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RNA-Seq analysis of Polyrhachis vicina Roger and insights into the heat shock protein 90 and 70 families.

ABSTRACT: The heat shock protein 90 (Hsp90) and heat shock cognate proteins (Hsc70) have been identified as chaperones of the ecdysone receptor (EcR)/ultraspiracle protein (USP) heterocomplex. However, little is known about the status of Hsp90 and Hsc70 in Polyrhachis vicina Roger. Here, we sequenced the transcriptomes of adult ants in P. vicina for the first time. Clean reads in female, male, and worker ants were annotated into 40,147 transcripts, and 37,488, 28,300, and 33,638 unigenes were assembled in female, male, and worker ants, respectively. According to RPKM, the numbers of differentially expressed genes between female and male ants, between female and worker ants, and between male and worker ants and the common differentially expressed genes were 12,657, 21,630, 15,112 and 3704, respectively. These results reveal that caste differentiation, caste specificity formation, and social divisions of P. vicina ants may be due to gene expression differences. Moreover, PvEcR and PvUSP were also detected as differentially expressed genes in the ants; specifically, PvUSP expression was higher than PvEcR expression in all castes. We speculate that PvUSP may have a role similar to that of juvenile hormone receptor. Four identified PvHsp90 family members and 23 identified PvHsp70 family members were found in the ants, and 2 PvHsp90 genes and 8 PvHsp70 genes were analyzed by qRT-PCR. Among those genes, the expression of 2 PvHsp90 genes and 5 PvHsp70 genes coincided with the expression profiles of PvEcR and PvUSP, which suggest that the characterization of PvHsp90 and PvHsc70 may be as EcR/USP molecular chaperones in P. vicina.

SUBMITTER: Zhang J

PROVIDER: S-EPMC6363624 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundThe tick Dermacentor silvarum Olenev (Acari: Ixodidae) is a vital vector tick species mainly distributed in the north of China and overwinters in the unfed adult stage. The knowledge of the mechanism that underlies its molecular adaptation against cold is limited. In the present study, genes of hsp70 and hsp90 cDNA, named Dshsp70 and Dshsp90, and tubulin were cloned and characterized from D. silvarum, and their functions in cold stress were further evaluated.MethodsThe genome of the heat shock proteins and tubulin of D. silvarum were sequenced and analyzed using bioinformatics methods. Each group of 20 ticks were injected in triplicate with Dshsp90-, Dshsp70-, and tubulin-derived dsRNA, whereas the control group was injected with GFP dsRNA. Then, the total RNA was extracted and cDNA was synthesized and subjected to RT-qPCR. After the confirmation of knockdown, the ticks were incubated for 24 h and were exposed to - 20 °C lethal temperature (LT50), and then the mortality was calculated.ResultsResults indicated that Dshsp70 and Dshsp90 contained an open reading frame of 345 and 2190 nucleotides that encoded 114 and 729 amino acid residues, respectively. The transcript Dshsp70 showed 90% similarity with that identified from Dermacentor variabilis, whereas Dshsp90 showed 85% similarity with that identified from Ixodes scapularis. Multiple sequence alignment indicates that the deduced amino acid sequences of D. silvarum Hsp90, Hsp70, and tubulin show very high sequence identity to their corresponding sequences in other species. Hsp90 and Hsp70 display highly conserved and signature amino acid sequences with well-conserved MEEVD motif at the C-terminal in Hsp90 and a variable C-terminal region with a V/IEEVD-motif in Hsp70 that bind to numerous co-chaperones. RNA interference revealed that the mortality of D. silvarum was significantly increased after injection of dsRNA of Dshsp70 (P = 0.0298) and tubulin (P = 0.0448), whereas no significant increases were observed after the interference of Dshsp90 (P = 0.0709).ConclusionsThe above results suggested that Dshsp70 and tubulin play an essential role in the low-temperature adaptation of ticks. The results of this study can contribute to the understanding of the survival and acclimatization of overwintering ticks.

Project description:BackgroundHeat shock proteins (HSPs) are found in all prokaryotes and most compartments of eukaryotic cells. Members of the HSP family mediate immune responses to tissue damage or cellular stress. However, little is known about the immune response induced by the oriental liver fluke, Clonorchis sinensis, even though this organism is carcinogenic to humans. We address this issue in the present study in mouse bone marrow dendritic cells (mBMDCs), using recombinant HSP70 and 90 from C. sinensis (rCsHSP70 and rCsHSP90).MethodsrCsHSP70 and rCsHSP90 were produced in an E. coli system. Purified recombinant proteins were treated in BMDCs isolated from C57BL/6 mice. T cells were isolated from Balb/c mice and co-cultured with activated mBMDCs. Expression of surface molecules was measured by flow cytometry and cytokine secretion was quantified using ELISA. C57BL/6 mice were divided into four groups, including peptide alone, peptide/Freund's adjuvant, peptide/CsHSP70, peptide/CsHSP90, and were immunized intraperitoneally three times. Two weeks after final immunization, antibodies against peptide were measured using ELISA.ResultsBoth proteins induced a dose-dependent upregulation in major histocompatibility complex and co-stimulatory molecule expression and increased secretion of pro-inflammatory cytokines including interleukin (IL)-1β, -6, and -12p70 and tumor necrosis factor-α in mBMDCs. Furthermore, when allogenic T cells were incubated with mBMDCs activated by rCsHSP70 and rCsHSP90, the helper T cell (Th)1 cytokine interferon-γ was up-regulated whereas the level of the Th2 cytokine IL-4 was unchanged. These results indicate that rCsHSPs predominantly induce a Th1 response. Over and above these results, we also demonstrated that the production of peptide-specific antibodies can be activated after immunization via in vitro peptide binding with rCsHSP70 or rCsHSP90.ConclusionThis study showed for the first time that the HSP or HSP/peptide complexes of C. sinensis could be considered as a more effective vaccine against C. sinensis infection as results of the activator of host immune response as well as the adjuvant for antigenic peptide conjugate to induce peptide-specific antibody response in mice.

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RNA-Seq analysis of Polyrhachis vicina Roger and insights into the heat shock protein 90 and 70 families.

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