No abstract

The constitutively produced polygalacturonase isoenzyme PG2 was isolated from culture filtrates of Botrytis cinerea, purified to homogeneity, and characterized. The shape of titration curves of PG2 and two other polygalacturonase isoenzymes explained the difficulties found in separating superimposed pectic enzyme activities during the purification procedure. PG2 hydrolyzed sodium polygalacturonate more quickly than pectin. The optimal pH for PG2 activity with polygalacturonate was 4.5 and with pectin, 4.0. PG2 activity was also influenced by the presence of NaCl or CaCl2 in the reaction mixture. Analysis of the breakdown products by paper chromatography and a comparison of the reaction rate by viscosimetry and reducing group assay revealed that PG2 has an endocatalytic mode of action on polygalacturonate. The isoelectric point and the molecular mass of PG2 were estimated to be 9.1 and 23.0 kDa, respectively. Key words: Botrytis cinerea, chromatofocusing, endopolygalacturonase, purification, substrate specificity, titration curve.

As part of an investigation of the cell wall structure of plant pathogenic, filamentous fungi, we set out to characterize covalently bound cell wall glycoproteins (CWPs) of the tomato pathogen Fusarium oxysporum. N-terminal sequencing of an abundant 60-kDa CWP led to the cloning of the corresponding gene, which we have designated FEM1 (Fusarium extracellular matrix protein). The gene contains an ORF encoding a primary translation product of 212 amino acids, including an N-terminal 17-amino acid secretion signal sequence. Furthermore, FEM1p contains two potential N-glycosylation sites, and is rich in serine and threonine residues (29%) that could serve as O-glycosyl addition sites. At its C-terminus the protein contains a 22-amino acid sequence with the characteristics of a glycosyl-phosphatidylinositol (GPI) anchor addition signal. A mutant FEM1 protein lacking this GPI anchor addition signal is not retained in the fungal cell wall but released into the culture medium, indicating that in the wild-type protein this sequence functions to anchor the protein to the extracellular matrix. Southern analysis shows that FEM1 is present as a single-copy gene in all formae speciales of F. oxysporum tested and in F. solani. Database searches show that FEM1p homologous sequences are present in other filamentous fungi as well.

Fungal polygalacturonase activity reflects susceptibility of carnation cultivars to fusarium wilt Baayen, R.P.; Schoffelmeer, E.A.M.; Toet, S.; Elgersma, D.M. General rightsIt is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s), other than for strictly personal, individual use, unless the work is under an open content license (like Creative Commons). Disclaimer/Complaints regulationsIf you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please let the Library know, stating your reasons. In case of a legitimate complaint, the Library will make the material inaccessible and/or remove it from the website. Please Ask the Library: http://uba.uva.nl/en/contact, or a letter to: Library of the University of Amsterdam, Secretariat, Singel 425, 1012 WP Amsterdam, The Netherlands. You will be contacted as soon as possible. Key words: breeding, Dianthus caryophyllus, Fusarium oxysporum, pectic enzymes, phytoalexins, resistance AbstractCarnation cultivars with different levels of partial resistance were inoculated with race 2 of Fusarium oxysporum f.sp. dianthi and monitored for accumulation of host phytoalexins, fungal escape from compartmentalization, production of fungal pectin-degrading enzymes and development of external disease symptoms. Accumulation of phytoalexins, assessed after 10 days in the first 5 cm above the inoculation site, was weakly (methoxydianthramide S) or not (hydroxydianthalexin B) correlated with resistance levels after 12 weeks. Fungal escape from compartmentalization, assessed after 3 weeks as percentages colonized plants at 8 cm above the inoculation site, was highly correlated with expression of susceptibility after 12 weeks. Polygalacturonase (PG) activity, assessed after 4 weeks in the first 5 cm above the inoculation site, was highly correlated to final disease development. Linear increases in disease severity were accompanied by quadratic increases in PG activity. In contrast to water-treated plants, that lacked any PG activity, inoculated plants contained two main groups of fungal PGs, the dominant forms of which had estimated pI values of 7.0 and minimally 9.5, respectively. Compared to those of the first group, enzymes of the second group were produced only in trace amounts in liquid media containing pectin or polygalacturonate as sole source of carbon. On these media, the fungus also produced a pectin methyl esterase (PME) with an estimated pI of 9.3. Besides PMEs of host origin, inoculated plants of susceptible cultivars contained the fungal PME while no more than traces were found in resistant ones.Assessment of phytoalexin production by the host during defense responses cannot replace monitoring of external symptoms as a resistance test. Assessment of fungal growth, whether by reisolations above the compartmentalization area or by measurement of PG activity, provides a both rapid and reliable prediction of disease development.

No abstract