Oxygen and glucose deprivation induces widespread alterations in mRNA translation within 20 minutes

Abstract

BackgroundOxygen and glucose metabolism play pivotal roles in many (patho)physiological conditions. In particular, oxygen and glucose deprivation (OGD) during ischemia and stroke results in extensive tissue injury and cell death.ResultsUsing time-resolved ribosome profiling, we assess gene expression levels in a neural cell line, PC12, during the first hour of OGD. The most substantial alterations are seen to occur within the first 20 minutes of OGD. While transcription of only 100 genes is significantly altered during one hour of OGD, the translation response affects approximately 3,000 genes. This response involves reprogramming of initiation and elongation rates, as well as the stringency of start codon recognition. Genes involved in oxidative phosphorylation are most affected. Detailed analysis of ribosome profiles reveals salient alterations of ribosome densities on individual mRNAs. The mRNA-specific alterations include increased translation of upstream open reading frames, site-specific ribosome pauses, and production of alternative protein isoforms with amino-terminal extensions. Detailed analysis of ribosomal profiles also reveals six mRNAs with translated ORFs occurring downstream of annotated coding regions and two examples of dual coding mRNAs, where two protein products are translated from the same long segment of mRNA, but in two different frames.ConclusionsThese findings uncover novel regulatory mechanisms of translational response to OGD in mammalian cells that are different from the classical pathways such as hypoxia inducible factor (HIF) signaling, while also revealing sophisticated organization of protein coding information in certain genes.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-015-0651-z) contains supplementary material, which is available to authorized users.

PHD2 Targeting Overcomes Breast Cancer Cell Death upon Glucose Starvation in a PP2A/B55α-Mediated Manner

“…Together with hypoxia, nutrient restriction is a major feature of the tumor microenvironment. If, on one hand, we show that oxygen shortage enables B55α accumulation, on the other hand, we speculate that in breast cancer cells glucose deprivation reduces the production of fumarate mainly because of a block of SDH (Andreev et al., 2015), tilting the balance toward an excess of α-KG at the expense of the PHD2 inhibitor fumarate, overall resulting in enhanced PHD2 activity and increased B55α degradation. In parallel, prolonged glucose starvation results in P70S6K reactivation (G.D.C.…”

“…In both cell lines, malate levels were tremendously reduced to a similar extent as observed for fumarate (Figures 2H and 2I). This, together with the fact that succinate levels were unaltered, suggests a block of succinate dehydrogenase (SDH) activity upon glucose starvation (Andreev et al., 2015). At least in MCF7, the amount of α-KG coming from glutamine was increasing under glucose deprivation versus fed conditions, as evidenced by U- 13 C-glutamine fractional labeling (Figure S2A).…”

Limitations of oxygen delivery to cells in culture: An underappreciated problem in basic and translational research

“…Luminescent or fluorescent oxygen based sensors are added to cell culture systems by way of plate coating, adhesive stickers, or even beads. As dissolved oxygen concentrations change, so does the specific light readout which is quantified by an external, noninvasive detector [65]. While this methodology does not enhance the oxygen diffusion in standard cell culture models, it does afford researchers the opportunity to detect diffused oxygen concentrations in their previously optimized systems.…”

Systemic Reprogramming of Translation Efficiencies on Oxygen Stimulus

“…In support of this model, another cap-binding protein, eIF4E3, has been suggested to regulate translation under other settings (Landon et al, 2014). Thus, it is likely that cells have evolved multiple alternative translation initiation machineries that allow them to activate stimulus-specific translation efficiency programs (Andreev et al, 2015; Baudin-Baillieu et al, 2014; Kronja et al, 2014; Lu et al, 2009; McKinney et al, 2014; Ventoso et al, 2012; Young et al, 2008). These alternative machineries would reprogram global mRNA translation efficiencies in order to generate distinct, adaptive translatomes/proteomes.…”