Many proteins require assistance from molecular chaperones at various stages to attain correctly folded states and functional conformations during protein synthesis.  In this study, the gene encoding T-complex polypeptide 1 (TCP-1), which belongs to the heat shock protein 60 (HSP60) family, was isolated and characterized from the rice stem borer, Chilo suppressalis, by RACE and qPCR, respectively.  The full-length cDNA of Tcp-1 was 2 144 bp and encoded a 1 635-bp ORF; the deduced translational product contained 545 amino acids with 5´- and 3´-UTRs and an isoelectric point of 5.29.  Cluster analysis confirmed that the deduced amino acid sequence shared high identity (60–99%) with TCP-1 from other insects.  To investigate Tcp-1 expression in response to abiotic stress, qPCR was used to analyze expression levels of Tcp-1 mRNA in C. suppressalis larvae exposed to temperatures ranging from –11 to 43°C.  With respect to heat shock, Tcp-1 expression was higher than the control after a 2-h exposure to 30 and 36°C and declined at 39 and 43°C.  Difference in Tcp-1 expression was observed at temperatures ranging from –11 to 27°C.  qPCR analyses revealed that Tcp-1 expression was the highest in hindgut tissue as compared to heads, epidermis, fat body, foregut, midgut, and malpighian tubules.  Our results indicated that Tcp-1 expression was differentially expressed in C. suppressalis tissues, and was impacted by temperature stress.

Heat shock protein 70 (HSP70) is one of the most important members in the heat shock protein family, and plays important roles in the thermotolerance of insect.  To explore the molecular mechanism of thermotolerance of Frankliniella occidentalis adults, the difference in the expression of HSP70s in F. occidentalis male or female adults under the thermal stress was studied under the laboratory conditions.  Two full length cDNAs of HSP70s gene (Fohsc704 and Fohsc705) were cloned from F. occidentalis by using RT-PCR and RACE.  The genomic sequence was demonstrated by genomic validation, and the position and size of the intron were analyzed by sequence analysis of cDNA.  Real-time PCR was used to analyze the HSP70 expression patterns.  The cDNA of Fohsc704 and Fohsc705 possessed 2 073 and 1 476 bp which encoded 690 and 491 amino acids (aa) with a calculated molecular weight of 75 and 54 kDa, respectively.  Four introns in Fohsc704 and six introns in Fohsc705 protein were found.  However, the HSP70 protein sequences in our study were ended with EKKN and GIFL, which were different from the reported FoHSP70s.  Various expression patterns of Fohsc704 and Fohsc705 were found in both genders of F. occidentalis under thermal stress.  The expression of Fohsc704 and Fohsc705 reached to the highest level at –12 and –8°C in male adults, respectively, and Fohsc705 expressed the highest level at 33°C in female adults.  In conclusion, HSP70s of F. occidentalis in our study are novel heat shock proteins.  There were difference in expression patterns of the two hsc70s in genders of F. occidentalis, and the two HSP70s play important roles in the thermotolerance of F. occidentalis.  

Five genes encoding heat shock proteins (HSPs), Cchsp40, Cchsp60, Cchsp70, Cchsc70 and Cchsp90, were cloned and sequenced from Cotesia chilonis using RT-PCR and RACE.  The cDNA sequences of Cchsp40, Cchsp60, Cchsp70, Cchsc70 and Cchsp90 were 1 265, 2 551, 2 094, 2 297 and 2 635 bp in length, respectively, with a molecular weight (MW) of 39.1, 60.6, 71.45, 70.19 and 82.92 kDa, respectively.  The predicted amino acid sequences of these proteins showed high similarities with published HSPs of other insects in Hymenoptera.  Analysis of genomic DNAs indicated that Cchsp40, Cchsp60, Cchsp70, Cchsc70 and Cchsp90 lacked introns, but Cchsc70 contained an intron.  The results also suggested that CcHSP40 in C. chilonis was the Type II HSP40, CcHSP60 was a member of the mitochondrial HSP60 family, and CcHSP90 was a part of cytoplasmic HSP90A family.  Expression patterns varied in the five Cchsps in response to temperature.  Expression of Cchsp40 and Cchsp60 was induced significantly by cold but not heat stress.  Cchsp70 and Cchsc70 showed similar response to the thermal stress and could be induced by both cold and heat, but their expression levels were consistently lower than that of Cchsp40 and Cchsp60.  Cchsp90 could be induced by heat stress and mild cold, but not cold stress.  In addition, the results demonstrated Cchsc70 might be constitutive and inducible protein that was expressed during normal cell functioning and also up-regulated in response to stressful stimuli while Cchsp70 was solely inducible protein induced by temperature changes.  Overall, results generated from this study could significantly advance the understanding of Cchsps in response to temperature and provide important biological information for C. chilonis insects that reared under different temperatures.