Hypoxia Strongly Affects Mitochondrial Ribosomal Proteins and Translocases, as Shown by Quantitative Proteomics of HeLa Cells

Abstract

Hypoxia is an important and common characteristic of many human tumors. It is a challenge clinically due to the correlation with poor prognosis and resistance to radiation and chemotherapy. Understanding the biochemical response to hypoxia would facilitate the development of novel therapeutics for cancer treatment. Here, we investigate alterations in gene expression in response to hypoxia by quantitative proteome analysis using stable isotope labeling with amino acids in cell culture (SILAC) in conjunction with LCMS/MS. Human HeLa cells were kept either in a hypoxic environment or under normoxic conditions. 125 proteins were found to be regulated, with maximum alteration of 18-fold. In particular, three clusters of differentially regulated proteins were identified, showing significant upregulation of glycolysis and downregulation of mitochondrial ribosomal proteins and translocases. This interaction is likely orchestrated by HIF-1. We also investigated the effect of hypoxia on the cell cycle, which shows accumulation in G1 and a prolonged S phase under these conditions. Implications. This work not only improves our understanding of the response to hypoxia, but also reveals proteins important for malignant progression, which may be targeted in future therapies.

Hypothesis: Hypoxia inducesde novosynthesis of NeuGc gangliosides in humans through CMAH domain substitute

“…However, we did not observe an increased expression of NeuGc GM3 using media supplemented with fetal bovine serum, which contains NeuGc ( Figure S2). Sialin (SLC17A5) was not identified in our recent SILAC study, not even at below-significance levels, even though 26 other proteins from the SCL-family were detected, of which four were upregulated [34]. One of them was the GM3 synthase (ST3GAL5/SLC35E1) responsible for catalyzing the covalent addition of sialic acid to lactosylceramide to generate the GM3 ganglioside.…”

“…In the same study, we found an iron-sulfur [Fe 2 S 2 ]-containing protein to be significantly up-regulated under hypoxic conditions (1.9 fold; p=0.0057), while many other mitochondrial proteins were down-regulated [34]. The protein in question is subunit B of the succinate dehydrogenase complex (which represents complex II of the respiratory chain).…”

“…Bioinformatics analysis was performed with STRING 9.1 (http://string.embl.de/) [35,36], on data acquired previously by NanoLC-LTQ Orbitrap mass spectrometry, as described [34].…”

“…SILAC is a quantitative proteomics method that reveals up-or down-regulated proteins upon exposure to different conditions [33]. The present work builds on a recent comprehensive SILAC study, where we analyzed protein regulation in HeLa cells in response to hypoxia [34].…”

Whole-Genome Association Analyses of Sleep-disordered Breathing Phenotypes in the NHLBI TOPMed Program

“…Additional genes were significantly associated in population-specific analyses, including the components of the oxidative phosphorylation system, and a majority of the small subunit proteins have been implicated in disease [ 88 ]. The expression of several small and large subunit proteins are altered in a hypoxic environment [ 89 ]. MRPS33 expression varies with oxygen treatment in COPD [ 90 ].…”