Leitsch David, Mbouaka Alvie Loufouma, Köhsler Martina, Müller Norbert, Walochnik Julia
Institute of Specific Prophylaxis and Tropical Medicine, Medical University of Vienna, Kinderspitalgasse 15, 1090, Vienna, Austria.
Institute of Parasitology, Vetsuisse Faculty, University of Bern, Länggassstrasse 122, 3012, Bern, Switzerland.
Cell Mol Life Sci. 2021 Apr;78(7):3673-3689. doi: 10.1007/s00018-021-03786-x. Epub 2021 Feb 18.
The free-living amoeba Acanthamoeba castellanii occurs worldwide in soil and water and feeds on bacteria and other microorganisms. It is, however, also a facultative parasite and can cause serious infections in humans. The annotated genome of A. castellanii (strain Neff) suggests the presence of two different thioredoxin reductases (TrxR), of which one is of the small bacterial type and the other of the large vertebrate type. This combination is highly unusual. Similar to vertebrate TrxRases, the gene coding for the large TrxR in A. castellanii contains a UGA stop codon at the C-terminal active site, suggesting the presence of selenocysteine. We characterized the thioredoxin system in A. castellanii in conjunction with glutathione reductase (GR), to obtain a more complete understanding of the redox system in A. castellanii and the roles of its components in the response to oxidative stress. Both TrxRases localize to the cytoplasm, whereas GR localizes to the cytoplasm and the large organelle fraction. We could only identify one thioredoxin (Trx-1) to be indeed reduced by one of the TrxRases, i.e., by the small TrxR. This thioredoxin, in turn, could reduce one of the two peroxiredoxins tested and also methionine sulfoxide reductase A (MsrA). Upon exposure to hydrogen peroxide and diamide, only the small TrxR was upregulated in expression at the mRNA and protein levels, but not the large TrxR. Our results show that the small TrxR is involved in the A. castellanii's response to oxidative stress. The role of the large TrxR, however, remains elusive.
自由生活的阿米巴原虫卡氏棘阿米巴在全球范围内的土壤和水中均有发现，以细菌和其他微生物为食。然而，它也是一种兼性寄生虫，可导致人类严重感染。卡氏棘阿米巴（菌株Neff）的注释基因组表明存在两种不同的硫氧还蛋白还原酶（TrxR），其中一种是小细菌类型，另一种是大脊椎动物类型。这种组合非常不寻常。与脊椎动物的TrxRases类似，卡氏棘阿米巴中编码大TrxR的基因在C末端活性位点含有UGA终止密码子，表明存在硒代半胱氨酸。我们结合谷胱甘肽还原酶（GR）对卡氏棘阿米巴的硫氧还蛋白系统进行了表征，以更全面地了解卡氏棘阿米巴的氧化还原系统及其成分在应对氧化应激中的作用。两种TrxRases均定位于细胞质，而GR定位于细胞质和大细胞器部分。我们只能鉴定出一种硫氧还蛋白（Trx-1）确实被其中一种TrxRases还原，即被小TrxR还原。反过来，这种硫氧还蛋白可以还原所测试的两种过氧化物酶之一以及甲硫氨酸亚砜还原酶A（MsrA）。在暴露于过氧化氢和二酰胺后，只有小TrxR在mRNA和蛋白质水平上的表达上调，而大TrxR则没有。我们的结果表明，小TrxR参与了卡氏棘阿米巴对氧化应激的反应。然而，大TrxR的作用仍然难以捉摸。
