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A Novel Pathogenesis-Related Class 10 Protein Gly m 4l, Increases Resistance upon Phytophthora sojae Infection in Soybean (Glycine max [L.] Merr.).

Fan S, Jiang L, Wu J, Dong L, Cheng Q, Xu P, Zhang S - PLoS ONE (2015)

Bottom Line: HQ913577.1).The recombinant Gly m 4l protein showed RNase activity and displayed directly antimicrobial activity that inhibited hyphal growth and reduced zoospore release in P. sojae.Further analyses showed that the RNase activity of the recombinant protein to degrading tRNA was significantly affected in the presence of zeatin.

View Article: PubMed Central - PubMed

Affiliation: Soybean Research Institute, Key Laboratory of Soybean Biology of Chinese Education Ministry, Northeast Agricultural University, Harbin, 150030, Heilongjiang, People's Republic of China.

ABSTRACT
Phytophthora root and stem rot of soybean, caused by Phytophthora sojae (P. sojae), is a destructive disease in many soybean planting regions worldwide. In a previous study, an expressed sequence tag (EST) homolog of the major allergen Pru ar 1 in apricot (Prunus armeniaca) was identified up-regulated in the highly resistant soybean 'Suinong 10' infected with P. sojae. Here, the full length of the EST was isolated using rapid amplification of cDNA ends (RACE). It showed the highest homology of 53.46% with Gly m 4 after comparison with the eight soybean allergen families reported and was named Gly m 4-like (Gly m 4l, GenBank accession no. HQ913577.1). The cDNA full length of Gly m 4l was 707 bp containing a 474 bp open reading frame encoding a polypeptide of 157 amino acids. Sequence analysis suggests that Gly m 4l contains a conserved 'P-loop' (phosphate-binding loop) motif at residues 47-55 aa and a Bet v 1 domain at residues 87-120 aa. The transcript abundance of Gly m 4l was significantly induced by P. sojae, salicylic acid (SA), NaCl, and also responded to methyl jasmonic acid (MeJA) and ethylene (ET). The recombinant Gly m 4l protein showed RNase activity and displayed directly antimicrobial activity that inhibited hyphal growth and reduced zoospore release in P. sojae. Further analyses showed that the RNase activity of the recombinant protein to degrading tRNA was significantly affected in the presence of zeatin. Over-expression of Gly m 4l in susceptible 'Dongnong 50' soybean showed enhanced resistance to P. sojae. These results indicated that Gly m 4l protein played an important role in the defense of soybean against P. sojae infection.

No MeSH data available.


Related in: MedlinePlus

Expression patterns analysis of Gly m 4l by quantitative real-time PCR.(A) The transcript abundance of Gly m 4l in the root, stem and leaf. The Ct value of each sample was normalized to the Ct value of GmEF1B. (B) The transcript abundance of Gly m 4l in response to P. sojae infection. The Ct value of each sample was normalized to the Ct value of GmActin4, and the relative expression of Gly m 4l was compared with mock plants at the same time point. (C) and (D), The transcript abundance of Gly m 4l in response to various stresses. Exogenous chemicals are SA (0.5 mM), MeJA (100 μM), ET (0.2 mM ethephon), ABA (50 mM), GA3 (50 mg.L-1), NaCl (100 mM), PEG (20%), and cold (4°C). The Ct value of each sample was normalized to the Ct value of GmActin4 and the relative expression of Gly m 4l was compared with mock plants at the same time point. The experiments were performed on three biological replicates with their respective three technical replicates and statistically analysed using Student’s t-test (*P<0.05, **P<0.01). Bars indicate standard error of the mean (SE).
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pone.0140364.g003: Expression patterns analysis of Gly m 4l by quantitative real-time PCR.(A) The transcript abundance of Gly m 4l in the root, stem and leaf. The Ct value of each sample was normalized to the Ct value of GmEF1B. (B) The transcript abundance of Gly m 4l in response to P. sojae infection. The Ct value of each sample was normalized to the Ct value of GmActin4, and the relative expression of Gly m 4l was compared with mock plants at the same time point. (C) and (D), The transcript abundance of Gly m 4l in response to various stresses. Exogenous chemicals are SA (0.5 mM), MeJA (100 μM), ET (0.2 mM ethephon), ABA (50 mM), GA3 (50 mg.L-1), NaCl (100 mM), PEG (20%), and cold (4°C). The Ct value of each sample was normalized to the Ct value of GmActin4 and the relative expression of Gly m 4l was compared with mock plants at the same time point. The experiments were performed on three biological replicates with their respective three technical replicates and statistically analysed using Student’s t-test (*P<0.05, **P<0.01). Bars indicate standard error of the mean (SE).

