Roles Of MAPK Signaling Pathway In Whitefly Defense Against Environmental Stresses

The whitefly(Bemisia tabaci) is one of the most important agricultural pests in tropics and subtropics. With the invasion of Middle East-Asia Minor1(MEAM1) and Mediterranean (MED), whitefly caused significant losses to agriculture industry. Whiteflies ingest phloem sap, which causes malnutrition of plants. What’s more, they transmit a large number of viruses, mainly the geminivirus.The global invasion of the two B. tabaci species is associated with their high capacity of surviving under various stresses, such as unsuitable host plants, extreme temperature stresses, fungal and bacterial infection. However, the molecular mechanisms underlying their remarkable adaptability are still largely unknown. Mitogen-activated protein kinase (MAPK) cascade is one of the most ancient and evolutionarily conserved signaling pathways that control a vast array of physiological processes, especially stress responses. In this study, we cloned and characterized MAPK genes in MED whiteflies. To reveal their biological function, the gene expression and functional activation of MAPKs were analyzed under various stresses. Our results are listed as follows:(1) Cloning and analyses of Bemisia tabaci MAPKWe cloned cDNA of three MAPK genes (JNK, p38and ERK) from the MED species of the B. tabaci complex, and compared their amino acid sequences among different species. The full-length cDNA of MED B. tabaci JNK consists of1176bp open reading frame (ORF) which encodes392amino acids. The full-length p38cDNA from MED contains a1077-nucleotide ORF, which encodes a protein of359amino acids. Sequence analysis revealed that MED ERK is a41.71-kDa protein with362amino acids residues. Amino acid sequence analyses indicate that these MAPK proteins are highly conserved among different species, and each has a strictly conserved dual phosphorylation motif (JNK:TPY; P38:TGY; ERK:TEY). (2) Effects of different temperatures on MAPK phosphorylationTo test the effects of temperatures on MAPK signaling pathway, we treated whitefly under six temperatures ranging from0to40℃. The activation of MAPK was analyzed by immunoblotting with antiphospho-p38,-ERK, and-JNK antibodies. After cold stress, the level of phospho-p38increased but no significant change was observed in the phosphorylation of ERK and JNK, thus suggesting that the p38might be responsible for the defense response to low temperature stress. To future analyze the changes of p38signaling pathway under low temperatures; we collected whitefly exposed to0℃for different durations. The result shows that p38phosphorylated at the third minute, indicated that p38signaling pathway can be activated rapidly at0℃.(3) Effect of host shift on MAPK activationWhiteflies were transferred from cotton (a favorable host plant) to tobacco (an unfavorable host plant). The relative expression of ERK decreased rapidly, while JNK and p38changed little. Phosphorylation of p38, ERK and JNK had no significant change, indicated that host shift didn’t activate the MAPK signaling pathway in MED whiteflies.(4) Effect of fungal and bacterial infection on MAPK activationTo find the role of MAPK in B. tabaci immune response, the effect of fungus (Beauveria bassiana) and bacterium(Pseudomonas aeruginosa) infection on MAPK were examined. The gene expression and phosphorylation levels of MAPK showed no significant difference when treated with P. aeruginosa, suggesting that MAPK pathway has little or no involvement in the response to bacterial infection. However, we found that the amount of phosphorylated JNK increased significantly after fungal infection, while there is no obvious change for phosphorylated p38and ERK. Our results indicate that the whitefly JNK plays an important role in whitefly’s immune responses to fungal infection.Overall, we find that the MAPK signaling pathway takes part in the responses of whitefly to stresses. While host shift and bacterial infection have little to do with MAPK pathway, low temperature and fungus can activate MAPK pathway in MED. Among these, p38was specifically activated by low temperature, and JNK signaling was triggered by fungal infection. All these indicated that MAPK signaling pathway plays an important role in the adaptation of whitefly to various environmental stresses.