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The plant defensin NaD1 induces tumor cell death via a non-apoptotic, membranolytic process.

ABSTRACT: Cationic anti-microbial peptides (CAPs) have an important role in host innate defense against pathogens such as bacteria and fungi. Many CAPs including defensins also exhibit selective cytotoxic activity towards mammalian cells via both apoptotic and non-apoptotic processes, and are being investigated as potential anticancer agents. The anti-fungal plant defensin from ornamental tobacco, Nicotiana alata Defensin 1 (NaD1), was recently shown to induce necrotic-like cell death in a number of tumor cell types within 30?min of treatment, at a concentration of 10??M. NaD1-mediated cell killing within these experimental parameters has been shown to occur via binding to the plasma membrane phosphatidylinositol 4,5-bisphosphate (PIP2) in target cells to facilitate membrane destabilization and subsequent lysis. Whether NaD1 is also capable of inducing apoptosis in tumor cells has not been reported previously. In this study, treatment of MM170 (melanoma) and Jurkat T (leukemia) cells with subacute (<10??M) concentrations of NaD1 over 6-24?h was investigated to determine whether NaD1 could induce cell death via apoptosis. At subacute concentrations, NaD1 did not efficiently induce membrane permeabilization within 30?min, but markedly reduced cell viability over 24?h. In contrast to other CAPs that have been shown to induce apoptosis through caspase activation, dying cells were not sensitive to a pancaspase inhibitor nor did they display caspase activity or DNA fragmentation over the 24?h treatment time. Furthermore, over the 24?h period, cells exhibited necrotic phenotypes and succumbed to membrane permeabilization. These results indicate that the cytotoxic mechanism of NaD1 at subacute concentrations is membranolytic rather than apoptotic and is also likely to be mediated through a PIP2-targeting cell lytic pathway.

SUBMITTER: Baxter AA

PROVIDER: S-EPMC5253418 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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The plant defensin NaD1 induces tumor cell death via a non-apoptotic, membranolytic process.

Baxter Amy A AA Poon Ivan Kh IK Hulett Mark D MD

Cell death discovery 20170123

Cationic anti-microbial peptides (CAPs) have an important role in host innate defense against pathogens such as bacteria and fungi. Many CAPs including defensins also exhibit selective cytotoxic activity towards mammalian cells via both apoptotic and non-apoptotic processes, and are being investigated as potential anticancer agents. The anti-fungal plant defensin from ornamental tobacco, <i>Nicotiana alata</i> Defensin 1 (NaD1), was recently shown to induce necrotic-like cell death in a number o ...[more]

PMID: 28179997

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Project description:Defensins are a large family of small, cationic, cysteine-rich proteins that are part of the defense arsenal that plants use for protection against potentially damaging fungal infections. The plant defensin NaD1 from Nicotiana alata is a potent antifungal protein that inhibits growth and kills a variety of fungal pathogens that affect both plant and animal (human) hosts. Some serine protease inhibitors have also been reported to be antifungal molecules, while others have no inhibitory activity against fungi. Here we describe the synergistic activity of the plant defensin NaD1 with a selection of serine protease inhibitors against the plant pathogens Fusarium graminearum and Colletotrichum graminicola and the animal pathogen Candida albicans. The synergistic activity was not related to the protease inhibitory activity of these molecules but may arise from activation of fungal stress response pathways. The bovine pancreatic trypsin inhibitor (BPTI) displayed the most synergy with NaD1. BPTI also acted synergistically with several other antifungal molecules. The observation that NaD1 acts synergistically with protease inhibitors provides the foundation for the design of transgenic plants with improved resistance to fungal disease. It also supports the possibility of naturally occurring accessory factors that function to enhance the activity of innate immunity peptides in biological systems. IMPORTANCE This work describes the increased activity of a natural antifungal peptide in the presence of another antifungal peptide from a different family. This is termed antifungal synergy. Synergy is important for decreasing the amount of antifungal molecule needed to control the disease. Traditionally, naturally occurring antifungal molecules are assayed in isolation. Identification of synergistic interactions between antifungal peptides means that their activities in a complex biological system are likely to be different from what we observe when examining them individually. This study identified synergy between an antifungal peptide and a group of peptides that do not affect fungal growth in vitro. This provides the foundation for generation of transgenic plants with increased resistance to fungal disease and identification of antifungal accessory factors that enhance the activity of innate immune molecules but do not have an antifungal effect on their own.

Project description:Some symbiotic bacteria cause remarkable reproductive phenotypes like cytoplasmic incompatibility and male-killing in their host insects. Molecular and cellular mechanisms underlying these symbiont-induced reproductive pathologies are of great interest but poorly understood. In this study, Drosophila melanogaster and its native Spiroplasma symbiont strain MSRO were investigated as to how the host's molecular, cellular and morphogenetic pathways are involved in the symbiont-induced male-killing during embryogenesis. TUNEL (terminal deoxynucleotidyl transferase dUTP nick end labeling) staining, anti-cleaved-Caspase-3 antibody staining, and apoptosis-deficient mutant analysis unequivocally demonstrated that the host's apoptotic pathway is involved in Spiroplasma-induced male-specific embryonic cell death. Double-staining with TUNEL and an antibody recognizing epidermal marker showed that embryonic epithelium is the main target of Spiroplasma-induced male-specific apoptosis. Immunostaining with antibodies against markers of differentiated and precursor neural cells visualized severe neural defects specifically in Spiroplasma-infected male embryos as reported in previous studies. However, few TUNEL signals were detected in the degenerate nervous tissues of male embryos, and the Spiroplasma-induced neural defects in male embryos were not suppressed in an apoptosis-deficient host mutant. These results suggest the possibility that the apoptosis-dependent epidermal cell death and the apoptosis-independent neural malformation may represent different mechanisms underlying the Spiroplasma-induced male-killing. Despite the male-specific progressive embryonic abnormality, Spiroplasma titers remained almost constant throughout the observed stages of embryonic development and across male and female embryos. Strikingly, a few Spiroplasma-infected embryos exhibited gynandromorphism, wherein apoptotic cell death was restricted to male cells. These observations suggest that neither quantity nor proliferation of Spiroplasma cells but some Spiroplasma-derived factor(s) may be responsible for the expression of the male-killing phenotype.

Dataset Information

The plant defensin NaD1 induces tumor cell death via a non-apoptotic, membranolytic process.

Publications

The plant defensin NaD1 induces tumor cell death via a non-apoptotic, membranolytic process.

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