Yiming Wang

Publications (15) View all

• Article: Depletion of abundant plant RuBisCO protein using the protamine sulfate precipitation method.

  Yu Ji Kim, Hye Min Lee, Yiming Wang, Jingni Wu, Sang Gon Kim, Kyu Young Kang, Ki Hun Park, Yong Chul Kim, In Soo Choi, Ganesh Kumar Agrawal, Randeep Rakwal, Sun Tae Kim
  [show abstract] [hide abstract]
  ABSTRACT: Ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) is the most abundant plant leaf protein, hampering deep analysis of the leaf proteome. Here, we describe a novel protamine sulfate precipitation (PSP) method for the depletion of RuBisCO. For this purpose, soybean leaf total proteins were extracted using Tris-Mg/NP-40 extraction buffer. Obtained clear supernatant was subjected to the PSP method, followed by 13% SDS-PAGE analysis of total, PS-supernatant and -precipitation derived protein samples. In a dose-dependent experiment, 0.1% (w/v) PS was found to be sufficient for precipitating RuBisCO large and small subunits (LSU and SSU). Western-blot analysis confirmed no detection of RuBisCO LSU in the PS-supernatant proteins. Application of this method to Arabidopsis, rice, and maize leaf proteins revealed results similar to soybean. Furthermore, 2-DE analyses of PS-treated soybean leaf displayed enriched protein profile for the protein sample derived from the PS-supernatant than total proteins. Some enriched 2-D spots were subjected to MALDI-TOF-TOF analysis and were successfully assigned for their protein identity. Hence, the PSP method is: (i) simple, fast, economical, and reproducible for RuBisCO precipitation from the plant leaf sample; (ii) applicable to both dicot and monocot plants; and (iii) suitable for down-stream proteomics analysis.
  Proteomics 04/2013; · 4.43 Impact Factor
• Article: Secretome analysis of the rice bacterium Xanthomonas oryzae (Xoo) using in vitro and in planta systems.

  Yiming Wang, Sang Gon Kim, Jingni Wu, Hyun-Hye Huh, Su-Ji Lee, Randeep Rakwal, Ganesh Kumar Agrawal, Zee-Yong Park, Kyu Young Kang, Sun Tae Kim
  [show abstract] [hide abstract]
  ABSTRACT: Xanthomonas oryzae pv. oryzae (Xoo) causes bacterial blight disease in rice, and that severely affects yield loss (upto 50%) of total rice production. Here, we report a proteomics investigation of Xoo (compatible race K3)-secreted proteins, isolated from its in vitro culture and in planta infected rice leaves. Two-dimensional gel electrophoresis (2-DE) coupled with MALDI-TOF-MS and/or nLC-ESI-MS/MS approach identified 139 protein spots (out of 153 differential spots), encoding 109 unique proteins. Identified proteins belonged to multiple biological and molecular functions. Metabolic and nutrient uptake proteins were common up to both in vitro and in planta secretomes. However, pathogenicity, protease/peptidase, and host defense-related proteins were highly- or specifically-expressed during in planta infection. A good correlation was observed between protein and transcript abundances for nine proteins-secreted in planta as per semi-quantitative RT-PCR analysis. Transgenic rice leaf sheath (carrying PBZ1 promoter::GFP cell death reporter), when used to express a few of the identified secretory proteins, showed a direct activation of cell death signaling, suggesting their involvement in pathogenicity-related with secretion effectors. This work furthers our understanding of rice bacterial blight disease, and serves as a resource for possible translation in generating disease resistant rice plants for improved seed yield.
  Proteomics 03/2013; · 4.43 Impact Factor
• Article: Characterization of a newly identified rice chitinase-like protein (OsCLP) homologous to xylanase inhibitor.

