Mechanism Of Herbivorous Insect Resistance And Salt Tolerance In Mangrove Plant Avicennia Marina

Mangrove,which is mainly distributed in the intertidal zone of tropical and subtropical coastal areas,is the main primary producer of the mangrove ecosystem and possess various ecosystem function,such as to prevent coastal erosion,to maintain water quality and to support a wide range of wildlife.Recently,the mangrove forest in China have been frequently attacked by various herbivorous insects leading to the death of a large number of mangrove plants.Noteworthy,Avicennia marina is more susceptible by herbivorous insects but little is known about the defensive mechanism of A.marina respond to insect attack.As a pioneer species of mangrove ecosystem,A.marina has a higher salt tolerance under high salinity condition.NO,which is a small signaling molecule in plant,play an important role in improving the salt tolerance in plants,but little is known about the regulation mechanism of NO-mediated the increase of salt tolerance in A.marina leaves.In addition,the salt secretion in leaves is one of the most important strategy for A.marina to growth under high salinity condition.However,the mechanism of salt secreting in A.marina leaves is still not clear.Therefore,this study aims to investigate the mechanism of herbivorous insect resistant and salt tolerance in A.marina.The conclusions as following:1.Based on the morphological observation,the insect isolated from A.marina leaves was preliminary identified as a moth in Phyllocnistis Genus.Subsequently,the results of DNA barcoding COI gene showed that the bootstrap values between our sample and Phyllocnistis ampelopsiella was 100,suggesting that the insect isolated from A.marina is Phyllocnistis ampelopsiella.To further understand the mechanism of mangrove plant in response to the leaf miner attack,the RNA of A.marina leaves attacked by leaf miner was extracted and used for transcriptional profile analysis.After sequencing,78,791 unigenes were obtained.For the expression level analysis,a total of 1654 unigenes were identified as differentially expressed genes(DEGs)including 767 and 887 unigenes that were up-regulated and down-regulated,respectively,under the attack by leaf miner.The KEGG enrichment analysis of DEGs showed that the pathways of photosynthesis including photosynthesis-antenna proteins,biosynthesis of amino acids including cysteine and methionine metabolism,ribosome of biogenesis in eukaryotes and other glycan degradation were significantly affected.In addition,the DEGs participated in jasmonic acid(JA)signaling pathway were identified,such as MAK1/3 in signaling transduction pathway,transcriptional factors MYBs and GLs in the downstream of JA signaling pathway and ISPH,GPPS,TPS14 in volatile organic compounds(VOCs)biosynthesis pathway.According to these results,we speculated that the stress signaling induced by insect attack was perceived and transducted from MAPK pathway to JA signaling pathway,then to activate the defensive response in A.marina leaves,such as to promote the degradation of chlorophyll,to reduce the biosynthesis of available carbonhydrate and protein,to strengthen the regulation of circadian rhythm and to enhance the synthesis of volatile organic compounds(VOCs).Besides,the activation of fatty acid degradation is conducive to make up for the lack of acetyl coenzyme A caused by the low efficiency of photosynthesis,thus provide material for VOCs synthesis.2.The analysis of photosynthesis in A.marina leave under high salinity and SNP addition treatments showed that high salinity treatment could induce not only the stomatal limitation but also non-stomatal limitation on photosynthetic reduction,while SNP addition could restore the non-stomatal limitation.Our proteomic data also found that NO alleviated the reduction of photosynthesis caused by non-stomatal limitation by enhancing the accumulation of RBCL,QOR and ATPase.Moreover,high salinity also inhibited the energy metabolism,primary metabolism,RNA transcription and protein translation in cell,while NO could partically alleviate the salt induced damage by promoting the accumulation of some proteins in these biological processes.Besides,the protein-protein interaction(PPI)analysis of identified protein in present study showed that ENO,HSP and ADK may play an important role in NO-mediated salt tolerance in A.marina.3.Our previous study has found that there were more salt crystals occurred on the surface of A.marina leaves under 400 mM NaCl treatment.In present study,more salt glands were obaserved under 400 mM NaCl treatment than that in 100 mM NaCl treatment.Besides,we also found that the secreting cells produced more vesicles or vesicles-like structure under 400 mM NaCl treatment.We suggested that vesicles play a very important role in salt secretion.To further understand the mechanism of salt secretion in A.marina leaves,iTRAQ was performed to analyze the protein profile in the epidermis of A.marina leaves.A total of 1596 proteins was successful identified and 123 proteins were defined as different expressed proteins(DEPs).Among them,the abundance of 57 proteins were increased and 66 proteins were decreased.After annotation in databases,these protein were divide into 8 catagorie:protein metabolism,secondary metabolism,carbohydrate metabolism,antioxidation proteins,signaling transdution,stress respond proteins,RNA transcription,ion transport and vesicle transport proteins and unknown proteins.The long-term treatment of high salinity seriously inhibited the growth of A.marina,and the abundance of proteins involve in RNA transcription,protein biosynthesis and carbonhydrate metabolism were decreased.To adapt to the long-term high salinity treatment,the abundance of proteins participated in signaling transduction were increased to control the protein synthesis in the downstream.As the salt signal was perceived by LRR-RPKs and CDPKs then transducted to CRT,CML and BSL.These three protein were mainly function in controlling RNA transcription and protein translation,subsequently the stress response proteins were produced.Noteworthy,the abundance of most proteins in ion transport and vesicle transport proteins were increased and most of them were involved in vesicles formation and transportation.The PPI network analysis of these protein exhibited a close correlation with ion transportation and vesicles transportation.In addition,we also found that high salinity enhanced the stabilization of cell wall.Moreover,the antioxidative enzyme(PrxR/TRX and AsA/GSH)or stress response proteins(HSP,ABA and ET signalling pathway)also have a positive role in salt tolerance.According to these results,we speculated a mechanism of vesicles-mediated salt secretion as following.The RABs and COPs are induced in Golgi,then budding to form vesicles.Meanwhile,V-ATPases and NHXs are embedding into vesicles.During the vesicular trafficking,V-ATPase will provide a electrochemical proton gradient for NHX to accumulate Na+ in vesicles.By the regulation of tubulins,PTI,V-ATPase,KIN14A and annexins,the vesicles are trafficking to plasma membrane.After that,vesicles are tethering,docking and fusing with plasma membrane by the interaction between SECs,VAMPs and KAT1/KC1.