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Identification of a Spotted Leaf Sheath Gene Involved in Early Senescence and Defense Response in Rice

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Identification of a Spotted Leaf Sheath Gene Involved in Early Senescence and Defense Response in Rice

Dongryung Lee et al. Front Plant Sci.

Abstract

Lesion mimic mutants (LMMs) commonly exhibit spontaneous cell death similar to the hypersensitive defense response that occurs in plants in response to pathogen infection. Several lesion mimic mutants have been isolated and characterized, but their molecular mechanisms remain largely unknown. Here, a spotted leaf sheath (sles) mutant derived from japonica cultivar Koshihikari is described. The sles phenotype differed from that of other LMMs in that lesion mimic spots were observed on the leaf sheath rather than on leaves. The sles mutant displayed early senescence, as shown, by color loss in the mesophyll cells, a decrease in chlorophyll content, and upregulation of chlorophyll degradation-related and senescence-associated genes. ROS content was also elevated, corresponding to increased expression of genes encoding ROS-generating enzymes. Pathogenesis-related genes were also activated and showed improved resistance to pathogen infection on the leaf sheath. Genetic analysis revealed that the mutant phenotype was controlled by a single recessive nuclear gene. Genetic mapping and sequence analysis showed that a single nucleotide substitution in the sixth exon of LOC_Os07g25680 was responsible for the sles mutant phenotype and this was confirmed by T-DNA insertion line. Taken together, our results revealed that SLES was associated with the formation of lesion mimic spots on the leaf sheath resulting early senescence and defense responses. Further examination of SLES will facilitate a better understanding of the molecular mechanisms involved in ROS homeostasis and may also provide opportunities to improve pathogen resistance in rice.

Keywords: Mitogen-Activated Protein Kinase Kinase Kinase (MAPKKK); blast resistance; early senescence; leaf sheath; lesion mimic mutant (LMM); reactive oxygen species (ROS).

Figures

Figure 1
Figure 1
Morphological comparisons between wild-type and sles mutant plants. (A) Wild type (left) and sles mutant (right) at 60 day after germination. (B) 90-day-old and 120-day-old wild type (left) and sles mutant (right). (C) Root color in wild type (left) and sles mutant (right).
Figure 2
Figure 2
Light microscopic analysis of spotted and non-spotted leaf sheath from sles mutant and wild type plants. Transverse sections of penultimate leaf sheaths were observed under white light. (D–F) are magnified view of (A–C), respectively. (A,D) are wild type leaf sheath sections. (B,E) are non-spotted, and (C,F) are spotted leaf sheath sections from the sles mutant. Indications in (A) are ARC, aerenchyma; B, bundle sheath; P, phloem; PP, palisade parenchyma; SP, spongy parenchyma; X, xylem.
Figure 3
Figure 3
Early senescence in the sles mutant leaf sheath. (A) Abundance of major plant pigments in non-spotted (nsp) and spotted (sp) leaf sheaths from the sles mutant and wild-type (WT) leaf sheaths. (B) Expression of chlorophyll degradation-related genes. (C) Expression of senescence transcription factors. (D) Expression of senescence-associated genes. Real-time PCR (three biological replicates and three technical replicates) was performed with WT leaf sheath samples and sles leaf sheath samples from regions with legion mimic spots. Asterisks indicate the statistical significance levels according to Student's t-test: **P < 0.01 and *P < 0.05.
Figure 4
Figure 4
ROS accumulation in the sles mutant leaf sheath. (A) Trypan blue, NBT and DAB staining of penultimate leaves and leaf sheaths in wild-type and sles mutant plants after heading. (B) Expression of genes encoding ROS-generating enzymes in wild-type and sles mutant (C–E) Expression levels of ROS detoxification-related genes in wild-type and sles mutant. Real-time PCR (three biological replicates and three technical replicates) was performed with leaf sheath samples from wild-type and from areas with legion mimic spots in the sles mutant. Asterisks indicate the statistical significance level according to Student's t-test: **P < 0.01 and *P < 0.05.
Figure 5
Figure 5
Blast resistance in the sles mutant leaf sheath. (A) Expression of pathogenesis-related marker genes. Real-time PCR was performed with leaf sheath samples from wild-type and from areas with legion mimic spots in the sles mutant. Asterisks indicate the statistical significance level according to Student's t-test: **P < 0.01 and *P < 0.05. (B) The excised leaf sheath from 50-day-old rice seedlings of WT and sles mutant was inoculated with conidial suspension (1 × 104 conidia/ml). Samples were harvested and observed 48 h after inoculation. Three biological replicates and three technical replicates were performed for each experiment. Bar = 25 μm.
Figure 6
Figure 6
Fine-mapping and identification of SLES. (A) Fine-mapping of SLES. The sles locus was mapped to a 66 kb region on chromosome 7. (B) Schematic diagram of SLES. Black rectangles represent exons and the black inverted triangle represents the mutation site. (C) Expression of SLES gene on leaf and leaf sheath. (D–F) Phenotypic comparison of wild-type Dongjinbyeo (left) and the homozygous T-DNA insertion line (right). (D) Seedling. (E) Leaf sheath. (F) Root. Real-time PCR (three biological replicates and three technical replicates) was performed. Asterisks indicate the statistical significance levels according to Student's t-test: **P < 0.01 and *P < 0.05.
Figure 7
Figure 7
Protein sequence analysis of SLES. (A) Predicted schematic of the SLES protein. (B) Amino acid sequence alignment of the SLES kinase domain with that of other proteins indicated that SLES had a highly conserved kinase domain. Black boxes indicate identical residues and gray boxes indicate conservative substitutions. Roman numerals indicate the 11 characteristic sub-domains of protein kinases. The Raf specific motif is shown in red boxes. Asterisk indicates the position where a single amino acid change occurred in the sles mutant. Shown are: Oryza brachyantha (XP_006657649), Setaria italic (XP_004956079), Zea mays (XP_008664484), Brachypodium distachyon (XP_003560267), and Sorghum bicolor (XP_002466055).

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References

    1. Agrawal G. K., Jwa N. S., Rakwal R. (2000a). A novel rice (Oryza sativa L.) acidic PR1 gene highly responsive to cut phytohormones and protein phosphatase inhibitors. Biochem. Biophys. Res. Commun. 274, 157–165. 10.1006/bbrc.2000.3114 - DOI - PubMed
    1. Agrawal G. K., Rakwal R., Jwa N. S. (2000b). Rice (Oryza sativa L.) OsPR1b gene is phytohormonally regulated in close interaction with light signals. Biochem. Biophys. Res. Commun. 278, 290–298. 10.1006/bbrc.2000.3781 - DOI - PubMed
    1. Arnon D. (1949). Estimation of total chlorophyll. Plant Physiol. 24, 1–15. - PMC - PubMed
    1. Asai T., Tena G., Plotnikova J., Willmann M. R., Chiu W., Gomez-Gomez L., et al. . (2002). MAP kinase signalling cascade in Arabidopsis innate immunity. Nature 415, 977–983. 10.1038/415977a - DOI - PubMed
    1. Boch J., Verbsky M., Robertson T., Larkin J., Kunkel B. (1998). Analysis of resistance gene-mediated defense responses in Arabidopsis thaliana plants carrying a mutation in CPR5. Mol. Plant Microbe. Interact. 11, 1196–1206. 10.1094/MPMI.1998.11.12.1196 - DOI

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