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Mediated plastid RNA editing in plant immunity.

García-Andrade J, Ramírez V, López A, Vera P - PLoS Pathog. (2013)

Bottom Line: Furthermore, we observed that following a pathogenic challenge, wild type plants respond with editing inhibition of ndhB transcript.In parallel, rapid destabilization of the plastidial NDH complex is also observed in the plant following perception of a pathogenic cue.Therefore, NDH complex activity and plant immunity appear as interlinked processes.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Biología Molecular y Celular de Plantas, Universidad Politécnica de Valencia-C.S.I.C, Ciudad Politécnica de la Innovación, Ingeniero Fausto Elio, Valencia, Spain.

ABSTRACT
Plant regulatory circuits coordinating nuclear and plastid gene expression have evolved in response to external stimuli. RNA editing is one of such control mechanisms. We determined the Arabidopsis nuclear-encoded homeodomain-containing protein OCP3 is incorporated into the chloroplast, and contributes to control over the extent of ndhB transcript editing. ndhB encodes the B subunit of the chloroplast NADH dehydrogenase-like complex (NDH) involved in cyclic electron flow (CEF) around photosystem I. In ocp3 mutant strains, ndhB editing efficiency decays, CEF is impaired and disease resistance to fungal pathogens substantially enhanced, a process recapitulated in plants defective in editing plastid RNAs encoding NDH complex subunits due to mutations in previously described nuclear-encoded pentatricopeptide-related proteins (i.e. CRR21, CRR2). Furthermore, we observed that following a pathogenic challenge, wild type plants respond with editing inhibition of ndhB transcript. In parallel, rapid destabilization of the plastidial NDH complex is also observed in the plant following perception of a pathogenic cue. Therefore, NDH complex activity and plant immunity appear as interlinked processes.

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Comparative immune responses of plastid PPR-related mutants to inoculation with P. cucumerina.(A) crr2, crr21, ppra, and pprb disease resistance responses to P. cucumerina compared with ocp3 and wild-type (Col-0) plants. Lesion diameter of 20 plants per genotype and four leaves per plant were determined 12 d following inoculation with P. cucumerina. Values are means and ± SE (n = 80). ANOVA detected significant differences at the P<0.05 level. Experiments were repeated three times with similar results. (B) Representative leaves from each genotype at 12 days following inoculation with P. cucumerina. Bar represents 5 mm. (C) Aniline blue staining and epifluorescence microscopy was applied to visualize callose accumulation. Micrographs indicate P. cucumerina inoculation and infection site in the different Arabidopsis genotypes at 0 h.p.i (right panel) and at 48 h.p.i. (left panel). Bar represents 500 µm. (D) The number of yellows pixels (corresponding to pathogen-induced callose) per million on digital photographs of infected leaves were used as a means to express arbitrary units (i.e. to quantify the image) at 48 h.p.i. Data are visible microscopy averages from Col-0 and mutant plants (±SE). Different letters above bars indicate statistically significant differences between genotypes, according to one-way ANOVA (P<0.05, n = 15).
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ppat-1003713-g006: Comparative immune responses of plastid PPR-related mutants to inoculation with P. cucumerina.(A) crr2, crr21, ppra, and pprb disease resistance responses to P. cucumerina compared with ocp3 and wild-type (Col-0) plants. Lesion diameter of 20 plants per genotype and four leaves per plant were determined 12 d following inoculation with P. cucumerina. Values are means and ± SE (n = 80). ANOVA detected significant differences at the P<0.05 level. Experiments were repeated three times with similar results. (B) Representative leaves from each genotype at 12 days following inoculation with P. cucumerina. Bar represents 5 mm. (C) Aniline blue staining and epifluorescence microscopy was applied to visualize callose accumulation. Micrographs indicate P. cucumerina inoculation and infection site in the different Arabidopsis genotypes at 0 h.p.i (right panel) and at 48 h.p.i. (left panel). Bar represents 500 µm. (D) The number of yellows pixels (corresponding to pathogen-induced callose) per million on digital photographs of infected leaves were used as a means to express arbitrary units (i.e. to quantify the image) at 48 h.p.i. Data are visible microscopy averages from Col-0 and mutant plants (±SE). Different letters above bars indicate statistically significant differences between genotypes, according to one-way ANOVA (P<0.05, n = 15).

