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Gene expression profiling and silencing reveal that monolignol biosynthesis plays a critical role in penetration defence in wheat against powdery mildew invasion.

Bhuiyan NH, Selvaraj G, Wei Y, King J - J. Exp. Bot. (2008)

Bottom Line: Co-silencing led to greater penetration of Bgt or Bgh than when the genes were silenced separately.Fluorescence emission spectra analyses revealed that gene silencing hampered host autofluorescence response at fungal contact sites.These results illustrate that monolignol biosynthesis is critically important for host defence against both appropriate and inappropriate pathogen invasion in wheat.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Saskatchewan, Saskatoon, SK S7N 5E2, Canada. nazmul.bhuiyan@yahoo.com

ABSTRACT
Cell wall apposition (CWA) formation is one of the first lines of defence used by plants to halt invading fungi such as powdery mildew. Lignin is a complex polymer of hydroxylated and methoxylated phenylpropane units (monolignols) and lignification renders the cell wall more resistant to pathogen attack. The role of monolignol biosynthesis in CWA-mediated defence against powdery mildew penetration into cereals is demonstrated here using RNA interference (RNAi)-mediated gene silencing and enzyme-specific inhibitors. Thirteen cDNAs representing eight genes involved in monolignol biosynthesis were cloned from an expression sequence tag (EST) library derived from the epidermis of diploid wheat (Triticum monococcum) infected with Blumeria graminis f. sp. tritici (Bgt). Differential expression patterns were found for these genes in susceptible and resistant plants after infection. Transcripts of phenylalanine ammonia lyase (PAL), caffeic acid O-methyltransferase (CAOMT), ferulic acid hydroxylase (FAH), caffeoyl-CoA O-methyltransferase (CCoAMT), and cinnamyl alcohol dehydrogenase (CAD) were accumulated, particularly in the epidermis. RNAi-mediated transient gene silencing in the epidermis led to a higher penetration efficiency of Bgt than in the controls. Gene silencing also compromised penetration resistance to varying degrees with different genes against an inappropriate pathogen, B. graminis f. sp. hordei (Bgh). Co-silencing led to greater penetration of Bgt or Bgh than when the genes were silenced separately. Fluorescence emission spectra analyses revealed that gene silencing hampered host autofluorescence response at fungal contact sites. These results illustrate that monolignol biosynthesis is critically important for host defence against both appropriate and inappropriate pathogen invasion in wheat.

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Effects of PAL and CAOMT inhibitors on the penetration ability of Bgt (A, B) and Bgh (C, D). (A, C) PAL inhibitor; (B, D) CAOMT inhibitor; (E) autofluorescence and (F) callose deposition in response to fungal interaction in CAOMT inhibitor-treated (right panel) and untreated (left panel) sample leaves. Con, conidia; AGT, appressorial germ tube. Scale bar corresponds to 10 μm. Wheat leaves were exposed to various concentrations of PAL and CAOMT inhibitors for 6 h prior to inoculation with Bgh. After inoculation, leaves were again placed on inhibitor solutions for 36 h. For observation of autofluorescence and callose deposition, leaves were collected at 20 hpi. Penetration efficiency was scored as described in the Materials and methods. Data shown represent mean ±standard deviation from three independent experiments in which 100 interaction sites were evaluated. Asterisks besides the columns indicate P <0.05 (Student's t test) compared with the negative control (0 mM inhibitor). (This figure is available in colour at JXB online.)
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fig7: Effects of PAL and CAOMT inhibitors on the penetration ability of Bgt (A, B) and Bgh (C, D). (A, C) PAL inhibitor; (B, D) CAOMT inhibitor; (E) autofluorescence and (F) callose deposition in response to fungal interaction in CAOMT inhibitor-treated (right panel) and untreated (left panel) sample leaves. Con, conidia; AGT, appressorial germ tube. Scale bar corresponds to 10 μm. Wheat leaves were exposed to various concentrations of PAL and CAOMT inhibitors for 6 h prior to inoculation with Bgh. After inoculation, leaves were again placed on inhibitor solutions for 36 h. For observation of autofluorescence and callose deposition, leaves were collected at 20 hpi. Penetration efficiency was scored as described in the Materials and methods. Data shown represent mean ±standard deviation from three independent experiments in which 100 interaction sites were evaluated. Asterisks besides the columns indicate P <0.05 (Student's t test) compared with the negative control (0 mM inhibitor). (This figure is available in colour at JXB online.)

Mentions: PAL is the first enzyme in the phenylpropanoid and monolignol biosynthesis pathways. Previous studies by others showed that the PAL-specific inhibitors, α-aminooxy-β-phenylpropaonic acid and 2-aminoindan-2-phosphonic acid, at a concentration of 1 mM, led to a reduction in papilla autofluorescence and, at the same time, increased the penetration rate of powdery mildew fungi into barley, oat, and wheat (Carver et al., 1992; Zeyen et al., 2002). Another PAL-specific inhibitor, 4-hydroxybenzoic hydrazide (HBH) (Sigma), was used here and its effect on the penetration efficiency of powdery mildew into wheat leaves was followed. The penetration efficiency of Bgt increased at all concentrations of inhibitor up to 1 mM, the highest being from 44% in the control to 60.3% at 0.5 mM inhibitor concentration (Fig. 7A). At 2 mM, the efficiency of penetration decreased, suggesting that this concentration of inhibitor was toxic to the plant.


