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Destruxin A induces and binds HSPs in Bombyx mori Bm12 cells HUANHUAN ZHANG, Weina Hu, Miaomiao XIAO, Shiyi Ou, and Hu Qiongbo J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.7b03734 • Publication Date (Web): 19 Oct 2017 Downloaded from http://pubs.acs.org on October 20, 2017
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Destruxin A induces and binds HSPs in Bombyx mori Bm12 cells Huanhuan Zhang1, Weina Hu1, Miaomiao Xiao1, Shiyi Ou2, Qiongbo Hu1*
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1. Key Laboratory of Bio-Pesticide Innovation and Application of Guangdong Province, College of
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Agriculture, South China Agricultural University, Guangzhou 510642, China.
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2. Department of Food Science and Engineering, Jinan University, Guangzhou 510632, China.
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* Correspondence:
[email protected]; Tel.: +86-20-85280308; Fax: +86-20-85280292
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Abstract: Destruxin A (DA) is a cyclodepsipeptidic mycotoxin isolated from the entomopathogenic fungus,
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Metarhizium anisopliae. It has insecticidal activity against host insect’s innate immunity system, but the
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molecular mechanism is not elucidated yet. In our previous experiment, four HSPs (heat shock proteins,
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BmHSP70-3, BmHSP75, BmHSP83 and BmHSCP) were characterized from the specific protein
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electrophoretic bands of Bombyx mori Bm12 cell line treated with DA in the test of drug affinity responsive
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target stability (DARTS), which implied that these HSPs might be kinds of DA-affinity proteins, or DA
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induces them up-regulated expression. Therefore, in current research, the interactions of DA and HSPs were
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explored through analysis of Bio-Layer Interferometry (BLI) employing FortBio OcteteQK. The expression
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levels of HSPs genes were surveyed by quantitative real-time PCR (qPCR). The results indicated that DA
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had no interactions with BmHSP70-3, BmHSP75 and BmHSP83, but had affinity to BmHSCP with a KD
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value of 88.1 µM, in BLI analysis. However, the expression levels of all HSPs genes were significantly
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up-regulated after the Bm12 cells were treated by DA. In conclusion, DA can induce the four HSPs
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expression in Bm12 cells, but DA only binds to BmHSCP. Our research provides new insights on
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understanding of the action mechanisms of destruxins.
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Keywords: destruxin; heat shock protein; molecular interaction; gene expression
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INTRODUCTION Metarhizium anisoplyae is a well-known entomopathogenic fungus distributing worldwide, which plays
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an important role in biological control of insect pests 1. Destruxins, a group of cyclo-hexadepsipeptide and
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bio-synthesized by non-ribosomal peptide synthetase, were firstly isolated from M. anisopliae 2. Chemically,
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destruxin compounds comprise five amino acids and an α-hydroxy acid, forming a cyclic hexadepsipeptid.
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To date, 39 destruxin analogs have been isolated and identified 3. Destruxin A (DA), the most common
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analogue of destruxins, is the key pathogenic factor of M. anisopliae and exhibits substantial insecticidal
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activity including contact toxicity, antifeedant and growth regulating against insects. Thus, DA was
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considered as a new lead compound of insecticides and a potential food pollution source of biological control
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agents 1, 3-4.
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There have been some experiments exploring the mechanisms of DA killing pests. It was reported that
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DA damaged the morphological characters of insect’s cells such as the B. mori Bm12, Drosophila
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melanogaster S2, Ostrinia furnacalis SYSU-Of He-C, Spodoptera exigua Sf9, etc. 5. Meanwhile, the tissues
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of midguts, visceral muscles, renal epithelia and Malpighian tubules were injured after the insects, D.
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melanogaster, Locusta migratoria and Rhodnius prolixus were treated with DA 6. However, more research
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results supported that DA damages the innate immunity of insects. In fact, DA even at a dosage of
