| Recently, the emerging gasotransmitter hydrogen sulfide （H2S）, acting as an important mediator in regulating plants physiological pathway, has attracted more and more attention. H2S is involved in regulating roots growth and morphogenesis, promoting seed germination, enhancing plant photosynthesis, promoting flowering, delaying senescence. Additionally, H2S is also a critical supporter in protecting plants against environmental stresses.H2S relieved toxic symptoms caused by heavy metals （HMs） and enhanced plants tolerance to HMs stress. Reports have showed that cadmium （Cd） stress activated the endogenous H2S production in plants, then the H2S was employed to enhance the contents of reduced glutathione （GSH） and ascorbic acid （AsA）, as well as regulate the activities of antioxidant enzymes, maintaining the balance of intracellular redox state and alleviating the Cd stress induced oxidative damage. H2S improved Na+-ATPase, K+-ATPase and H+-ATPase activities in plants to inhibit the absorption of HM ions, while the absorption of S, P, Ca, Mg and Fe element was increased simultaneously. Moreover, H2S also inhibited the activity of H2O2 mediated Ca2+ channels to block the Cd uptake and activated some transporters in the vacuole membrane to transport the cytoplasm Cd2+ to the vacuole, thereby reduced the content of Cd2+ in the cytoplasm. Exogenous H2S enhanced the tolerance to Cd stress in plants by improving the content of chlorophyll, enhancing photosynthesis, promoting the development of chloroplast thylakoid, increasing the amount of mitochondria. In addition, the crosstalk between H2S and other signaling molecules or plant hormones in H2S alleviating HMs stress has been reported successively.The cabbage （Brassica rapa L. ssp. Pekinensis）was employed in this study. We used pharmacological, physiological, biochemical and cytological techniques to explore the mechanism of H2S in plants fighting against Cd stress. The positive effects of H2S in relieving the Cd stress induced inhibition of root elongation in cabbage were supported from phenotype, cytogenetic and molecular levels. The main results in this study are as follows:1. During Cd stress, the roots growth of cabbage seedlings was inhibited in Cd2+ contentration-dependent manner, the death rate of root cells, simultaneously, was increased rapidly. What’s more, the length of roots and the death rate of root cells had negative correlation, suggesting that there might be some connection between the root cells death and the roots elongation during Cd stress. Subsequent results showed that the production rate of endogenous H2S and the expression of H2S synthase coding genes in cabbage seedlings were activated significantly by Cd stress. Exogenous H2S fumigation, as NaHS being the H2S donor, enhanced the cabbage tolerance to Cd stress. It was found that H2S with 5 μM relieved the inhibition of roots growth caused by Cd stress, significantly reduced the root cells death and attenuated the DNA degradation in Cd stressed cabbage seedling. H2S reduced the Cd stress caused reactive oxygen species （ROS） accumulation in cabbage roots, then relieved the ROS-induced cell death of roots and lipid peroxidation, maintaining the integrity of the plasma membrane structure. These results suggested that H2S involved in the roots of cabbage seedlings response to Cd stress, and reduced Cd stress induced cell death by inhibiting the accumulation of excessive ROS in cabbage roots.2. H2S reduced the content of hydrogen peroxide （H2O2）, which is the key member of ROS, in the roots of Cd stressed cabbage. The content of malondialdehyde （MDA）, one of the decomposition products of lipid peroxidation, and the content of protein carbonylation product （PCO）, were also significantly reduced by H2S. Further results showed that H2S regulated the ROS metabolic system, down-regulated the expression of BrarohD and BrarohF, which encoded the O2·- generation enzyme NADPH oxidase （NOX） located in plasma membrane, and thus suppressed the activity of NOX. During Cd Stress, the antioxidant enzymes superoxide dismutase （SOD） and catalase （CAT）, which contribute to scavenging ROS, can also be regulated by H2S. Both SOD and CAT transcription levels and enzymatic activities were enhanced by H2S. However, the peroxidase （POD） and ascorbate peroxidase （APX） were hardly regulated by H2S. Furthermore, H2S pretreatment increased the synthesis of Cd stress mediated glutathione （GSH） and reduced the content of oxidized glutathione GSSG, and thus enhanced the GSH/GSSG ratio, which kept a high AsA content by regenerating the reduced AsA through GSH-AsA cycle signaling pathway. In summary, H2S mitigated the Cd stress caused oxidative damage in plants mainly through keeping the ROS content at the normal level.3. The roots growth of cabbage seedlings and cell mitosis in root tips were obviously inhibited by Cd stress, while H2S enhanced cell mitosis in root tips of Cd stressed seedlings, moreover, H2S significantly reduced the chromosome breakage and hysteresis as well as the chromosome bridges during cell mitosis, meanwhile the incidence of cell micronuclei can also be reduced by H2S. The Cd stress induced DNA damage can be mitigated by H2S, with the DNA-protein crosslink and genomic DNA cleavage decreased. The expression of PARP1 and PARP2, which encode the poly adenosine diphosphate-ribose polymerase （PARP）, related to DNA repair, were significantly up-regulated by H2S pretreatment. Overall, H2S increased mitosis of cells in root tips and decreased the incidence of cell micronuclei in Cd stressed cabbage roots, mainly mediated through restraining the DNA damage and chromosomal abnormalities during cell mitosis.4. With the extension of Cd stress, the lipid peroxidation product MDA and the relative conductivity of cells as well as the permeability of cell membrane increased widely. While H2S pretreatment weakened the relative conductivity of cells increased by Cd stress and improved the relative water content in the roots of cabbage. Moreover, the Cd stress increased soluble sugar and proline contents were strengthened by H2S pretreatment but weakened by the H2S production inhibitor HA pretreatment, while the soluble proteins and betaine contents were hardly affected by H2S.In summary, H2S synthesis was activated by Cd stress, then the H2S was utilized to participate in complicated pathways to regulate the generation and scavenging of ROS and thus protect the cells of root tips from being injured by Cd stress. In addition, H2S relieved DNA damage mediated by Cd stress and promoted mitosis in Cd stressed cabbage seedlings. H2S can also help cabbage resist to Cd stress by regulating the contents of some substances related to osmotic adjustment. Therefore, H2S participated in complicated defense and regulation pathways to mitigate the inhibition of root growth in cabbage seedlings caused by Cd stress. |
