Induced Thermotolerance and Expression of Three Key Hsp Genes (Hsp70, Hsp21, and sHsp21) and Their Roles in the High Temperature Tolerance of Agasicles hygrophila

Abstract

Thermal adaptation plays a fundamental role in the expansion and distribution of insects, and heat shock proteins (Hsps) play important roles in the temperature adaptation of various organisms. To determine the roles of Hsp genes (Hsp70, Hsp21, and sHsp21) on the high temperature tolerance of Agasicles hygrophila, we obtained complete cDNA (complementary DNA) sequences for Hsp70, Hsp21, and sHsp21 by rapid amplification of cDNA ends (RACE), analyzed their expression profiles under different high temperature treatments by quantitative reverse transcription polymerase chain reaction (RT-qPCR), and performed functional verification by RNA interference (RNAi). The open reading frames of Hsp70, Hsp21, and sHsp21 were 1940, 543, and 567 bp, encoding 650, 180, and 188 amino acids, respectively. Their molecular weights (MWs) were 71.757, 20.879, and 21.510 kDa, and the isoelectric points were 5.63, 6.45, and 6.24, respectively. Phylogenetic tree analysis showed that the Hsp70, Hsp21, and sHsp21 genes of A. hygrophila were relatively conserved in evolution. The Hsp70 and Hsp21 genes in A. hygrophila were homologous to those in Leptinotarsa decemlineata (87 and 79% similarity, respectively), and the sHsp21 gene in A. hygrophila was homologous to that in Lissorhoptrus oryzophilus (74% similarity). The amino acid polypeptide chain had highly conserved sequences of DLGGGTFD, VLVGGSTR, and GPTIEEVD. The sequence of EEVD was the characteristic motif of cytoplasmic Hsp70, and the highly conserved sequences of MALFR and MSLLP were characteristic sequences of Hsp2 and sHsp21, respectively. Relative quantitative real time PCR showed that the three Hsps could be induced by 4-h treatment at high temperatures. Significant upregulation of these Hsps was observed when the temperature was further increased. The RNAi results showed that the injection of the three Hsps' dsRNA could suppress the expression at the gene level significantly. Compared with the control group,

Transcriptome Analysis Reveals Genes of Flooding-Tolerant and Flooding-Sensitive Rapeseeds Differentially Respond to Flooding at the Germination Stage

“…Proteomic analysis performed in soybean roots supported our findings, where abundance of several heat shock proteins in response to flooding was found [ 62 ]. Studies have reported diverse roles of HSPs in regulating Rubisco activity under cold temperature stress in sunflower and barley and in chloroplast development even under abnormal growth conditions [ 63 , 64 , 65 , 66 , 67 ]. Under flooding stress, both light and carbon supplies are limited due to the slower diffusion rates in water, hence ATP synthesis is compromised, which causes carbohydrate starvation and disruption in mineral absorption and root hydraulic conductance [ 68 ].…”

Heat Shock Factor Is Involved in Regulating the Transcriptional Expression of Two Potential Hsps (AhHsp70 and AhsHsp21) and Its Role in Heat Shock Response of Agasicles hygrophila

“…Isolated RNA was stored at −80 • C until first-strand cDNA synthesis was performed using a commercial reverse transcription kit (AT341-02, TransGen Biotech, China). Full-length cDNA of AhHsp70 and AhsHsp21 of A. hygrophila was produced by RT-PCR and rapid amplification of cDNA-ends (RACE)-PCR, and the resulting sequences were submitted to NCBI (GenBank accession numbers: MN138034 and MN163038, respectively, Supplementary Figures S2, S3; Jin et al, 2020a). We used A. hygrophila transcriptome data to obtain expressed sequence tags (ESTs) that showed similarities to other insect Hsfs.…”

“…The heat shock response was first described by Ritossa (1962) who observed an induction of specific chromosome puffs on the polytene chromosomes in Drosophila melanogaster following heat or chemical treatment; this constitutes an efficient defense system against detrimental effects of protein denaturation that are predominantly associated with heat stress. Subsequent studies elucidated the nature of induced RNAs and proteins at a molecular level and led to the isolation and characterization of Hsp genes (Lindquist, 1986;Lindquist and Craig, 1988;Nover, 1991;Lü et al, 2014;Deng et al, 2018;Jin et al, 2020a). Central to the heat shock response is the induction of Hsps that effectively counteract the effects of stress by stabilizing, re-folding, or degrading denatured proteins (Parsell and Lindquist, 1993).…”

“…Previous studies reveal that the three Hsp genes play important roles for thermotolerance in A. hygrophila (Jin et al, 2020a), however, the respective molecular mechanisms affecting thermotolerance are poorly understood. In the present study, we identified a heat shock factor (AhHsf) and two of its potential downstream target genes, AhHsp70 and AhsHsp21 via gene functional verification.…”

Characterization of Inducible HSP70 Genes in an Antarctic Yeast, Glaciozyma antarctica PI12, in Response to Thermal Stress

“…At 0 • C, the expression of GaHsp70-1 was 6-fold higher than that of GaHsp70-2. The importance of HSP70 in bacteria to eukarya has been demonstrated in numerous studies [50][51][52][53][54][55]. One study showed the abundance of HSP70 in the psychrophilic bacterium Shewanella frigidimarina when cells were exposed to heat stress at 28 • C [56].…”