Chitinases are considered to act as defense proteins when plants are exposed to heavy metal stresses. Typical class III chitinase genes were cloned from Kandelia obovate, Bruguiera gymnorrhiza, and Rhizophora stylosa by using RT-PCR and RACE and named KoCHI III, BgCHI III, and RsCHI III. Bioinformatics analysis revealed that the three genes encoding proteins were all typical class III chitinases with the characteristic catalytic structure belonging to the family GH18 and located outside the cell. In addition, there are heavy metal binding sites in the three-dimensional spatial structure of the type III chitinase gene. Phylogenetic tree analysis indicated that CHI had the closest relationship with chitinase in Rhizophora apiculata. In mangrove plants, the balance of the oxidative system in the body is disrupted under heavy metal stress, resulting in increased H2O2 content. Real-time PCR illustrated that the expression level under heavy metal stress was significantly higher than that in the control group. Expression levels of CHI III were higher in K. obovate than in B. gymnorrhiza and R. stylosa. With the increase in heavy metal stress time, the expression level increased continuously. These results suggest that chitinase plays an important role in improving the heavy metal tolerance of mangrove plants.
Chitinases are believed to act as defense proteins when plants are exposed to heavy metal stress. Typical Class I chitinase genes were cloned from Bruguiera gymnorrhiza, Rhizophora stylosa, Kandelia obovata, and Avicennia marina using the methods of reverse-transcription–polymerase chain reaction and rapid amplification of cDNA ends. All four cDNA sequences of chitinase from the mangrove plants were 1092 bp in length and consisted of an open reading frame of 831 bp, encoding 276 amino acids. However, there were differences in the sequences among the four mangrove species. Four gene proteins have a signal peptide, are located in the vacuole, and belong to the GH19 chitinase family. The sequence of chitinase was highly similar to the protein sequences of Camellia fraternal chitinases. A real-time polymerase chain reaction was used to analyze the chitinase expressions of the above four mangrove species exposed to different concentrations of heavy metal at different times. The gene expression of chitinase was higher in Bruguiera gymnorrhiza leaves than in other mangrove plant species. With an increase in heavy metal stress, the expression level of Bruguiera gymnorrhiza increased continuously. These results suggest that chitinase plays an important role in improving the heavy metal tolerance of mangrove plants.
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