Characterization Of Chitin Metabolism Related Genes In Puccinia Striiformis F.Sp. Tritici And Sequence Analysis Of Fungal Polysaccharide Deacetylase Genes

The coordinated synthesis and modification of chitin, a key component of fungal cell wall, is essential for plant fungal pathogens to maintain cell structure integrity and full pathogenicity. Researches have indicated that fungal pathogens could utilize CDA (Chitin Deacetylase) to evade host recognization and downstream defense responses by modifying the chitin component exposed at the cell wall surface during the infection process. A typical characteristic of CDA is a signal peptide at the N-terminal of the protein and a PDA domain (Polysaccharide Deacetylase domain) inside the protein. Several fungal species pocess many copies of PDA proteins, yet it is unclear how they have evolved and whether they are all CDAs or other functional proteins.To understand the role of chitin metabolism in the differentiation and infection processes of Puccinia striiformis f. sp. tritici and the evolutionary characteristics of fungal PDAs (Polysaccharide Deacetylase), a class II Chs (Chitin Synthase) gene PstChsII and several PstPDA genes were isolated and characterized from wheat stripe rust;two PstPDAs were also expressed in E.coli and purified;the sequences of 90 fungal PDAs were compared and analysized. The main results are listed as follows:1. Cloned PstChsII and PstPDAs encoded proteins with domains and motifs in accordance with reported Chs and PDA proteins; PstChsII may function in the synthesis of cell wall chitin during the urediospore germination process considering its expressional up-regulation at this stage; different The expression of PstPDAs were differently regulated and thus may function at different differentiation or infection stages; Alternative splicing and dikaryotic nature of urediospore contributed to the exsistance of three different transcripts corresponding to one PstPDA gene - PstPDA1.2. 22 members of 90 fungal PDAs were re-annotated based on protein and intron splicing site characteristics. This re-annotation identified 11 signal peptides out of 18 PDAs originally annotated as non-secreting proteins.3. The evolutionary characteristics of 90 fungal PDAs were analyzed. 13 lineage specific duplication groups were identified in PDAs from Puccinia graminis, Coprinus cinera and Rhizopus oryzae, demonstrating the role of gene duplication in gene family expansion. Diverse changes at critical amino acid in conserved motifs were observed in members of the same duplication group in C. cinera and R. oryzae, implying a functional diversification of these duplicated genes. Fungal PDAs were classified into 3 subgroups based on their phylogenetic relationship. All subgroup III PDAs did not have complete catalytic-related motifs; Subgroup II did not have representative from ascomycete fungal species Botrvtis cinerea and Fusarium graminearum;PDAs from all basidiomycete fungal species in subgroup I showed amino acid changes at the tryptophan residue in"DXXDW"motif.4. 2 supercontig pairs possessing highly similar regions were identified from P. graminis genome when analyzing PDA genes in P. graminis. A synteny region of at least 80 kb was discovered between supercontig 34 and supercontig 100. Sequence characteristics indicating DNA segment duplication and insertion events were identified in the regions.