gene discovery in EST sequences from the wheat leaf rust fungus puccinia triticina sexual spores, asexual spores and haustoria, compared to other rust and corn smut fungi

Authors: XU, JUNHUAN - Agriculture And Agri-Food Canada; Fellers, John; ZHU, WENHAN - Georgia Institute Of Technology; JOLY, DAVID - Agriculture And Agri-Food Canada; EILAM, TAMAR - Tel Aviv University; BANKS, TRAVIS - Agriculture And Agri-Food Canada; MAYO, MICHAEL - Provincial Health Services Authority; ALI, JOHAR - Provincial Health Services Authority; MCCALLUM, BRENT - Agriculture And Agri-Food Canada; SAVILLE, BARRY - Trent University; LINNING, ROB - Agriculture And Agri-Food Canada; DICKENSON, MATT - University Of Nottingham; ANTONOV, IVAN - Georgia Institute Of Technology; DONALDSON, MICHAEL - Trent University; ANIKSTER, YEHOSHUA - Tel Aviv University; MONRO, SARAH - Provincial Health Services Authority; WHNHOVEN, BRIAN - Provincial Health Services Authority; MOORE, RICHARD - Provincial Health Services Authority; BORODOVSKY, MARK - Georgia Institute Of Technology; BAKKEREN, GUUS - Agriculture And Agri-Food Canada

Submitted to: BMC Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/11/2011
Publication Date: 3/24/2011
Citation: Xu, J., Fellers, J.P., Zhu, W., Joly, D., Eilam, T., Banks, T., Mayo, M., Ali, J., Mccallum, B., Saville, B., Linning, R., Dickenson, M., Antonov, I., Donaldson, M., Anikster, Y., Monro, S., Whnhoven, B., Moore, R., Borodovsky, M., Bakkeren, G. 2011. Gene discovery in EST sequences from the wheat leaf rust fungus puccinia triticina sexual spores, asexual spores and haustoria, compared to other rust and corn smut fungi. Biomed Central (BMC) Genomics. 12:161.

Interpretive Summary: Wheat leaf rust is one of the most economically important diseases of wheat worldwide. The leaf rust fungus has repeatedly shown the ability to rapidly adapt to resistant varieties that have been periodically released to producers. In order to design more durable types of resistance, novel approaches are needed. The objective of this work was to obtain a better understanding of the fungal genes that are expressed at various stages in the life cycle of the pathogen. A database of 13,328 fungal genes was constructed and compared to databases from four other related fungi. Many genes were evolutionarily conserved between five species, but over 40% of the genes from the leaf rust fungus were unique. This database will be invaluable for the identification of the key genes that control the virulence of the pathogen or trigger the resistance response of the host.

Technical Abstract: Background: Rust fungi are biotrophic basidiomycete plant pathogens that cause major diseases on plants and trees world-wide, affecting agriculture and forestry. Their biotrophic nature precludes many established molecular genetic manipulations and lines of research. The generation of genomic resources for these microbes is leading to novel insights into their biology such as interactions with their hosts and guiding directions for break-through research in plant pathology. Results: In anticipation of and to support gene discovery and gene model verification in the wheat leaf rust fungus, Puccinia triticina (Pt), genome, we have generated Expressed Sequence Tags (ESTs) by sampling several life cycle stages. In this study we focused on several spore stages and isolated haustorial structures from infected wheat, generating 17,684 ESTs. We produced sequences from both the sexual (pycniospores, aeciospores and teliospores) and asexual (germinated urediniospores) stages of the life cycle. From pycniospores and aeciospores, produced by infecting the alternate host, meadow rue (Thalictrum speciosissimum), 4,869 and 1,292 reads, respectively, were generated. From teliospores produced on the senescent primary wheat host, we generated 3,703 ESTs. In addition, haustoria isolated from infected wheat yielded 6,817 reads and we generated 1,003 sequences from germinated urediniospores. These sequences were combined with a previous set of 25,558 ESTs into a database of 13,328 non-redundant sequences (4,506 singlets and 8,822 contigs). Fungal genes were predicted using the EST version of the GeneMarkS algorithm trained on wheat stem rust fungus, P. graminis f. sp. tritici (Pgt), predicted genes. To sanitize the EST database further, we compared sequences by BLASTN to a preliminary set of 454 pyrosequencing-generated contigs and Sanger BAC-end sequences derived from the Pt genome and the wheat genome. A collection of 6,308 most likely fungal genes was established which was compared to genomic resources of related cereal rusts Pgt and stripe rust P. striiformis f. sp. tritici (Pst), poplar leaf rust Melampsora species, and the corn smut fungus, Ustilago maydis (Um). Extensive homologies were found but significant sets of genes seemed species-specific and over 40% did not match any known sequences in various databases. Focusing on spore stages, direct comparison to Um identified potential functional homologs, possibly allowing heterologous functional analysis in that model fungus. Many potentially secreted protein genes were identified using comparative searches against predicted sets in Pgt and Melampsora spp., many belonging to groups of apparent orthologs. Conclusions: The current set of Pt unigenes contributes to gene discovery in this major cereal pathogen and will be invaluable for gene model verification in the anticipated genome sequence.