Differental Proteomic Analysis Of Wheat Root Plasma Membrane Proteins Under Stress By Polycyclic Aromatic Hydrocarbons

Polycyclic aromatic hydrocarbons(PAHs)are priority pollutants due to their potential toxicity,mutagenicity and carcinogencity.It is of great importance to investigate the root plasma membrane(PM)proteome of PAHs for elucidating the bioprocess of PAHs transport across cell membrane.However,reports on the PAHs absorbing mechanism are few.In this paper,wheat was employed to study the effects of temperature,light intensity and incubation time on root plasma membrane protein content.In order to establish two-dimensional elecreophoresis(2-DE)system for wheat root plasma membrane proteomic analysis,SDS-PAGE gel concentration,isoelectric focusing parameter and loading amount of samples were investigated and optimized.The differential expression of proteome in wheat root PM stressed by phenanthrene(Phe)was analyzed with the established 2-DE system,and the differential expression protein spots were identified by MALDI-TOF/TOF-MS.This study provides both a novel insight into response of wheat root PM proteome to Phe exposure and methodological support for wheat root PM proteomic analysis.The main results obtained are shown as follows:1.The protein content was highest at 25?,18 000 Lux and 3-d incubation.The results of orthogonal test further proved that the optimum culture conditions for wheat root plasma membrane protein were 25?,18000 Lux and 3 d of incubation,respectively,with plasma membrane protein of 3.7891 ?g/(10 g).Our results can provide incubation conditions for the investigation of root plasma membrane proteome.2.High-quality,good-repeatability 2-DE maps were obtained under conditions with 10%SDS-PAGE gel concentration,18 h isoelectric focusing time and 450 ?g/gel loading quantities.The orthogonal test further proved this result.The system of 2-DE was suitable for plasma membrane proteomic analysis of wheat root.3.About 200 protein spots were distinguished on each 2-DE map of wheat root PM treated with Phe for 4 h and without Phe.14 differential expression proteins were identified through MALDI-TOF/TOF-MS analysis combined with retrieving NCBI database.These identified proteins,including members of albumin,enolase,cobalamin-independent methionine synthase,adenosine kinase,beta-glucosidase,apha-fetoprotein,heat shock proteins,glyceraldehyde-3-phosphate dehydrogenase and lactoylglutathione lyase were involved in coordinated regulation,signal transduction,stress response and energy metabolism,indicating that Phe stress has a great impact on wheat root PM proteome,and forced root PM to resist Phe stress.