Cell wall is a dynamic structure that plays crucial roles in: maintaining fungal cell morphology, protecting mycelia from environmental stresses, and allowing interactions with substrates and living organisms. Starting from the availability of the genome sequence of Tuber melanosporum Vitt., an ectomycorrhizal (ECM) fungus, we performed an in silico analysis focusing on cell-wall related genes. The results offered a first glimpse on cell wall-related and secreted proteins, allowing the identification of several members of wall-associated genes (i.e., chitin synthases, chitinases, hydrophobins). As a second step, microarray data, available from the T.melanosporum genome (Martin et al. 2010), were validated selecting the symbiotic stage as the main focus. In detail, we performed qRT-PCR for 27 genes in order to verify their expression during the ECM formation versus the free living mycelium (FLM). The results demonstrated that genes coding for fungal cell wall degrading enzymes (i.e., some chitinases and glucanases) are up-regulated during the symbiosis, suggesting that these enzymes may control cell wall modifications required for the transition from the linear growth of the mycelium to the branched hyphae of the ECMs. Interestingly, also a gene encoding a protein involved in beta-1,6-glucans synthesis resulted to be up-regulated in qRT-PCR, suggesting that the synthesis of polysaccharides may also has a role during the interaction with plants. Lastly, a laser microdissection protocol was set up to verify whether cell wall related genes were differentially expressed in the mantle and in the Hartig net of the ECMs.
Genome-wide analysis of Tuber melanosporum secretome and cell wall genes: their implication in symbiosis
SILLO, FABIANO;BONFANTE, Paola
2010-01-01
Abstract
Cell wall is a dynamic structure that plays crucial roles in: maintaining fungal cell morphology, protecting mycelia from environmental stresses, and allowing interactions with substrates and living organisms. Starting from the availability of the genome sequence of Tuber melanosporum Vitt., an ectomycorrhizal (ECM) fungus, we performed an in silico analysis focusing on cell-wall related genes. The results offered a first glimpse on cell wall-related and secreted proteins, allowing the identification of several members of wall-associated genes (i.e., chitin synthases, chitinases, hydrophobins). As a second step, microarray data, available from the T.melanosporum genome (Martin et al. 2010), were validated selecting the symbiotic stage as the main focus. In detail, we performed qRT-PCR for 27 genes in order to verify their expression during the ECM formation versus the free living mycelium (FLM). The results demonstrated that genes coding for fungal cell wall degrading enzymes (i.e., some chitinases and glucanases) are up-regulated during the symbiosis, suggesting that these enzymes may control cell wall modifications required for the transition from the linear growth of the mycelium to the branched hyphae of the ECMs. Interestingly, also a gene encoding a protein involved in beta-1,6-glucans synthesis resulted to be up-regulated in qRT-PCR, suggesting that the synthesis of polysaccharides may also has a role during the interaction with plants. Lastly, a laser microdissection protocol was set up to verify whether cell wall related genes were differentially expressed in the mantle and in the Hartig net of the ECMs.
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