title: Evidence for involvement of cytosolic thioredoxin peroxidase in the excessive resistance of Sf9 Lepidopteran insect cells against radiation-induced apoptosis.
abstract: Lepidopteran insect cells display 50-100 times higher radioresistance compared to human cells, and reportedly have more efficient antioxidant system that can significantly reduce radiation-induced oxidative stress and cell death. However, the antioxidant mechanisms that contribute substantially to this excessive resistance still need to be understood thoroughly. In this study, we investigated the role of thioredoxin peroxidase (TPx) in high-dose γ-radiation response of Sf9 cell line derived from Spodoptera frugiperda, the Fall armyworm. We identified a TPx orthologue (Sf-TPx) in Spodoptera system, with primarily cytosolic localization. Gamma-irradiation at 500 Gy dose significantly up-regulated Sf-TPx, while higher doses (1000 Gy-2000 Gy) had no such effect. G2/M checkpoint induced following 500 Gy was associated with transition of Sf-TPx decamer into enzymatically active dimer. Same effect was observed during G2/M block induced by 5 nM okadaic acid or 10 µM CDK1 (cycline dependent kinase-1) inhibitor roscovitine, thus indicating that radiation-induced Sf-TPx activity is mediated by CDKs. Accumulation of TPx dimer form during G2/M checkpoint might favour higher peroxidase activity facilitating efficient survival at this dose. Confirming this, higher lethal doses (1000 Gy-2000 Gy) caused significantly less accumulation of dimer form and induced dose-dependent apoptosis. A ∼50% knock-down of Sf-TPx by siRNA caused remarkable increase in radiation-induced ROS as well as caspase-3 dependent radiation-induced apoptosis, clearly implying TPx role in the radioresistance of Sf9 cells. Quite importantly, our study demonstrates for the first time that thioredoxin peroxidase contributes significantly in the radioresistance of Lepidopteran Sf9 insect cells, especially in their exemplary resistance against radiation-induced apoptosis. This is an important insight into the antioxidant mechanisms existing in this highly stress-resistant model cell system.
authors.affiliation: Natural Radiation Response Mechanisms Group, Division of Radiation Biosciences, Institute of Nuclear Medicine and Allied Sciences, Delhi, India.
authors.forename: Shashank
authors.initials: S
authors.lastname: Hambarde
authors.forename: Vijaypal
authors.initials: V
authors.lastname: Singh
authors.forename: Sudhir
authors.initials: S
authors.lastname: Chandna
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics* / Q000528:radiation effects*; D059447:Cell Cycle Checkpoints / Q000528:radiation effects; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D005047:Etoposide / Q000494:pharmacology; D005720:Gamma Rays; D005786:Gene Expression Regulation / Q000528:radiation effects; D055785:Gene Knockdown Techniques; D007915:Lepidoptera; D008969:Molecular Sequence Data; D018384:Oxidative Stress / Q000235:genetics / Q000528:radiation effects; D054464:Peroxiredoxins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D055503:Protein Multimerization; D011836:Radiation Tolerance / Q000235:genetics*; D017382:Reactive Oxygen Species / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: D017382:Reactive Oxygen Species; D005047:Etoposide; D054464:Peroxiredoxins