Characterization of a 20 kDa DNase elicitor from Fusarium solani f. sp. phaseoli and its expression at the onset of induced resistance in Pisum sativum

Abstract

Summary DNase released from Fusarium solani f. sp. phaseoli (Fsph DNase) has previously been reported to induce pathogenesis-related (PR) genes, phytoalexin accumulation and disease resistance against subsequent challenge with the true pea pathogen, Fusarium solani f. sp. pisi (Fspi). This report is a further analysis of DNase production with probes specific for both the gene and protein. N-terminal analysis of the approximately 20 kDa Fsph DNase protein facilitated both the development of anti-Fsph DNase antiserum and the cloning of the Fsph DNase gene. Utilizing the anti-Fsph DNase antiserum to prepare an affinity column, we demonstrated that the retention and recovery of the DNase activity was associated with this protein. Fsph DNase protein was detectable by Western analysis in both the fungi and plant cytoplasm within 6-8 h following inoculation of the pea endocarp surface. Partially purified DNase detected via catalytic activity began accumulating within pea tissue at 3 h post-inoculation. Enhanced fragmentation of pea DNA occurred within 5 h following treatment of pods with Fsph DNase or inoculations with the two fungi. DNA cleavage within the nuclei of endocarp pea cells was detectable via a TUNEL assay at 3 h post-inoculation. As a result of these findings, we propose that the entrance of Fsph DNase into the pea cell and the signalling of plant defence responses is temporally associated with the damage of host DNA.

Extracellular DNA Is Required for Root Tip Resistance to Fungal Infection

“…Like human pathogens that produce extracellular DNase activities now known to be involved in virulence (Sherry and Goeller, 1950;Sumby et al, 2005;Buchanan et al, 2006), rootassociated bacteria and fungi also produce extracellular DNase activities (Klosterman et al, 2001;Tavares and Sellstedt, 2001;Balestrazzi et al, 2007). Klosterman et al (2001) have proposed that such enzymes in pathogenic fungi function by entering the nuclei of plant cells and causing DNA damage that triggers defense responses in nonhost tissues. It will be of interest in future studies to examine the alternative hypothesis that, as with human pathogens, these enzymes play a role in plant cell exDNA degradation as part of the infection process.…”

Anatomy of a nonhost disease resistance response of pea to Fusarium solani: PR gene elicitation via DNase, chitosan and chromatin alterations

“…Enzymes such as cutinase (Woloshuk and Kollatukudy, 1986), and DNase (Klosterman et al, 2001) are released as the inoculum contacts pea tissue. Alternately, the pea tissue has constitutive levels of enzymes such as chitinase and β-glucanase (Mauch et al, 1984), each with N-terminal “SignalP” peptides that can enable their immediate transfer through membranes (Hadwiger, 2009).…”

“…Additionally, these patterns were dramatically different from those found in heat-shock and heavy metal treated endocarp tissue. The biotic gene-inducing agents deemed important in the pea/Fusarium interactions were the chitosan oligomers (Kendra et al, 1989) and the FsphDNase (Klosterman et al, 2001) discussed above.…”

Characterization of Human Antigenic Proteins SchS21 and SchS34 from <i>Stachybotrys chartarum</i>

“…DNases from the extracellular fraction of filamentous fungi also function in autolysis when fungal cells reach maximal dry weight [31]. Other functions of DNases of filamentous fungi, including that of N. haematococca, may also result in the stimulation of pathogenesis-related genes and phytoalexin accumulation in plants [20,29,32]. The DNase from N. haematococca shares 58% identity with SchS21.…”

“…These enzymes can be divided into 2 classes: endonucleasesthat hydrolyse the deoxyribose phosphodiester backbone within the DNA strand and exonucleasesthat hydrolyze the phosphodiester bonds at the DNA ends [26,27]. Differing in their substrate specificities, chemical mechanisms and biological functions, a wide variety of DNases are known [26,28,29]. An extracellular DNase (AAD53090.1) from N. haematococca was homologous and has similar properties to SchS21, such as a molecular weight of 20 kDa, isoelectric point of 4.2 and optimal temperature of 45°C [5,20].…”