The Regulation Mechanism Of Tauroursodesoxycholic Acid On Cell Death Mediated By Endoplasmic Reticulum Stress In Wheat Under Osmotic Stress

Endoplasmic reticulum stress(ER Stress)pathway is one of the important ways to respond to adversity in plant.The exploration of the the drought response from the perspective of ER stress has important theoretical significance for revealing the drought response mechanism in wheat.TUDCA,as an ER stress relieving agent,is widely used in the study of ER stress related research and treatment of clinical protein folding diseases in mammals.However,TUDCA is rarely studied in plants,especially in grain crop wheat.In order to explore the regulation mechanism of TUDCA on ER stress induced by osmotic stress simulated drought stress in wheat,we employed the methods of pharmacology,ecology,plant physiology,molecular biology,cytology and quantitative ubiquitination protein groups technology to carry out the study from multi angle and multi-level research.This study explored the effects of TUDCA on the growth and cell death of wheat under osmotic stress and its regulatory mechanism through a series of plant physiology,molecular biology and cell biology technology.Our results show that TUDCA: 1)Ameliorates the impact of osmotic stress on wheat height,fresh weight,and water content;2)Alleviates the decrease in chlorophyll content as well as membrane damage caused by osmotic stress;3)Decreases the accumulation of reactive oxygen species(ROS)by increasing the activity of antioxidant enzymes under osmotic stress,and;4)To some extent alleviates osmotic stress induced cell death probably by regulating endoplasmic reticulum(ER)stress related genes expression,for example the basic leucine zipper genes b ZIP60 B and b ZIP60 D,the binding proteins BiP1 and BiP2,the protein disulfide isomerase PDIL8-1,and glucose regulated protein GRP94.We also propose a model that illustrates how TUDCA alleviates osmotic stress-related wheat cell death,which provides an important theoretical basis for improving plant stress adaptation and elucidates the mechanisms of ER stress-related plant osmotic stress resistance.In this study,the function of apoptosis inhibitor TaBI-1 was studied by virus induced gene silencing(VIGS)technology and transgenic technology.1)With the VIGS technology,BSMV:asTaBI-1,the vector of virus induced gene silencing,was successfully constructed,and wheat that TaBI-1 gene the silenced were obtained through rubbing wheat leaves to inoculate virus with recombinant plasmid BSMV:asPDS.The natural drought treatment was carried out with WT,BSMV:0 and BSMV:asPDS wheat.The results showed that: campared with WT and empty BSMV:0wheat,TaBI-1 silenced wheat were more sensitive to drought stress.2)With transgenic technology,the TaBI-1 gene overexpression vector was successfully constructed,and the pure line of OE-TaBI-1Arabidopsis seeds were obtained.WT and 3 line of OE-TaBI-1 Arabidopsis thaliana conducted drought treatment and high temperature treatment respectively.The results show that: OE-TaBI-1Arabidopsis has stronger resistance and adaptability under drought stress and high temperature stress.In other words,the apoptosis inhibitor TaBI-1 can improve the stress resistance in wheat,especially drought and high temperature resistance,which will lay the foundation for further study of the mechanism of cell death mediated by drought induced wheat ER stress.In this study,the ubiquitination proteomics technology was used to analyze the effects of osmotic stress on wheat and the effects of TUDCA on cell death induced osmotic stress in wheat..The results show that: 1)osmotic stress changes the ubiquitination of wheat proteins,and TUDCA regulated the proteins ubiquitination under osmotic stress;2)ubiquitin modification is widely involved in multiple pathways of plant response to osmotic stress: cell stress and stimulation,signal transduction,cytochemical balance,glucose metabolism,unfolded protein response,endoplasmic reticulum stress response,ER related degradation(ERAD)pathway and other pathways.Under osmotic stress,some proteins in these pathways have significant ubiquitination modification changes;3)TUDCA regulates plant response to drought through proteins ubiquitination modification: TUDCA mainly affects DNA damage and repair,glycometabolism,calcium homeostasis,signaling and signal transduction pathways to modulate the osmotic stress response in wheat;4)we propose a regulated mechanism of TUDCA to alleviate PEG induced cell death from the perspective of protein ubiquitination in wheat:Osmotic stress changed the levels of ubiquitination of VCP/p97,Dsk2,HSP90,HMGB1,RAB7,PCNA and other cell death related proteins,which hindered the ERAD,DNA damage repair and autophagy pathway.It aggravated the occurrence of cell death to a certain extent in wheat.Yet,TUDCA pretreatment inhibited the changes of these proteins ubiquitin modification,which maintain the normal work of these pathways,and delayed or slowed down the occurrence of cell death.