Activity of endo-polygalacturonases in mirid bugs (Heteroptera: Miridae) and their inhibition by plant cell wall proteins (PGIPs)

Endo-polygalacturonases (PGs) are hydrolytic enzymes involved in the degradation of pectin, one of the major components of plant cell wall. While PGs from fungi, bacteria and plants have been extensively studied, PGs from insects are much less known, although they are likely to play an important role in insect-plant interactions. Presence of PGs has been reported for both piercing-sucking and chewing insect species, and possibly more commonly in mirid bugs (Heteroptera: Miridae). A screening of some common mirid species and other insects, belonging to different orders and families, was conducted using agarose diffusion assays run at different pHs. All the mirid species tested [Lygus rugulipennis Popp., L. pratensis (L.), Orthops kalmi (L.), Adelphocoris lineolatus (Goeze) and Closterotomus norwegicus (Gmelin)] showed PG activity, mainly at pH 7–8, whereas no activity was recorded for the other insect species, except Sitophilus sp. (Coleoptera: Curculionidae). PG activity in females of L. pratensis was significantly higher than in males, whereas there were no differences between the sexes in the other species. In all these species, PGs were present both in the salivary glands and the gut, with a higher activity in the salivary glands, confirming the role of these enzymes in the feeding behaviour of mirid bugs. Inhibition of mirid PGs by polygalacturonase-inhibiting proteins (PGIPs) from different plant sources was analysed at pH 7. PGIPs are extracellular plant proteins known for their ability to inhibit fungal PGs and restrict fungal colonization. Two PGIPs from Phaseolus vulgaris (PvPGIP3 and PvPGIP4) inhibited PGs of all the mirid bugs tested. This information may be helpful for the development of innovative insect-resistant plant varieties, for use in low-impact IPM. 515 * Corresponding author; e-mail: [email protected] polygalacturonase-inhibiting proteins (PGIPs), which prevent fungal invasion and growth in plants (reviewed by De Lorenzo et al., 2001; De Lorenzo & Ferrari, 2002; Bellincampi et al., 2004). PGIPs are ineffective against bacterial PGs and pectic enzymes of plant origin (De Lorenzo & Ferrari, 2002). PGIPs are cell wall glycoproteins widespread in flowering plants with great potential for developing of plant resistance to fungi (De Lorenzo et al., 2001). Upon attack by fungal hypha, the control of PG activity by PGIPs favours the accumulation of oligalacturonides (OG) able to elicit defence responses such as phytoalexin accumulation, induction of phenylalanine ammonia lyase (PAL), glucanase, chitinase and lignification (Ridley at al., 2001). OGs may be released not only by exogenous PGs during pathogen infection, but also by endogenous PGs, systemically induced after wounding (De Lorenzo et al., 2001; De Lorenzo & Ferrari, 2002). Recent in vitro experiments using both a chewing insect, Diaprepes abbreviatus L. (Coleoptera: Curculionidae) (Doostdar et al., 1997) and the piercing-sucking species, Lygus spp. and Adelphocoris lineolatus (Goeze) (Heteroptera: Miridae) (Conti et al., 2004; D’Ovidio et al., 2004; Shackel et al., 2005), have shown that PGIPs also inhibit the PGs of insects. In this paper, through an initial investigation on the presence of PGs in several insect pest species of considerable economic importance, belonging to different orders, it is shown that these enzymes are present in both sexes of all the five mirid bugs tested, in a weevil but not in some other species. In mirid bugs, PGs are more abundant in the salivary glands than guts, which confirms the results of previous investigations on other species. Finally, evidence is provided that mirid PGs are inhibited by PGIPs from bean. The significance of these results for the development of innovative insect-resistant plant varieties is