  Jingni Wu, Yiming Wang, Sun Tae Kim, Sang Gon Kim, Kyu Young Kang
  [show abstract] [hide abstract]
  ABSTRACT: BACKGROUND: During rice blast fungal attack, plant xylanase inhibitor proteins (XIPs) that inhibit fungal xylanase activity are believed to act as a defensive barrier against fungal pathogens. To understand the role of XIPs in rice, a xylanase inhibitor was cloned from rice. The expression of this gene was examined at the transcriptional/translational levels during compatible and incompatible interactions, and the biochemical activity of this protein was also examined. RESULTS: Sequence alignment revealed that the deduced amino acid sequence of OsCLP shares a high degree of similarity with that of other plant TAXI-type XIPs. However, recombinant OsCLP did not display inhibitory activity against endo-1,4-beta-xylanase enzymes from Aureobasidium pullulans (A. pullulans) or Trichoderma viride (T. viride). Instead, an in-gel activity assay revealed strong chitinase activity. The transcription and translation of OsCLP were highly induced when rice was exposed to pathogens in an incompatible interaction. In addition, exogenous treatment with OsCLP affected the growth of the basidiomycete fungus Rhizoctonia solani through degradation of the hyphal cell wall. These data suggest that OsCLP, which has chitinase activity, may play an important role in plant defenses against pathogens. CONCLUSIONS: Taken together, our results demonstrate that OsCLP may have antifungal activity. This protein may directly inhibit pathogen growth by degrading fungal cell wall components through chitinase activity.
  BMC Biotechnology 01/2013; 13(1):4. · 2.35 Impact Factor
• Article: In-depth insight into in vivo apoplastic secretome of rice-Magnaporthe oryzae interaction.

  Sang Gon Kim, Yiming Wang, Kyung Hee Lee, Zee-Yong Park, Jongsun Park, Jingni Wu, Soon Jae Kwon, Yong-Hwan Lee, Ganesh Kumar Agrawal, Randeep Rakwal, Sun Tae Kim, Kyu Young Kang
  [show abstract] [hide abstract]
  ABSTRACT: The in vivo apoplastic fluid secretome of rice-blast fungus interaction remains largely uncharacterized. Here, we report a proteomics investigation of in vivo secreted proteins of rice leaves infected with incompatible (KJ401) and compatible (KJ301) races of Magnaporthe oryzae (M. oryzae) using 2-DGE and MudPIT coupled with MALDI-TOF-MS and/or nESI-LC-MS/MS analyses. Prepared fractions of secretory proteins were essentially free from cytoplasmic contamination. Two-DGE and MudPIT identified 732 secretory proteins, where 291 (40%) and 441 (60%) proteins were derived from rice and M. oryzae, respectively. Of these, 39.2% (rice) and 38.9% (M. oryzae) of proteins were predicted by SignalP as retaining signal peptides. Among these, rice secreted more proteins related to stress response, ROS and energy metabolism, whereas, M. oryzae secreted more proteins involved in metabolism and cell wall hydrolyses. Semi-quantitative RT-PCR revealed their differential expression under compatible/incompatible interactions. In vivo expression of M. oryzae glycosyl hydrolase (GH) protein family members using particle bombardment driven transient expression system showed that four GH genes could act as effectors within host apoplast possibly via interaction with host membrane bound receptor. The established in vivo secretome serves as a valuable resource toward secretome analysis of rice-M. oryzae interaction.
  Journal of proteomics 11/2012; · 5.07 Impact Factor
• Article: Physiological and proteomic analysis of young rice leaves grown under nitrogen-starvation conditions

  Sang Gon Kim, Yiming Wang, Jingni Wu, Kyu Young Kang, Sun Tae Kim
  [show abstract] [hide abstract]
  ABSTRACT: Rice grown in anaerobic waterlogged soil accumulates ammonium as a major source of nitrogen (N). We have compared the physiological symptoms of rice seedlings subjected to N-starvation stress with those receiving sufficient N, based on measurements of shoot/root length and weight and an analysis of protein expression patterns. N starvation marginally increased root growth but notably decreased shoot biomass. N uptake was reduced by >50% in the roots and shoots of N-starved seedlings. To better understand the mechanism of N starvation in rice, we performed a comparative proteome analysis of proteins isolated from rice leaves. Twenty-five differentially expressed proteins were analyzed by matrix-assisted laser desorption/ionization time-of-flight (TOF) mass spectrometry and electron spray ionization quadrupole TOF. Functional analysis of the N-starvation response proteins suggested their involvement in protein synthesis and fate, metabolism, and defense. These results indicate that these proteins may play important roles in regulating the plant’s complex adaptation responses for N use during N starvation. The proteins may be useful for further characterization of protein function in plant N nutrition. KeywordsNitrogen starvation–Proteomics–Rice–2-DGE
  Plant Biotechnology Reports 04/2012; 5(4):309-315. · 1.19 Impact Factor