Secretome analysis of differentially induced proteins in rice suspension-cultured cells triggered by rice blast fungus and elicitor.
Secreted proteins were investigated in rice suspension-cultured cells treated with rice blast fungus Magnaporthe grisea and its elicitor using biochemical and 2-DE coupled with MS analyses followed by their in planta mRNA expression analysis. M. grisea and elicitor successfully interacted with suspension-cultured cells and prepared secreted proteins from these cultures were essentially intracellular proteins free. Comparative 2-D gel analyses identified 21 differential protein spots due to M. grisea and/or elicitor over control. MALDI-TOF-MS and microLC-ESI-MS/MS analyses of these protein spots revealed that most of assigned proteins were involved in defense such as nine chitinases, two germin A/oxalate oxidases, five domain unknown function 26 (DUF 26) secretory proteins, and beta-expansin. One chitin binding chitinase protein was isolated using chitin binding beads and strong enzymatic activity was identified in an in-gel assay. Interestingly, their protein abundance correlated well at transcript levels in elicitor-treated cultures as judged by semi-quantitative RT-PCR. Each identified differentially expressed protein group was compared at transcript levels in rice leaves inoculated with incompatible (KJ401) and compatible (KJ301) races of M. grisea. Time-course profiling revealed their inductions were stronger and earlier in incompatible than compatible interactions. Identified secreted proteins and their expression correlation at transcript level in suspension-cultured cells and also in planta suggest that suspension-cultured cells can be useful to investigate the secretome of rice blast-pathogen interactions.
Kim
Sun Tae
ST
Environmental Biotechnology National Core Research Center, Gyeongsang National University, Chinju, Korea.
Kang
Young Hyun
YH
Wang
Yiming
Y
Wu
Jingni
J
Park
Zee Yong
ZY
Rakwal
Randeep
R
Agrawal
Ganesh Kumar
GK
Lee
Sang Yeol
SY
Kang
Kyu Young
KY
D015153:Blotting, Western; D002478:Cells, Cultured; D002688:Chitinases / Q000235:genetics / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D018506:Gene Expression Regulation, Plant; D020082:Magnaporthe; D012275:Oryza / Q000166:cytology* / Q000378:metabolism / Q000382:microbiology*; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism; D018515:Plant Leaves / Q000235:genetics / Q000378:metabolism; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D012639:Seeds / Q000235:genetics / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D010940:Plant Proteins; C095825:expansin protein, plant; D010088:Oxidoreductases; C020598:oxalate oxidase; D002688:Chitinases