Iron-dependent cleavage of ribosomal RNA during oxidative stress in the yeast Saccharomyces cerevisiae

Abstract

Stress-induced strand breaks in rRNA have been observed in many organisms, but the mechanisms by which they originate are not well understood. Here, we show that a chemical rather than enzymatic mechanism initiates rRNA cleavages during oxidative stress in yeast (Saccharomyces cerevisiae). We used cells lacking the mitochondrial glutaredoxin Grx5 to demonstrate that oxidant-induced cleavage formation in 25S rRNA correlates with intracellular iron levels. Sequestering free iron by a chemical or genetic means decreased the extent of rRNA degradation and relieved the hypersensitivity of grx5 cells to the oxidants. Importantly, subjecting purified ribosomes to an in vitro iron/ascorbate reaction precisely recapitulated the 25S rRNA cleavage pattern observed in cells, indicating that redox activity of the ribosome-bound iron is responsible for the strand breaks in the rRNA. In summary, our findings provide evidence that oxidative stress-associated rRNA cleavages can occur through rRNA strand scission by redox-active, ribosomebound iron that potentially promotes Fenton reaction-induced hydroxyl radical production, implicating intracellular iron as a key determinant of the effects of oxidative stress on ribosomes. We propose that iron binding to specific ribosome elements primes rRNA for cleavages that may play a role in redox-sensitive tuning of the ribosome function in stressed cells.

Cutting in-line with iron: ribosomal function and non-oxidative RNA cleavage

“…Thus, aerobic cells tightly regulate intracellular Fe to maintain low Fe 2+ levels in the cytosol (58), minimizing Fe 2+ availability for incorporation into ribosomes. We detect only around 1-2 Fe 2+ per ribosome in aerobic growth conditions, consistent to what was previously seen for yeast ribosomes (59). In anoxic conditions, Fe 2+ is more bioavailable and is less harmful because anaerobic growth generates fewer reactive oxygen species and there is no threat from O 2 diffusion into cells.…”

“…Moreover, iron likely decreases ribosome longevity within the cell, as we have illustrated the potency of Fe 2+ in inducing rRNA cleavage. In fact, rRNA cleavage events commonly linked to Fe 2+ oxidation, such as in the human ribosome factoring in Alzheimer’s disease (69), or in Saccharomyces cerevisiae rRNA where cleavage is tied to downstream oxidative stress response (59) could be in some measure attributable to Fe 2+ in-line cleavage. The rapid Fe 2+ in-line cleavage phenomena can be expected to hold to any RNAs regardless of the presence of an oxidant, given they do not specifically coordinate or shield themselves from the iron to prevent cleavage.…”

Multiple prebiotic metals mediate translation

“…What was the role of Fe 2+ and Mn 2+ in the origin and early evolution of the translational system? Finally, what are the implications for ribosome-bound Fe 2+ in oxidative damage and disease ( 50 , 51 )?…”

Effects of Oxidative Stress on Protein Translation: Implications for Cardiovascular Diseases

“…For example, the Cys-Cys-switch found in the Yap1 transcription factor master regulator of Saccharomyces cerevisiae leads to a conformational change upon Cys oxidation of Yap1, promoting the factor's translocation into the nucleus to activate expression of~100 stress-responsive genes [198]. Low-dose oxidative stress catalyzes chemical Fe(II)-dependent hydrolysis of 25S rRNA within the regulatory region of the yeast ribosome (expansion segment 7, ES7L), generating translationally active ribosomal species that likely participate in stress adaptation [199,200]. These examples demonstrate that ROS are not always damaging, but rather, in biological systems like eukaryotic cells, can play dual roles, adaptive at low concentrations and destructive at high concentrations.…”