Mentions: We hypothesized that via NDH complex inhibition, plants could develop an alerted immune status. This might explain why ocp3 plants exhibited enhanced disease resistance to fungal pathogens resulting from earlier and more intense callose synthesis and deposition following pathogen exposure [25], [26]. If so, then mutants showing similar chloroplast NDH complex defects would activate the same immune status, and become resistant to fungal attack. Consequently, we challenged crr21 and crr2 mutants with P. cucumerina, and studied disease susceptibility in comparison to the resistant ocp3 plants, and the susceptible Col-0 plants. CRR2 is a distinct PPR protein that functions in the intergenic RNA cleavage between rps7 and ndhB, which is essential for subunit B translation, and crr2 mutants are compromised in NDH activity [35]. ppra, a previously uncharacterized T-DNA mutant, defective in the expression of PPRa (Figure S8) encoding a PPR protein of unknown function that is highly co-expressed with CRR21 and OCP3 (Figure 3A), was also evaluated. Similarly, pprb, a T-DNA mutant defective in another co-expressed PPR of unknown function (Figure S7) was included in these experiments for comparison. Following inoculation with P. cucumerina, disease was scored 12 d after inoculation by following necrosis and chlorosis extent present in inoculated leaves. As expected, Col-0 plants were highly susceptible to P. cucumerina, and all inoculated plants showed extended necrosis accompanied by extensive proliferation of fungal mycelia (Figure 6A–B). The same disease susceptibility was observed in the pprb mutant, indicating this PPR gene is not essential in plant's defense activation (Figure 6A–B). In marked contrast, the inoculated crr21, crr2, and ppra plants responded with a substantial increase in disease resistance to P. cucumerina infection that was of a magnitude similar to that attained in ocp3 plants (Figure 6A–B). Comparative cytological observations were performed at the sites of attempted fungal infection and the degree of induced callose deposition induction in inoculated leaves was monitored after staining with aniline blue, and examination by fluorescence microscopy. Results indicated none of the mutants exhibited aniline blue staining in control leaves (Figure 6C). Col-0 and pprb plants deposited callose locally at sites demarcating the zones of extended fungal growth. In marked contrast, crr21, crr2, ppra, and ocp3 plants all exhibited intensified and highly localized callose deposition in response to fungal infection, which occurred at zones where fungal growth and colonization was impeded (Figure 6C–D). Consequently, heightened disease resistance, and increased callose deposition were concurring traits in mutants defective in the correct editing of RNAs encoding subunits of the chloroplast NDH complex.


Mediated plastid RNA editing in plant immunity.

García-Andrade J, Ramírez V, López A, Vera P - PLoS Pathog. (2013)

Comparative immune responses of plastid PPR-related mutants to inoculation with P. cucumerina.(A) crr2, crr21, ppra, and pprb disease resistance responses to P. cucumerina compared with ocp3 and wild-type (Col-0) plants. Lesion diameter of 20 plants per genotype and four leaves per plant were determined 12 d following inoculation with P. cucumerina. Values are means and ± SE (n = 80). ANOVA detected significant differences at the P<0.05 level. Experiments were repeated three times with similar results. (B) Representative leaves from each genotype at 12 days following inoculation with P. cucumerina. Bar represents 5 mm. (C) Aniline blue staining and epifluorescence microscopy was applied to visualize callose accumulation. Micrographs indicate P. cucumerina inoculation and infection site in the different Arabidopsis genotypes at 0 h.p.i (right panel) and at 48 h.p.i. (left panel). Bar represents 500 µm. (D) The number of yellows pixels (corresponding to pathogen-induced callose) per million on digital photographs of infected leaves were used as a means to express arbitrary units (i.e. to quantify the image) at 48 h.p.i. Data are visible microscopy averages from Col-0 and mutant plants (±SE). Different letters above bars indicate statistically significant differences between genotypes, according to one-way ANOVA (P<0.05, n = 15).
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Related In: Results  -  Collection

