Differential in vitro and in vivo effect of barley cysteine and serine protease inhibitors on phytopathogenic microorganisms

Abstract

Keywords:Cysteine protease inhibitor Serine protease inhibitor Barley Plant pathogen Transgenic plant Protease inhibitors from plants have been involved in defence mechanisms against pests and pathogens. Phytocystatins and trypsin/a-amylase inhibitors are two of the best characterized protease inhibitor families in plants. In barley, thirteen cystatins (HvCPI-1 to 13) and the BTI-CMe trypsin inhibitor have been previously studied. Their capacity to inhibit pest digestive proteases, and the negative in vivo effect caused by plants expressing these inhibitors on pests support the defence function of these proteins. Barley cystatins are also able to inhibit in vitro fungal growth. However, the antifungal effect of these inhibitors in vivo had not been previously tested. Moreover, their in vitro and in vivo effect on plant pathogenous bacteria is still unknown. In order to obtain new insights on this feature, in vitro assays were made against different bacterial and fungal pathogens of plants using the trypsin inhibitor BTI-CMe and the thirteen barley cystatins. Most barley cystatins and the BTI-CMe inhibitor were able to inhibit mycelial growth but no bacterial growth. Transgenic Arabidopsis plants independently expressing the BTI-CMe inhibitor and the cystatin HvCPI-6 were tested against the same bacterial and fungal pathogens. Neither the HvCPI-6 expressing transgenic plants nor the BTI-CMe ones were more resistant to plant pathogen fungi and bacteria than control Arabidopsis plants. The differences observed between the in vitro and in planta assays against phytopathogenic fungi are discussed.

Characterization of a cDNA encoding cystatin with antifungal activity from Siam tulip Curcuma alismatifolia

“…The growth of P. grisea was inhibited up to 90% in the presence at 5 µM CaCPI in the culture medium but others needed more CaCPI concentration for inhibit at the same level. Our results agreed with a previous report on barley cystatins 40 . In vitro, most barley cystatins provided a greater growth inhibition in P. grisea than F. oxysporum or Plectosphaerella cucumerina.…”

“…It is possible that cystatin was targeted to another type of proteolytic enzymes or that growth inhibition was not mediated by inhibition of cystein proteinases. However, the mechanisms responsible for these differences are still unknown 40 . When a sensitivity of fungi to CaCPI was compared between C. capsici and F. oxysporum.…”

“…In the case of P. grisea, there was no report on the maximum cystatin concentration for growth inhibition but there was some reports on barley cystatins (HvCPI-1 to HvCPI-12). The 50% growth inhibition (EC 50 ) against P. grisea was 0.18-5.08 µM 40 . This concentration was also similar to our result (Fig.…”

Castor bean cake contains a trypsin inhibitor that displays antifungal activity against Colletotrichum gloeosporioides and inhibits the midgut proteases of the dengue mosquito larvae

“…At 5 g mL −1 concentration (0.31 M), the trypsin inhibitor from Helianthus annuus flower completely inhibited the Sclerotinia sclerotiorum ascospore germination (Giudici et al, 2000). Moreover, the barley trypsin inhibitor (BTI-CMe) inhibited the spore germination of the fungi Magnaporthe grisea, Plectosphaerella cucumerina and F. oxysporum at 0.5, 1.0, and 1.5 M concentrations, respectively (Carrillo et al, 2011). In general, the effects of protease inhibitors against phytopathogenic fungi are associated with inhibition of fungus proteases needed for growth and differentiation.…”

“…In plant tissues, protease inhibitors, besides being considered storage proteins (Mandal et al, 2002) are associated with regulation and control of endogenous proteases during seed development and germination (Xavier-Filho and Campos, 1889;Richardson, 1991), regulation of programmed cell death (Solomon et al, 1999), response to abiotic stresses (Franco and Melo, 2000) and protection against pathogens and insects (Ryan, 2000;Carrillo et al, 2011).…”