Mentions: Quantitative real-time PCR was performed to assess the tissue-specific expression of Gly m 4l in ‘Suinong 10’ soybean. The results showed that Gly m 4l was constitutively and highly expressed in the roots, followed by the leaves and stems (Fig 3A). The transcript abundance of Gly m 4l was investigated in the fully expanded unifoliolate leaves of ‘Suinong 10’ soybean in response to biotic and abiotic stresses. For P. sojae infection, a significant induction of Gly m 4l was detected from 6–72 h after the treatment, and the transcripts reached a maximum level at 24 h (Fig 3B). The transcript abundance of Gly m 4l under abiotic stresses was also investigated, such as NaCl, PEG, and cold (Fig 3C). NaCl treatment induced a significant up-regulation of Gly m 4l transcripts from 12–72 h. Cold induced an upregulation of Gly m 4l expression significantly only at 72 h after the treatment. There was little change for Gly m 4l transcript under the PEG stress. Meanwhile, the transcript abundance of Gly m 4l in response to exogenous chemicals including SA, MeJA, ET, ABA or GA3 was then carried out (Fig 3D). Sprayed with SA, the transcript of Gly m 4l was induced significantly from 3–24 h after the treatment, and reached a maximum level at 6 h. The Gly m 4l transcripts were also induced by MeJA and ET. There was almost down-regulation of the Gly m 4l transcript with the treatments of ABA and GA3.


A Novel Pathogenesis-Related Class 10 Protein Gly m 4l, Increases Resistance upon Phytophthora sojae Infection in Soybean (Glycine max [L.] Merr.).

Fan S, Jiang L, Wu J, Dong L, Cheng Q, Xu P, Zhang S - PLoS ONE (2015)

Expression patterns analysis of Gly m 4l by quantitative real-time PCR.(A) The transcript abundance of Gly m 4l in the root, stem and leaf. The Ct value of each sample was normalized to the Ct value of GmEF1B. (B) The transcript abundance of Gly m 4l in response to P. sojae infection. The Ct value of each sample was normalized to the Ct value of GmActin4, and the relative expression of Gly m 4l was compared with mock plants at the same time point. (C) and (D), The transcript abundance of Gly m 4l in response to various stresses. Exogenous chemicals are SA (0.5 mM), MeJA (100 μM), ET (0.2 mM ethephon), ABA (50 mM), GA3 (50 mg.L-1), NaCl (100 mM), PEG (20%), and cold (4°C). The Ct value of each sample was normalized to the Ct value of GmActin4 and the relative expression of Gly m 4l was compared with mock plants at the same time point. The experiments were performed on three biological replicates with their respective three technical replicates and statistically analysed using Student’s t-test (*P<0.05, **P<0.01). Bars indicate standard error of the mean (SE).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4608668&req=5

