Molecular cloning, expression detection of IcHsp104 and IcHsp78 genes under different stress conditions in Isaria cateinannulata

Abstract:

[Background] Isaria cateinannularis is an important entomogenous fungus, and its biocontrol efficiency in the field is affected by the stress of many environmental factors. [Objective] Hsp100/ClpB is a class of ATP-dependent Hsp100 family protein, to clone two crucial Hsp100/ClpB (heat shock protein, casein lytic proteinase B) family genes IcHsp104 and IcHsp78 from Isaria cateinannulata, and to explore its role in response to different temperatures and concentrations salt stress. [Methods] The Hsp100 family genes were analyzed and screened by BLAST method using transcriptomic database obtained in early stage. RT-PCR technologies were used to verify the open reading frame (ORF) sequence of Isaria cateinannulata Hsp100 genes. The amino acid structure, phylogenetic tree, functional domain and tertiary structure of Isaria cateinannulata Hsp100 genes were analyzed by molecular bioinformatics analysis software. The expression of Isaria cateinannulata Hsp100 genes were detected by RT-qPCR technology after the fungus were treated with different temperature and salt concentration. [Results] A pair of Hsp100/ClpB genes Hsp104 and Hsp78 were screened and named IcHsp104 and IcHsp78, which encoded 923 and 805 amino acids, and their molecular weights were 103.199 kD and 88.805 kD, respectively. The relationship of both genes were close to Isaria spp., Beauveria spp. and Cordyceps spp., but it showed lower similarity between the two genes. The proteins encoded by the two genes were classic AAA+-ATPase family proteins, and the tertiary structure was mainly by alpha helix. In addition, after the treatment of high and low temperature, the expression levels of both genes were up-regulated, and increased more significantly with the extension of treatment time. Additionally, the effect of high temperature stress was significantly stronger than the low temperature stress. Then, in terms of the treatment of different concentrations of sodium chloride, the expression levels of both genes were up-regulated in the low concentration stress, while they were inhibited in the high concentration stress. [Conclusion] The IcHsp104 and IcHsp78 genes from Isaria cateinannulata play an important role in the response to external temperature and salt stress, those findings provide a theoretical basis for the further mechanism studies on Isaria cateinannulata responding to environmental stress.