Tomassini, Alessia (2008) Looking for a role of polygalacturonase of Fusaria during cereal infection. [Tesi di dottorato]
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To penetrate and colonize host tissue most pathogenic fungi produce enzymes degrading the cuticle and the plant cell wall. Among these, pectinases and in particular endo-polygalacturonases (PGs), are expressed in the early stages of host infection and contribute to the virulence or the pathogenicity of several phytopathogenic fungi by degrading the pectin component of the cell wall and middle lamella. Pectic enzymes are considered the main factors responsible for the maceration of the pectin rich cell wall of dicotyledonous plants. Recently, the importance of these enzymes has been shown also in the pathogenesis of some Graminaceous, although these plants have a cell wall consisting of small amount of pectin.
Fusarium graminearum and Fusarium verticillioides are two relevant pathogens of cereal species and are know to produce PG activity in liquid culture. The demonstration that the PGs of these fungi are virulence factors might contribute to develop strategies aimed to increase the resistance of host plants to infection by these pathogens. To clarify the importance of these enzymes during pathogenesis, firstly the two endo-PGs secreted in vitro by F. graminearum were purified and characterized. These PGs showed different biochemical properties, like optimum pH and substrate cleavage mechanism. The expression of their encoding genes was analysed also during wheat infection compared to the expression of pectin lyase and xylanase encoding genes: since pectinases genes were expressed earlier than xylanase gene, pectinases, and in particular PGs, might play a role during the early stage of infection.
To establish the importance in pathogenesis of the two purified and characterized F. graminearum PGs, and of F. verticillioides PG, previously characterized (Sella et al., 2004; Raiola et al., in press), their encoding genes were disrupted by targeted homologous recombination. The pathogenicity of each mutant was tested by inoculating host plants. Single F. graminearum mutants maintained the capability to infect wheat plants. However, since the loss of PG activity due to the knock-out of a single pg gene could be compensated by the activity of the remaining PG, a double knock-out mutant should be obtained and tested in infection experiments. Also the F. verticillioides mutant maintained the capability to infect maize plants, but in this case the pg gene disruption caused a reduction of virulence. In particular, the necrotic symptom observed during infection with the wild-type strain might be related to the presence of the PG in the infected tissue.
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