H₂S Improves Salt Tolerance Through Persulfidation Of PM H⁺-ATPase In Zea Mays And Arabidopsis Thaliana

Plasma membrane(PM)H+-ATPase plays a pivotal role in regulating ion transport in PM of plant cells.Under salt stress,PM H+-ATPase can maintain the Na+/K+ homeostasis in plant cells,thus PM H+-ATPase plays a vital role in plant cells against salt stress.Hydrogen sulfide(H2S),a small gasotransmitter,plays an important and positive physiological function in regulating plant adaptation to environmental stress and growth.H2S mediates post-translational modification of proteins that is named persulfidation.Persulfidation is the main way that H2S regulates protein activity.It has been reported that H2S can improve salt tolerance in plants,but the molecular mechanism of salt tolerance needs further research.In this study,model plants Arabidopsis thaliana and model maize(Zea mays)were used as research materials.The effect of H2S on plant salt resistance was studied,and the molecular mechanism of H2S to improve plant salt resistance was explored with the persulfidation of H2S as the entry point.The main research results obtained are as follows.(1)Treating plants with H2S donor NaHS can alleviate the inhibitory effect of salt stress on the growth of Zea mays and Arabidopsis seedlings,while increasing the chlorophyll content in plant cells and reducing the malondialdehyde(MDA)content and ion permeability.In addition,NaHS pretreatment can reduce Na+ content in plants under salt stress,inhibit K+ efflux,and maintain ion balance under stress conditions;These results indicate that H2S increases the tolerance of Arabidopsis seedlings and Zea mays seedlings to salt stress.(2)To determine persulfidation of plant PM H+-ATPase protein by H2S.In this study,the persulfidation domain peptides of PM H+-ATPase proteins in Arabidopsis thaliana and Zea mays were prokaryotically expressed,and purified recombinant proteins were obtained.NaHS pretreatment increased the persulfidation level of PM H+-ATPase recombinant protein,and it increases with increaseing concentration.In addition,the recombinant protein was treated with persulfidation removal substance dithiothreitol(DTT),and it was found that DTT treatment eliminated persulfidation of the protein,which induced by NaHS.The above results proved that hydrogen sulfide can regulate the persulfidation level of PM H+-ATPase protein in Arabidopsis and Zea mays.(3)H2S-mediated persulfidation can increase the PM H+-ATPase activity of Zea mays and Arabidopsis seedling,accelerate the movement of Na+/H+reverse transport proteins,promote Na+efflux,and alleviate the damage caused by salt stress to plants.The plasma membrane microcapsules of plants were pretreated with NaHS,and it was found that NaHS can increase the PM H+-ATPase activity of plants;Non-invasive Micro-test Technology(NMT)was used to measure the H+flow on the surface of the plant root cell membrane under different treatments.It was found that the transient addition of NaHS could cause H+efflux on the plant root cell surface,and NaHS pretreatment could accelerate H+influx on the root cell surface under salt stress.In summary,this study reveals the molecular mechanism of H2S-mediated PM H+-ATPase in Zea mays and Arabidopsis plant cells modified by persulfidation in response to plant salt stress.The results show that H2S can improve PM H+-ATPase activity through persulfidation,promote Na+efflux in plants,reduce K+ efflux,maintain plant Na+,K+homeostasis,and participate in plant responses to salt stress.