Figure 1.
Traps of Dionaea muscipula in response to mechanical stimulation.
An open trap (A), the same trap 1 min after double mechanical stimulation of trigger hairs (B) and 3 hours later after repeated mechanical stimulation in narrowed phase (C).
Figure 2.
Narrowing response after different treatments.
As a control wet (distilled H2O) and dry filter papers were applied. M – mechanical stimulation, Different letters denote significant differences at P < 0.05 (ANOVA, Tukey-test), means ± s.e., n = 5–9.
Figure 3.
Electrical activity measured by extracellular electrode on trap surface.
Mechanical irritation delivered in 10 minutes intervals (A), detailed view on action potentials (B) and chemical stimulation with NH4Cl and KH2PO4 did not trigger any AP (C).
Figure 4.
Enzymatic activities in response to mechanical and chemical stimulation after 24 (black bars) and 48 hours (white bars).
Acid phosphatase activity (A) and proteolytic activity (B). Different letters denote significant differences at the same time interval at P < 0.05 (ANOVA, Tukey-test), means ± s.e., n = 7–9.
Figure 5.
Inhibition of cysteine peptidase activity after 48 hours.
Relative proteolytic activity measured after inhibition with 10 µM cysteine proteinase inhibitor E-64. Activity without inhibitor of given variant is 100 %. Different letters denote significant differences at P < 0.05 (ANOVA, Tukey-test), means ± s.e., n = 6.
Figure 6.
Protein pattern, Western blot and zymogram in response to mechanical and chemical stimulation.
Silver-stained SDS-polyacrylamide gel containing proteins in D. muscipula digestive fluid after mechanical (M), P(K) and N(Cl) stimulation (A). The same amount of proteins was electrophoresed in 12% (v/v) SDS-polyacrylamide gel and subjected to Western blot analysis using antibodies against dionain-1 (B). Detection of protease activity in 12% (v/v) SDS-polyacrylamide gel with casein as a substrate (C). The clear bands against background indicate protease activity. Representative gels at least from 4 repetitions are shown.
Figure 7.
Endogenous concentration of phytohormones (JA, JA-Ile, IAA, ABA, cis-OPDA and SA) in trap tissue 36 - 48 hours after stimulation.
Note different scale on y-axis for cis-OPDA and SA. Control (C), mechanical (M), P(K) and N(Cl) stimulation. Different letters denote significant differences at P < 0.05 of particular phytohormone among treatments (ANOVA, Tukey test), means ± s.e., n = 9–10.
Figure 8.
Correlation between endogenous phytohormone level and proteolytic activity.
Correlation between endogenous concentration of JA (A), JA-Ile (B), IAA (C), ABA (D), cis-OPDA (E), SA (F) and proteolytic activity induced by mechanical and chemical stimulation. The linear relationship is significant only for JA (P = 0.048, significance test for linear regression), means ± s.e.
Figure 9.
Enzymatic activities in response to external JA application.
Acid phosphatase activities (A), proteolytic activities (B) and activities measured after inhibition with 10 µM cysteine proteinase inhibitor E-64. Activity without inhibitor of given variant is 100 % (C). ** - significant differences at P = 0.01 (Student's t-test), means ± s.e., n = 6–7.
Figure 10.
Protein pattern, Western blot and zymogram in response to external application of jasmonic acid.
Silver-stained SDS-polyacrylamide gel containing proteins in D. muscipula digestive fluid after mechanical stimulation (M) and external application of jasmonic acid (JA), (A). The same amount of proteins was electrophoresed in 12% (v/v) SDS-polyacrylamide gel and subjected to Western blot analysis using antibodies against dionain-1 (B). Detection of protease activity in 12% (v/v) SDS-polyacrylamide gel with casein as a substrate (C). The clear bands against background indicate protease activity. Representative gels at least from 4 repetitions are shown.
Figure 11.
Double trigger mechanism for protein digestion.
Three control points in prey capture and digestion of Venus flytrap, which ensure effective production of digestive enzymes. For detailed description, see discussion. AP – action potential, the increase of phytohormone level is indicated by arrows: no increase (-), small (↑), moderate (↑↑), high (↑↑↑).