Analysis Of Differently Expressed Proteins In Agaricus Bisporus Under Heat Stress

Agaricus bisporus is the manual cultivation most widespread,the output highest, the consumption quantity biggest edible fungus in the present world.It belongs to stable-temperature fruitfulness fungi,temperature is the main factor affecting its growth.The enforced and deeply research of thermotolerance moleculer biology of A.bisporus,especially researching on different expression proteins of heat shock and heat-resistant molecular mechanism by the modern biological technology,is important to breeding of A.bisporus for heat-tolerance,high yield,high quality in theory and practice.First,the ITS test are carried out to identify strains 02 and 8213.strains 02 and 8213 are identified to different strains of A.bisporus by ITS test.Strains 02 and 8213 are grouped into four groups:cultured on 24℃;cultured on 32℃for 6h,then cultured on 37℃for 6h;cultured on 37℃;cultured on 42℃.From the morphotype of A.bisporus in different temperature conditions we found:strain 02 is more thermotolerance than strain 8213;lower sublethal temperature is helpful to thermotolerance.Secondly,We have used two dimensional electrophoresis(2-DE) and Mass spectrometry(MALDI-TOF/MS) to identify proteins that are differentially expressed in group of strain 02 under heat stress and under normal temperature and group of strains 02 and 8213 under heat stress.From 2-DE data of strain 02 under heat stress and under normal temperature,32 resolved differentially expressed proteins were detected,5 of those were induced,14 of those were up-regulated,4 of those disappear,9 of those were down-regulated.From 2-DE data of strains 02 and 8213 under heat stress,41 resolved differentially expressed proteins were detected,18 of those were only found in strain 02,17 of those were were up-regulated in strain 02,3 of those were only found in strain 8213,3 of those were up-regulated in strain 8213.All the 73 spots were subjected to tryptic digestion followed by MALDI-TOF-MS, compared in Mascot.Total 58 proteins were identified and classified into different functional categories by COGs.They are classified into molecular chaperon,energy, metabolism,cellular processes and signaling,information storage and processing, function unknown.Six of these are molecular chaperon proteins,accounting for 10.3%, showed that molecular chaperon proteins(including HSP) are helpful to thermotolerance.Fourteen of these are energy and metabolism proteins,accounting for 24.1%,showed that the normal energy metabolism is interfered.Eleven of these are cellular processes and signaling,information storage and processing proteins, accounting for 19%,showed that A.bisporus regulate heat shock response on gene transcription level.Heat shock protein HSS1 is identified up-regulated in strain 02 under heat stress. HSP70 is identified up-regulated in strain 02 caompared to strain 8213.From real-time PCR detection,we find HSS1 and HSP70 are up-regulated to 5 times;in strains 02 and 8213 under heat stress,HSS1 and HSP70 are up-regulated to 1.6 and 2.8 in strain 02,showed that HSS1 and HSP70 are up-regulated induced by gene transcription level.The study found differentially expressed proteins in group of strain 02 under heat stress and under normal temperature and group of strains 02 and 8213 under heat stress by 2-DE,then identify them by MALDI-TOF-MS and PMF.Analyze the identified differentially expressed proteins and understand the mechanism of heat stress is helpful to reveal heat-resistant molecular mechanism of A.bisporus and breed heat-tolerance strain by genetic engineering.