Differential expression of defense/stress-related marker proteins in leaves of a unique rice blast lesion mimic mutant (blm)

J Proteome Res. 2006 Oct;5(10):2586-98. doi: 10.1021/pr060092c.

Abstract

We analyzed a unique rice (Oryza sativa L.) blast lesion mimic (blm) mutant for differentially expressed proteins in leaves of one- and two-week-old seedlings manifesting the lesion mimic phenotype. Gel-based one- and two-dimensional electrophoresis (1- and 2-DGE) was performed using leaves (blm and wild-type, WT) before (stage 1, S1) and after (stage 2, S2) lesion formation. 1-DGE immunoblotting revealed potent increase in the expression of a key pathogenesis-related (PR) marker biosynthetic enzyme, naringenin 7-O-methyltransferase, involved in rice phytoalexin sakuranetin biosynthesis, and three oxidative-stress-related marker proteins, catalase, ascorbate peroxidase (APX), and superoxide dismutase (SOD) in leaves of the blm mutant. 2-D gel immunoblotting analysis with anti-APX and anti-SOD antibodies revealed newly appearing cross-reacting protein spots in blm. 2-DGE analysis detected 50 Coomassie brilliant blue-stained protein spots differentially expressed in blm. A total of 23 and 44 protein spots was excised for analysis by N-terminal amino acid sequencing and nano-electrospray ionization liquid chromatography mass spectrometry, respectively; 26 nonredundant proteins were identified. The pathogenesis-related class 5 and 10 proteins, including a new OsPR10d protein, were significantly induced in blm. The OsPR5 protein spot was stained with Pro-Q Diamond phosphoprotein gel stain suggesting OsPR5 to be a putative phosphoprotein. Surprisingly, protein spot 20, a leaf OsPR10b, showed identity to a rice root-specific PR-10 (RSOsPR10). To resolve this discrepancy, we checked its expression in leaves of blm and WT (S1 and S2), respectively, using gene-specific primers and reverse transcriptase-polymerase chain reaction; RSOsPR10 mRNA was found to express in the leaves.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Ascorbate Peroxidases
  • Biomarkers / analysis
  • Catalase / analysis
  • Catalase / genetics
  • Electrophoresis, Gel, Two-Dimensional
  • Energy Metabolism
  • Flavonoids / metabolism
  • Heat-Shock Proteins / analysis*
  • Heat-Shock Proteins / genetics
  • Methyltransferases / analysis
  • Molecular Sequence Data
  • Mutation
  • Oryza / genetics
  • Oryza / metabolism*
  • Oxidative Stress / genetics
  • Peroxidases / analysis
  • Peroxidases / genetics
  • Phosphoproteins / analysis
  • Phosphoproteins / genetics
  • Photosynthesis
  • Phytoalexins
  • Plant Diseases* / genetics
  • Plant Leaves / genetics
  • Plant Leaves / metabolism*
  • Plant Proteins / analysis*
  • Plant Proteins / genetics
  • Proteome / analysis*
  • Proteome / genetics
  • RNA, Messenger / analysis
  • RNA, Messenger / genetics
  • Sesquiterpenes
  • Superoxide Dismutase / analysis
  • Superoxide Dismutase / genetics
  • Terpenes / metabolism

Substances

  • Biomarkers
  • Flavonoids
  • Heat-Shock Proteins
  • Phosphoproteins
  • Plant Proteins
  • Proteome
  • RNA, Messenger
  • Sesquiterpenes
  • Terpenes
  • sakuranetin
  • Peroxidases
  • Ascorbate Peroxidases
  • Catalase
  • Superoxide Dismutase
  • Methyltransferases
  • naringenin 7-O-methyltransferase
  • Phytoalexins