Study Of The Function And Regulation Mechanism Of Small Heat Shock Protein In The Immune Response Of Antheraea Pernyi

Small heat shock proteins(sHSPs) can regulate protein folding and protect cells from stress. To investigate the role of sHSPs in the silk-producing insect Antheraea pernyi(A.pernyi; Lepidoptera: Saturniidae), cDNA encoding HSP20.8 and 21.4 in A. pernyi, termed Ap-sHSP20.8, was identified as a 564 bp ORF. The translated amino acid sequence encoded 187 residues with a calculated molecular mass of 20.8 and 21.4 kDa and an isoelectronic point(pI) of 5.98 and 5.79; the sequence showed homology to sHSP chaperone proteins from other insects. Ap-sHSP20.8 and Ap-sHSP21.4 mRNA transcript expression was abundant in the midgut and fat body and found to be both constitutive and inducible by infectious stimuli.Ap-sHSP20.8 and Ap-sHSP21.4 were highly expressed in fat body and up-regulated in fat body and midgut of A. pernyi challenged with Escherichia coli, Beauveria bassiana and nuclear polyhedrosis virus(NPV), which was determined by quantitative real-time PCR and/or Western blot. Meanwhile, knock down of Ap-sHSP21.4 with dsRNA result in the decrease at the expression levels of several immune response-related genes(defensin,Dopa decarboxylase, Toll1, lysozyme and Kazal-type serine protease inhibitor).Additionally, to investigate the mechanism of immunity function of Ap-sHSPs, we analyzed the impact of eicosanoid biosynthesis on the expression of Ap-sHSP21.4 response to NPV was determined using qPCR and/or Western blot technology. And we found that inhibitors of eicosanoid biosynthesis significantly suppress Ap-sHSP20.8 and Ap-HSP21.4expression upon NPV challenge, while arachidonic acid could stimulate the up-regulation expression of Ap-sHSP20.8 and Ap-HSP21.4.On the other hand, after heat shock injury, a group of proteins that regulate protein-folding processes are synthesised to prevent infection. Caspase is an enzyme responsible for the execution of stress-induced apoptosis. Heat shock proteins(Hsp) are capable of modulating caspase activity. Therefore, we studied the effects of heat shock preconditioning on Ap-sHSP20.8 and on the final apoptotic effector enzyme caspase-1 to clarify whether these effects were modulated in the fat bodies of Antheraea pernyi injured by heat shock. We concluded that heat shock and eicosanoid biosynthesis inhibitors might be utilised to simultaneously increase Hsp 20.8 synthesis and to decrease caspase-1 activity.Modifications of the eicosanoid pathways might also be used to reduce caspase-1 activity under hyperthermic. These results will shed light in the understanding of the pathogen-host interaction in A.pernyi.