The Relationship Between H₂S Signal And MAPK In Response To Drought And Cold Stress In Arabidopsis

Hydrogen sulfide(H2S),the third gasotransmitter after nitric oxide(NO)and carbon oxide(CO),can be synthesized endogenously and performs important physiological functions in both animals and plants.H2 S is involved in growth and development,and biotic and abiotic stress resistance in plant.H2 S can increase the resistance ability to drought stress and cold stress in plant,however,the mechanism underlying still needs more exploration.MAPK is an essential and conserved component of the signaling transduction pathway that converts growth and environment signals into cellular responses in eukaryotes.MAPK plays an important role in plant responsing to drought stress and cold stress.However,the relationship between H2 S and MAPK has not been reported.In this study,we used Arabidopsis as material and explored the relationship between H2 S and MAPK under drought stress and cold stress.The main results are as follows:1.Drought stress activated both H2 S biosynthesis and gene expression of MAPKs.The increase in MAPK expression under drought stress was depressed in lcd/des1,a double mutant of H2 S synthesis.Na HS up-regulated the gene expression level of MAPK especially MPK4,so we selected MPK4 as our target and used mpk4 mutants for further studies.H2 S was able to alleviate the drought stress in wild-type(WT)but not in mpk4 mutant.Meanwhile,H2S-induced stomatal movement was impaired in mpk4 mutants.We then examined the role of H2 S and MPK4 in stomatal movements in response to abscisic acid(ABA)and hydrogen peroxide(H2O2).ABA-and H2O2-mediated stomatal movements were impaired in lcd/des1 and mpk4 mutants.H2S-induced stomatal closure was impaired in slac1-3 mutants.These results suggest that MPK4 is important downstream of H2 S in the drought stress response and in stomatal movement,and that the H2S-MPK4 cascade is involved in ABA-mediated stomatal movement to regulate the drought stress.2.The expression level of MAPK decreased in des1 and lcd,the mutant of H2 S synthesis.The induction of cold stress on MAPK gene was impaired in des1 and lcd.H2S's ability to alleviate cold stress was impaired in mpk4.Both H2 S and MPK4 regulated the expression levels of the cold response genes ICE1(inducer of CBF expression 1),CBF3(C-repeat-binding factors),COR15A(cold responsive 15A)and COR15B(cold responsive 15B).H2 S inhibited the opening of stomata under cold stress,and this function was impaired in mpk4.These results suggest that MPK4 is a downstream component of H2S-related cold-stress resistance,and H2 S and MPK4 both regulate the cold response genes and stomatal movement to response the cold stress.3.The ability of H2 S alleviating cold stress was not affected in mek2.MEK2 could not be S-sulfhydrated.MPK4 was basically S-sulfhydrated,and Na HS further increased the S-sulfhydration level of MPK4.These results suggeste that H2 S alleviates cold stress in an MEK2-independent way and H2 S modifies MPK4 through post-translational modification S-sulfhydration to increase the cold stress resistance ability.4.Na HS could up-regulate the gene expression level of CCA1(circadian clock associated 1)and PRR9(pseudo-response regulator 9)which are core circadian clock genes,and the peak expression of CCA1 and PRR9 was delayed in lcd/des1.The expression of CBFs,the cold stress response genes that are the downstream target of CCA1,was also regulated by H2 S.In lcd/des1,the peak expression of CBF1 and CBF3 was delayed and the expression level of CBF1,CBF2 and CBF3 was decreased.The seedlings of lcd/des1 were more sensitivity to cold stress than WT.Then the diurnal change of LCD(L-cysteine desulfhydrase)and DES1(desulfhydrase 1)expression level was explored,which encode H2 S production enzymes in Arabidopsis.The expression of LCD did not change significantly during 1 d,however,the expression of DES1 showed diurnal change with peak at 8 in the morning.The expression pattern of MPK3,MPK4 and MPK6 changed in lcd/des1.These results suggeste that H2 S can regulate the expression level of clock genes CCA1 and PRR9,as well as its downstream genes CBFs to increase resistance to cold stress.Above all,we found that H2 S is closely related to MAPK in response to drought stress and cold stress in Arabidopsis.MPK4,a member of MAPK,functions as a downstream component of H2 S,and H2 S can increase the resistance ability to drought stress and cold stress through S-sulfhydrating MPK4.