Download Nutritional control of p53 activity and its role in metabolic flexibility

Nutritional control of p53 activity and its role in metabolic
flexibility
Summary
Andreas Prokesch, Institute of Cell Biology, Histology and Embryology, Medical University of Graz
Supervisor:
Availability:
Offered by:
Application deadline:
PD Dr. Andreas Prokesch
This position is available.
Medical University of Graz
Applications are accepted between February 01, 2017 00:00 and March 19,
2017 23:59 (CEST)
Description
Background:
The tumor suppressor p53 is a transcription factor activated in cancerous cells by a variety of stress signals such
as DNA damage, oncogene activation, nutrient deprivation, and hypoxia. Once activated, the p53 pathway has a
wide range of downstream effects among which cell death, cell cycle arrest, autophagy, and regulation of cellular
metabolism are most prominent (1;2). While many functional aspects for the p53-mediated cellular stress response
during tumorigenesis (and therefore in rapidly dividing cells) are by now well established, much less is known about
the role of p53 in non-transformed, terminally differentiated cells. Targeting p53 has enormous potential in cancer
therapy since it is mutated in more than 50% of all human tumors (3). More recently, data have emerged that place
the p53 pathway in the center of the control of cancer metabolism showing that it is essential to re-program glucose
and lipid homeostasis (4;5). These functions of p53 in energy metabolism are largely consistent with its role as a
tumor suppressor, although some reports suggest pro-Warburg-effects of p53. Importantly, most studies focused on
the metabolic effects of p53-ablation during malignant transformation of cells. At present, little research has been
conducted on the role of p53 in post-mitotic cell types, the p53-mediated control mechanisms of substrate allocation
in healthy tissues, and its consequences for systemic energy homeostasis and metabolic flexibility (the capacity of
an organism to adapt utilization of nutrients to their availability).
In a recent publication (Prokesch et al., FASEB J, 2016 (6)) we showed that acute knock-out of p53 in mouse liver
disturbs glycogen storage and leads to lipid accumulation in hepatocytes. Furthermore we showed that the p53 protein accumulates in hepatocytes under starvation. Accordingly, in the acute liver knock-out model amino acid and
glucose homeostasis is disrupted. These and preliminary data in other tissues suggest that, beyond its role as tumor
suppressor, p53 plays a role as metabolic regulator in post-mitotic cells and tissues.
Hypothesis and Objectives:
With our preliminary data as foundation we recently acquired an FWF-DACH grant (lead applicant Andreas Prokesch).
This project is a collaboration with the Charite in Berlin (Prof. Michael Schupp) and the German Institute for Nutritional
Research (Prof. Tim Schulz). In this consortium we will utilize novel tissue-specific, inducible knock-out mouse models
to investigate the role of p53 in the metabolism of liver, white adipose tissue, brown adipose tissue, and skeletal
muscle.
The PhD student working in Graz will be focused on (i) elucidating the upstream mechanisms of starvation-induced
p53 stabilization (Aim 1 of the proposal) and (ii) work on the liver- and white adipose tissue (WAT)-specific aspects
of the project (aims 2 and 3 of the proposal). For the upstream mechanism we hypothesize regulation by the cellular
energy sensor AMPK. This was already shown in cultured cells (Prokesch et al., FASEB J, 2016 (6)) and will be
investigated in vivo by the PhD student. Another mechanism we will investigate is specific MDM2 (the canonical
endogenous p53 inhibitor) degradation under starvation. As proposed p53 downstream mechanisms in WAT and liver
we recently obtained data, suggesting a regulation of autophagy (and potentially lipophagy) by p53 under starvation.
Therefore, the PhD student will employ autophagy-related assays to the used model systems.
Methodology:
In addition to classical cell culture work (protein overexpression, Crispr/cas9, RNAi, proliferation assays) followed
by downstream analyses such as western blot, qPCR, cell cycle analyses, the student will perform immunoprecipGlowbase Graduate Recruitment Platform - http://www.glowbase.com - © Glowbase GmbH - 2017-01-31 16:30:22
itation assays followed by mass spectrometry (collaboration with Ruth Birner-Grünberger) to elucidate p53 interaction partner dynamics upon starvation. In the mouse models the student will perform metabolic phenotyping (e.g.
metabolic cages (collaboration with Dagmar Kratky), insulin and glucose tolerance tests) and will prepare RNA-seq
libraries (collaboration with Andrew Pospisilik, MPI, Freiburg) and samples for untargeted proteomics (collaboration
with Christoph Magnes, Joanneum Research) from selected tissues. Primary cells derived from these mouse models
will be used to measure mRNA and protein expression, oxygen consumption (Seahorse, collaboration with Wolfgang
Graier), ROS production, and autophagy (LC3B lipidation, fluorescence microscopy, and electron microscopy).
References:
1. Vousden, K. H., Prives, C. (2009) Blinded by the Light: The Growing Complexity of p53. Cell 137, 413-431
2. Junttila, M. R., Evan, G. I. (2009) p53--a Jack of all trades but master of none. Nat.Rev.Cancer 9, 821-829
3.
Muller, P. A., Vousden, K. H. (2014) Mutant p53 in cancer: new functions and therapeutic opportunities.
Cancer Cell 25, 304-317
4. Berkers, C. R., Maddocks, O. D., Cheung, E. C., Mor, I., Vousden, K. H. (2013) Metabolic regulation by p53
family members. Cell Metab 18, 617-633
5. Vousden, K. H., Ryan, K. M. (2009) p53 and metabolism. Nat.Rev.Cancer 9, 691-700
6. Prokesch, A., Graef, F. A., Madl, T., Kahlhofer, J., Heidenreich, S., Schumann, A., Moyschewitz, E., Pristoynik,
P., Blaschitz, A., Knauer, M., Muenzner, M., Bogner-Strauss, J. G., Dohr, G., Schulz, T. J., Schupp, M. (2016) Liver
p53 is stabilized upon starvation and required for amino acid catabolism and gluconeogenesis. FASEB J.
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Glowbase Graduate Recruitment Platform - http://www.glowbase.com - © Glowbase GmbH - 2017-01-31 16:30:22