HIF prolyl hydroxylase-3 regulates actin polymerisation and hypoxia-induced motility and invasion

Limited oxygen availability (hypoxia) influences cell migration and invasion, but the underlying mechanisms are poorly understood. Much of the cellular response to hypoxia is regulated by a family of Hypoxia Inducible Factor (HIF) prolyl hydroxylases (PHD1-3), each of which is thought to regulate specific pathways.Their activity is dependent on the availability of oxygen and alpha-ketoglutarate but despite intensive studies their activity in vivo and their substrates are poorly defined. In this study we performed a quantitative proteomic screen to identify new substrates of PHDs. Co-immunoprecipitations using FLAG-tagged PHDs were performed under hypoxia to trap the enzyme-substrate interactions, and binding partners were identified by mass spectrometry. Actin was identified to interact with PHD3 specifically under hypoxia. Subsequently two defined prolyl residues in beta-actin were shown to be hydroxylated. Hypoxia-induced rearrangement of the actin cytoskeleton was shown to be dependent on PHD3 activity as a knockdown of PHD3 was sufficient to increase the intracellular G- to F-actin ratio. An increase in cell migration and invasion was also found to be dependent on PHD3 activity. Mutation of both hydroxylated prolyl residues led to a similar phenotype regarding actin rearrangement and cell migration. Using constantly active HIF-mutants, we could show that these PHD3-dependent pathways are independent of HIF. All together, this study shows a pro-invasive pathway linking HIF-independent oxygen-sensing pathways and actin signalling. However, the mechanism of how hypoxia-induced actin rearrangement leads to increased migration and invasion remains to be elucidated

BRCA1 mislocalization leads to aberrant DNA damage response in heterozygous ABRAXAS1 mutation carrier cells

Peer reviewe

MCCP: are medium-chain chlorinated paraffins of concern for humans?

Glyphosate (N-[phosphonomethyl]-glycine) is the most widely used herbicide worldwide. Due to health concerns about glyphosate exposure, its continued use is controversially discussed. Biomonitoring is an important tool in safety evaluation and this study aimed to determine exposure to glyphosate and its metabolite AMPA, in association with food consumption data, in participants of the cross-sectional KarMeN study (Germany). Glyphosate and AMPA levels were measured in 24-h urine samples from study participants (n = 301). For safety evaluation, the intake of glyphosate and AMPA was calculated based on urinary concentrations and checked against the EU acceptable daily intake (ADI) value for glyphosate. Urinary excretion of glyphosate and/or AMPA was correlated with food consumption data. 8.3% of the participants (n = 25) exhibited quantifiable concentrations (> 0.2 µg/L) of glyphosate and/or AMPA in their urine. In 66.5% of the samples, neither glyphosate (< 0.05 µg/L) nor AMPA (< 0.09 µg/L) was detected. The remaining subjects (n = 76) showed traces of glyphosate and/or AMPA. The calculated glyphosate and/or AMPA intake was far below the ADI of glyphosate. Significant, positive associations between urinary glyphosate excretion and consumption of pulses, or urinary AMPA excretion and mushroom intake were observed. Despite the widespread use of glyphosate, the exposure of the KarMeN population to glyphosate and AMPA was found to be very low. Based on the current risk assessment of glyphosate by EFSA, such exposure levels are not expected to pose any risk to human health. The detected associations with consuming certain foods are in line with reports on glyphosate and AMPA residues in food

Demographic and clinical data of patients and healthy controls of which freshly isolated PBMCs were analyzed in this study.

<p>Demographic and clinical data of patients and healthy controls of which freshly isolated PBMCs were analyzed in this study.</p

γ-H2AX and 53BP1 levels in freshly isolated PBMCs of patients and healthy controls.

<p>(A) γ-H2AX foci per cell in patients with CIS/early RRMS and healthy controls. (B) Percentage of γ-H2AX positive cells in patients with CIS/early RRMS and healthy controls. (C) 53BP1 foci per cell in patients with CIS/early RRMS and healthy controls. (D) Percentage of 53BP1 positive cells in patients with CIS/early RRMS and healthy controls. Each data point represents the median of n = 6 separate measurements per individual. In each of the six separate measurements approximately n = 100 cells were scored. The horizontal bar indicates the median.</p

Demographic and clinical data of patients and healthy controls of which previously frozen PBMCs were analyzed in this study.

<p>Demographic and clinical data of patients and healthy controls of which previously frozen PBMCs were analyzed in this study.</p

Glutaminolysis activates Rag-mTORC1 signaling

Amino acids control cell growth via activation of the highly conserved kinase TORC1. Glutamine is a particularly important amino acid in cell growth control and metabolism. However, the role of glutamine in TORC1 activation remains poorly defined. Glutamine is metabolized through glutaminolysis to produce α-ketoglutarate. We demonstrate that glutamine in combination with leucine activates mammalian TORC1 (mTORC1) by enhancing glutaminolysis and α-ketoglutarate production. Inhibition of glutaminolysis prevented GTP loading of RagB and lysosomal translocation and subsequent activation of mTORC1. Constitutively active Rag heterodimer activated mTORC1 in the absence of glutaminolysis. Conversely, enhanced glutaminolysis or a cell-permeable α-ketoglutarate analog stimulated lysosomal translocation and activation of mTORC1. Finally, cell growth and autophagy, two processes controlled by mTORC1, were regulated by glutaminolysis. Thus, mTORC1 senses and is activated by glutamine and leucine via glutaminolysis and α-ketoglutarate production upstream of Rag. This may provide an explanation for glutamine addiction in cancer c