Cloning and identification of grass carp transcription factor HSF1 and its characterization involving the production of
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Cloning and identification of grass carp transcription factor HSF1 and its characterization involving the production of fish HSP70 Xinrui Yang & Yajun Gao & Minghui Zhao & Xinyan Wang & Hong Zhou & Anying Zhang
Received: 18 January 2020 / Accepted: 18 June 2020 # Springer Nature B.V. 2020
Abstract In mammals, heat shock transcription factor 1 (HSF1) is well documented as the critical transcript factor to regulate heat shock protein 70 (HSP70) expression under different stresses, such as heat shock or bacterial infection. In fish, Hsf1 responses to physiological and environmental stresses and regulates Hsp70 expression under thermal exposure. However, the functional role of Hsf1 in Hsp70 production is still elusive under bacterial infection. In the present study, a coding sequence of grass carp hsf1 (gchsf1) gene was cloned and identified. Using Ctenopharyngodon idellus kidney (CIK) cells as the model, we found that lipopolysaccharide (LPS) exerted stimulatory effects on the expression of grass carp hsp70 (gchsp70) and hsf1, implying possible relationship of Hsp70 and Hsf1 under immune stimulation in fish. To validate the hypothesis, overexpression of gcHsf1 was performed in CIK cells, and the effects of overexpressing gcHsf1 on the expression of gcHsp70 in the absence or presence of LPS were
examined. Results showed that LPS significantly upregulated the transcription and protein synthesis of gcHsp70, and these stimulatory effects were further amplified when overexpression of gcHsf1 was performed. Furthermore, luciferase reporter assays in CIK cells revealed that both overexpression of Hsf1 and LPS upregulated gchsp70 transcription, and their combined treatment further enhanced the gchsp70 promoter activity. Moreover, the regions responsive to these treatments were mapped to the promoter of gchsp70. Besides transcriptional level and cellular protein contents, gcHsp70 secretion was measured by competitive ELISA, uncovering that gcHsf1 enhanced the release of gcHsp70 induced by LPS in the same cells. These data not only demonstrated the enhancement of Hsf1 in Hsp70 production but also initially revealed the involvement of Hsf1-Hsp70 axis in mediating inflammatory response in fish. Keywords Grass carp . Heat shock factor 1 . Heat shock protein 70 . CIK cell . LPS
Electronic supplementary material The online version of this article (https://doi.org/10.1007/s10695-020-00842-4) contains supplementary material, which is available to authorized users. X. Yang : Y. Gao : M. Zhao : X. Wang : H. Zhou : A. Zhang (*) School of Life Science and Technology, University of Electronic Science and Technology of China, Chengdu 610054, People’s Republic of China e-mail: [email protected] X. Yang Department of Biology, Lawrence University, Appleton, WI, USA
Introduction Heat shock proteins (HSPs) are molecular chaperones that support protein folding and transport in response to a wide variety of physiological and environmental stresses such as heat shock or microbial infections (Kiang and Tsokos 1998; Liberek et al. 2008), among
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