Gene expression profiling and silencing reveal that monolignol biosynthesis plays a critical role in penetration defence in wheat against powdery mildew invasion.

Bhuiyan NH, Selvaraj G, Wei Y, King J - J. Exp. Bot. (2008)

Effects of PAL and CAOMT inhibitors on the penetration ability of Bgt (A, B) and Bgh (C, D). (A, C) PAL inhibitor; (B, D) CAOMT inhibitor; (E) autofluorescence and (F) callose deposition in response to fungal interaction in CAOMT inhibitor-treated (right panel) and untreated (left panel) sample leaves. Con, conidia; AGT, appressorial germ tube. Scale bar corresponds to 10 μm. Wheat leaves were exposed to various concentrations of PAL and CAOMT inhibitors for 6 h prior to inoculation with Bgh. After inoculation, leaves were again placed on inhibitor solutions for 36 h. For observation of autofluorescence and callose deposition, leaves were collected at 20 hpi. Penetration efficiency was scored as described in the Materials and methods. Data shown represent mean ±standard deviation from three independent experiments in which 100 interaction sites were evaluated. Asterisks besides the columns indicate P <0.05 (Student's t test) compared with the negative control (0 mM inhibitor). (This figure is available in colour at JXB online.)
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2651457&req=5

fig7: Effects of PAL and CAOMT inhibitors on the penetration ability of Bgt (A, B) and Bgh (C, D). (A, C) PAL inhibitor; (B, D) CAOMT inhibitor; (E) autofluorescence and (F) callose deposition in response to fungal interaction in CAOMT inhibitor-treated (right panel) and untreated (left panel) sample leaves. Con, conidia; AGT, appressorial germ tube. Scale bar corresponds to 10 μm. Wheat leaves were exposed to various concentrations of PAL and CAOMT inhibitors for 6 h prior to inoculation with Bgh. After inoculation, leaves were again placed on inhibitor solutions for 36 h. For observation of autofluorescence and callose deposition, leaves were collected at 20 hpi. Penetration efficiency was scored as described in the Materials and methods. Data shown represent mean ±standard deviation from three independent experiments in which 100 interaction sites were evaluated. Asterisks besides the columns indicate P <0.05 (Student's t test) compared with the negative control (0 mM inhibitor). (This figure is available in colour at JXB online.)
Mentions: PAL is the first enzyme in the phenylpropanoid and monolignol biosynthesis pathways. Previous studies by others showed that the PAL-specific inhibitors, α-aminooxy-β-phenylpropaonic acid and 2-aminoindan-2-phosphonic acid, at a concentration of 1 mM, led to a reduction in papilla autofluorescence and, at the same time, increased the penetration rate of powdery mildew fungi into barley, oat, and wheat (Carver et al., 1992; Zeyen et al., 2002). Another PAL-specific inhibitor, 4-hydroxybenzoic hydrazide (HBH) (Sigma), was used here and its effect on the penetration efficiency of powdery mildew into wheat leaves was followed. The penetration efficiency of Bgt increased at all concentrations of inhibitor up to 1 mM, the highest being from 44% in the control to 60.3% at 0.5 mM inhibitor concentration (Fig. 7A). At 2 mM, the efficiency of penetration decreased, suggesting that this concentration of inhibitor was toxic to the plant.

Bottom Line: Co-silencing led to greater penetration of Bgt or Bgh than when the genes were silenced separately.Fluorescence emission spectra analyses revealed that gene silencing hampered host autofluorescence response at fungal contact sites.These results illustrate that monolignol biosynthesis is critically important for host defence against both appropriate and inappropriate pathogen invasion in wheat.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Saskatchewan, Saskatoon, SK S7N 5E2, Canada. nazmul.bhuiyan@yahoo.com

ABSTRACT
Cell wall apposition (CWA) formation is one of the first lines of defence used by plants to halt invading fungi such as powdery mildew. Lignin is a complex polymer of hydroxylated and methoxylated phenylpropane units (monolignols) and lignification renders the cell wall more resistant to pathogen attack. The role of monolignol biosynthesis in CWA-mediated defence against powdery mildew penetration into cereals is demonstrated here using RNA interference (RNAi)-mediated gene silencing and enzyme-specific inhibitors. Thirteen cDNAs representing eight genes involved in monolignol biosynthesis were cloned from an expression sequence tag (EST) library derived from the epidermis of diploid wheat (Triticum monococcum) infected with Blumeria graminis f. sp. tritici (Bgt). Differential expression patterns were found for these genes in susceptible and resistant plants after infection. Transcripts of phenylalanine ammonia lyase (PAL), caffeic acid O-methyltransferase (CAOMT), ferulic acid hydroxylase (FAH), caffeoyl-CoA O-methyltransferase (CCoAMT), and cinnamyl alcohol dehydrogenase (CAD) were accumulated, particularly in the epidermis. RNAi-mediated transient gene silencing in the epidermis led to a higher penetration efficiency of Bgt than in the controls. Gene silencing also compromised penetration resistance to varying degrees with different genes against an inappropriate pathogen, B. graminis f. sp. hordei (Bgh). Co-silencing led to greater penetration of Bgt or Bgh than when the genes were silenced separately. Fluorescence emission spectra analyses revealed that gene silencing hampered host autofluorescence response at fungal contact sites. These results illustrate that monolignol biosynthesis is critically important for host defence against both appropriate and inappropriate pathogen invasion in wheat.

Show MeSH
Related in: MedlinePlus