Abstract
Protein biosynthesis is fundamental to cellular life and requires the efficient functioning of the translational machinery. At the center of this machinery is the ribosome, a ribonucleoprotein complex that depends heavily on Mg2+ for structure. Recent work has indicated that other metal cations can substitute for Mg2+, raising questions about the role different metals may play in the maintenance of the ribosome under oxidative stress conditions. Here, we assess ribosomal integrity following oxidative stress both in vitro and in cells to elucidate details of the interactions between Fe2+ and the ribosome and identify Mn2+ as a factor capable of attenuating oxidant-induced Fe2+-mediated degradation of rRNA. We report that Fe2+ promotes degradation of all rRNA species of the yeast ribosome and that it is bound directly to RNA molecules. Furthermore, we demonstrate that Mn2+ competes with Fe2+ for rRNA-binding sites and that protection of ribosomes from Fe2+-mediated rRNA hydrolysis correlates with the restoration of cell viability. Our data, therefore, suggest a relationship between these two transition metals in controlling ribosome stability under oxidative stress.
Highlights
All organisms require a number of metal elements in trace amounts, with manganese, iron, copper, zinc, selenium, cobalt, and molybdenum all considered essential for plants and animals, whereas larger amounts of magnesium, calcium, potassium, and sodium are required [1]
We have demonstrated in a previous study [41] that Fe21 has a direct role in cleaving the sugar-phosphate backbone of rRNA under oxidative stress conditions
Cellular lysates from exponentially growing WT cells were centrifuged through a sucrose cushion to obtain purified ribosomes, which were treated with ascorbic acid alone, ascorbic acid in combination with Fe(NH4)2(SO4)2, or ascorbic acid with both Fe(NH4)2(SO4)2 and the iron chelator deferoxamine (DFO)
Summary
All organisms require a number of metal elements in trace amounts, with manganese, iron, copper, zinc, selenium, cobalt, and molybdenum all considered essential for plants and animals, whereas larger amounts of magnesium, calcium, potassium, and sodium are required [1]. Divalent metal cations have a long-established involvement in biomolecules including stabilizing structures and participating in the active sites of enzymes operating across a vast catalytic range [2,3,4]. Metal cations have essential stabilizing roles in structures including phospholipid bilayers [13] and the ribosome [14]. The ribosome is a massive ribonucleoprotein complex that is dependent on Mg21 [14] to fold and maintain stability because of the negative charge of the RNA backbone, and as many as 200 Mg21 ions can be associated with just the large subunit, coordinated in six distinct geometries [23,24,25]. Abundant Mn21 was oxidized and precipitated out, resulting in vast amounts of both transition metals in sedimentary rocks
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