pone.0140364.g003: Expression patterns analysis of Gly m 4l by quantitative real-time PCR.(A) The transcript abundance of Gly m 4l in the root, stem and leaf. The Ct value of each sample was normalized to the Ct value of GmEF1B. (B) The transcript abundance of Gly m 4l in response to P. sojae infection. The Ct value of each sample was normalized to the Ct value of GmActin4, and the relative expression of Gly m 4l was compared with mock plants at the same time point. (C) and (D), The transcript abundance of Gly m 4l in response to various stresses. Exogenous chemicals are SA (0.5 mM), MeJA (100 μM), ET (0.2 mM ethephon), ABA (50 mM), GA3 (50 mg.L-1), NaCl (100 mM), PEG (20%), and cold (4°C). The Ct value of each sample was normalized to the Ct value of GmActin4 and the relative expression of Gly m 4l was compared with mock plants at the same time point. The experiments were performed on three biological replicates with their respective three technical replicates and statistically analysed using Student’s t-test (*P<0.05, **P<0.01). Bars indicate standard error of the mean (SE).
Mentions: Quantitative real-time PCR was performed to assess the tissue-specific expression of Gly m 4l in ‘Suinong 10’ soybean. The results showed that Gly m 4l was constitutively and highly expressed in the roots, followed by the leaves and stems (Fig 3A). The transcript abundance of Gly m 4l was investigated in the fully expanded unifoliolate leaves of ‘Suinong 10’ soybean in response to biotic and abiotic stresses. For P. sojae infection, a significant induction of Gly m 4l was detected from 6–72 h after the treatment, and the transcripts reached a maximum level at 24 h (Fig 3B). The transcript abundance of Gly m 4l under abiotic stresses was also investigated, such as NaCl, PEG, and cold (Fig 3C). NaCl treatment induced a significant up-regulation of Gly m 4l transcripts from 12–72 h. Cold induced an upregulation of Gly m 4l expression significantly only at 72 h after the treatment. There was little change for Gly m 4l transcript under the PEG stress. Meanwhile, the transcript abundance of Gly m 4l in response to exogenous chemicals including SA, MeJA, ET, ABA or GA3 was then carried out (Fig 3D). Sprayed with SA, the transcript of Gly m 4l was induced significantly from 3–24 h after the treatment, and reached a maximum level at 6 h. The Gly m 4l transcripts were also induced by MeJA and ET. There was almost down-regulation of the Gly m 4l transcript with the treatments of ABA and GA3.

Bottom Line: HQ913577.1).The recombinant Gly m 4l protein showed RNase activity and displayed directly antimicrobial activity that inhibited hyphal growth and reduced zoospore release in P. sojae.Further analyses showed that the RNase activity of the recombinant protein to degrading tRNA was significantly affected in the presence of zeatin.

View Article: PubMed Central - PubMed

Affiliation: Soybean Research Institute, Key Laboratory of Soybean Biology of Chinese Education Ministry, Northeast Agricultural University, Harbin, 150030, Heilongjiang, People's Republic of China.

ABSTRACT
Phytophthora root and stem rot of soybean, caused by Phytophthora sojae (P. sojae), is a destructive disease in many soybean planting regions worldwide. In a previous study, an expressed sequence tag (EST) homolog of the major allergen Pru ar 1 in apricot (Prunus armeniaca) was identified up-regulated in the highly resistant soybean 'Suinong 10' infected with P. sojae. Here, the full length of the EST was isolated using rapid amplification of cDNA ends (RACE). It showed the highest homology of 53.46% with Gly m 4 after comparison with the eight soybean allergen families reported and was named Gly m 4-like (Gly m 4l, GenBank accession no. HQ913577.1). The cDNA full length of Gly m 4l was 707 bp containing a 474 bp open reading frame encoding a polypeptide of 157 amino acids. Sequence analysis suggests that Gly m 4l contains a conserved 'P-loop' (phosphate-binding loop) motif at residues 47-55 aa and a Bet v 1 domain at residues 87-120 aa. The transcript abundance of Gly m 4l was significantly induced by P. sojae, salicylic acid (SA), NaCl, and also responded to methyl jasmonic acid (MeJA) and ethylene (ET). The recombinant Gly m 4l protein showed RNase activity and displayed directly antimicrobial activity that inhibited hyphal growth and reduced zoospore release in P. sojae. Further analyses showed that the RNase activity of the recombinant protein to degrading tRNA was significantly affected in the presence of zeatin. Over-expression of Gly m 4l in susceptible 'Dongnong 50' soybean showed enhanced resistance to P. sojae. These results indicated that Gly m 4l protein played an important role in the defense of soybean against P. sojae infection.

No MeSH data available.


Related in: MedlinePlus