Abstract
Our previous study showed that the srfABC operon, which was originally identified in Salmonella enterica as an SsrB-regulated operon clustered with the flagellar class 2 operon, exhibited significant cytotoxicity against insect midgut CF-203 cells and injectable insecticidal activity against Helicoverpa armigera larvae. The srfABC operon was widely distributed among bacteria, which raises the question of their biological roles in different species. In this study, we investigated the cytotoxic effect of SrfABC toxin on mammalian cell lines. When simultaneously expressed in the Escherichia coli cytoplasm, SrfABC exhibited cytotoxicity against all tested mammalian cancer cell lines (B16, 4T-1, Hep-3B, and HeLa) in a dose-dependent manner. Intracellular expression of SrfA–FLAG, SrfB–FLAG, or SrfC–FLAG also resulted in inhibition of proliferation and apoptosis on HeLa cells. When incubated with HeLa cells separately, SrfA, SrfB, and SrfC proteins alone could enter HeLa cells, then induce apoptosis and cytotoxicity. SrfC protein shifts its localization from cytoplasm to nucleus with the aid of SrfA and/or SrfB protein. Although SrfA, SrfB, and SrfC proteins alone exhibited a cytotoxic effect against HeLa cells, all three components were essential for the full cytotoxicity. Native PAGE and co-immunoprecipitation assay demonstrated that SrfA, SrfB, and SrfC proteins could interact with each other and form a heteromeric complex.
Highlights
Xenorhabdus spp. are symbionts of entomopathogenic nematodes (EPN) of the genus Steinernema, where they reside in a special vesicle in infective juveniles (IJ) [1]
B16, murine mammary carcinoma cell line 4T-1, human hepatoma cell line Hep-3B, and human cervical carcinoma cell line HeLa were treated with various amounts of the lysates of GB05/srfABC
As our previous study showed that each component of SrfABC exhibited cytotoxicity against insect CF-203 cells, we explored whether SrfA, SrfB, or SrfC alone could cause toxic effects on HeLa cells
Summary
Xenorhabdus spp. are symbionts of entomopathogenic nematodes (EPN) of the genus Steinernema, where they reside in a special vesicle in infective juveniles (IJ) [1]. A wide range of toxins with oral and/or injectable insecticidal activities have been identified and characterized in Xenorhabdus spp., such as Xpts (Xenorhabdus protein toxins) [4], XhlA (cell surface-associated hemolysin) [5], XaxAB (Xenorhabdus α-xenorhabdolysin) [6], Txp (40 kDa toxin from Xenorhabdus and Photorhabdus) [7], etc. SrfA, SrfB, and SrfC alone exhibited cytotoxicity against CF-203 cells, all three components of SrfABC toxin were essential for full cytotoxicity. SrfABC toxin exhibits highly injectable insecticidal activity against Helicoverpa armigera larvae [9]. The class B and C proteins (XptB1 and XptC1) form a binary complex and strongly bind to the tetrameric XptA2 to form the complete and fully active toxin complex [19]. We determined the cytotoxic effect of SrfABC toxin on mammalian cell lines, and its potential mechanism was investigated
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