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ppat-1003713-g006: Comparative immune responses of plastid PPR-related mutants to inoculation with P. cucumerina.(A) crr2, crr21, ppra, and pprb disease resistance responses to P. cucumerina compared with ocp3 and wild-type (Col-0) plants. Lesion diameter of 20 plants per genotype and four leaves per plant were determined 12 d following inoculation with P. cucumerina. Values are means and ± SE (n = 80). ANOVA detected significant differences at the P<0.05 level. Experiments were repeated three times with similar results. (B) Representative leaves from each genotype at 12 days following inoculation with P. cucumerina. Bar represents 5 mm. (C) Aniline blue staining and epifluorescence microscopy was applied to visualize callose accumulation. Micrographs indicate P. cucumerina inoculation and infection site in the different Arabidopsis genotypes at 0 h.p.i (right panel) and at 48 h.p.i. (left panel). Bar represents 500 µm. (D) The number of yellows pixels (corresponding to pathogen-induced callose) per million on digital photographs of infected leaves were used as a means to express arbitrary units (i.e. to quantify the image) at 48 h.p.i. Data are visible microscopy averages from Col-0 and mutant plants (±SE). Different letters above bars indicate statistically significant differences between genotypes, according to one-way ANOVA (P<0.05, n = 15).
Mentions: We hypothesized that via NDH complex inhibition, plants could develop an alerted immune status. This might explain why ocp3 plants exhibited enhanced disease resistance to fungal pathogens resulting from earlier and more intense callose synthesis and deposition following pathogen exposure [25], [26]. If so, then mutants showing similar chloroplast NDH complex defects would activate the same immune status, and become resistant to fungal attack. Consequently, we challenged crr21 and crr2 mutants with P. cucumerina, and studied disease susceptibility in comparison to the resistant ocp3 plants, and the susceptible Col-0 plants. CRR2 is a distinct PPR protein that functions in the intergenic RNA cleavage between rps7 and ndhB, which is essential for subunit B translation, and crr2 mutants are compromised in NDH activity [35]. ppra, a previously uncharacterized T-DNA mutant, defective in the expression of PPRa (Figure S8) encoding a PPR protein of unknown function that is highly co-expressed with CRR21 and OCP3 (Figure 3A), was also evaluated. Similarly, pprb, a T-DNA mutant defective in another co-expressed PPR of unknown function (Figure S7) was included in these experiments for comparison. Following inoculation with P. cucumerina, disease was scored 12 d after inoculation by following necrosis and chlorosis extent present in inoculated leaves. As expected, Col-0 plants were highly susceptible to P. cucumerina, and all inoculated plants showed extended necrosis accompanied by extensive proliferation of fungal mycelia (Figure 6A–B). The same disease susceptibility was observed in the pprb mutant, indicating this PPR gene is not essential in plant's defense activation (Figure 6A–B). In marked contrast, the inoculated crr21, crr2, and ppra plants responded with a substantial increase in disease resistance to P. cucumerina infection that was of a magnitude similar to that attained in ocp3 plants (Figure 6A–B). Comparative cytological observations were performed at the sites of attempted fungal infection and the degree of induced callose deposition induction in inoculated leaves was monitored after staining with aniline blue, and examination by fluorescence microscopy. Results indicated none of the mutants exhibited aniline blue staining in control leaves (Figure 6C). Col-0 and pprb plants deposited callose locally at sites demarcating the zones of extended fungal growth. In marked contrast, crr21, crr2, ppra, and ocp3 plants all exhibited intensified and highly localized callose deposition in response to fungal infection, which occurred at zones where fungal growth and colonization was impeded (Figure 6C–D). Consequently, heightened disease resistance, and increased callose deposition were concurring traits in mutants defective in the correct editing of RNAs encoding subunits of the chloroplast NDH complex.

Bottom Line: Furthermore, we observed that following a pathogenic challenge, wild type plants respond with editing inhibition of ndhB transcript.In parallel, rapid destabilization of the plastidial NDH complex is also observed in the plant following perception of a pathogenic cue.Therefore, NDH complex activity and plant immunity appear as interlinked processes.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Biología Molecular y Celular de Plantas, Universidad Politécnica de Valencia-C.S.I.C, Ciudad Politécnica de la Innovación, Ingeniero Fausto Elio, Valencia, Spain.

ABSTRACT
Plant regulatory circuits coordinating nuclear and plastid gene expression have evolved in response to external stimuli. RNA editing is one of such control mechanisms. We determined the Arabidopsis nuclear-encoded homeodomain-containing protein OCP3 is incorporated into the chloroplast, and contributes to control over the extent of ndhB transcript editing. ndhB encodes the B subunit of the chloroplast NADH dehydrogenase-like complex (NDH) involved in cyclic electron flow (CEF) around photosystem I. In ocp3 mutant strains, ndhB editing efficiency decays, CEF is impaired and disease resistance to fungal pathogens substantially enhanced, a process recapitulated in plants defective in editing plastid RNAs encoding NDH complex subunits due to mutations in previously described nuclear-encoded pentatricopeptide-related proteins (i.e. CRR21, CRR2). Furthermore, we observed that following a pathogenic challenge, wild type plants respond with editing inhibition of ndhB transcript. In parallel, rapid destabilization of the plastidial NDH complex is also observed in the plant following perception of a pathogenic cue. Therefore, NDH complex activity and plant immunity appear as interlinked processes.

Show MeSH
Related in: MedlinePlus