Linking the unfolded protein response to oxygen sensing - A phosphoproteomic approach to identify novel substrates of the ER kinase PERK

Description

The aging process is characterised by tissue decline and the onset of age-associated diseases. It is not, however, immutable, and can be modulated by manipulating the cell signalling pathways perturbed as cells and organisms age. A key characteristic of aging is the progressive accumulation of damaged/unfolded proteins in the endoplasmic reticulum (ER) leading to activation of the unfolded protein response (UPR). The UPR is the collective name for a number of cell signalling pathways that act in concert to restore normal cellular function by halting protein synthesis, clearing misfolded proteins and increasing the production of molecular chaperones. PERK, an ER-resident protein kinase, is critical to the UPR and acts to stall global protein synthesis. However inappropriate PERK activity is associated with the pathology of several age-related disorders including degenerative neurological diseases and cancer. Despite this key role for PERK very few direct substrates have been identified. 

 

Project Description and Experimental Approach

We have identified a new role for PERK as a regulator of the hypoxia-inducible factor (HIF)-dependent response to low oxygen – another key signalling pathway disrupted in human diseases1. In this project you will define how the UPR, and PERK specifically, communicates with the HIF-dependent response to low oxygen. Initially, you will identify new PERK-dependent phosphorylation events by performing phosphoproteomic mass-spectrometry experiments2. You will then determine the functional outcome of deleting/ mutating candidate PERK substrates on both the UPR and HIF signalling using state of the art CRISPR/Cas9-mediated genome editing techniques and relevant kinase assays. Completion of these experiments will lead to a greater understanding of how these signalling pathways communicate and contribute to the aging process.

 

Training and Environment

This Liverpool-based studentship provides an opportunity to use cutting edge mass-spectrometry coupled with genome editing techniques to dissect the function of important stress-responsive pathways. The placement in Newcastle University will provide excellent training in validating kinase-substrate relationships. The depth of expertise at both institutions and the world class facilities available, will provide an excellent training environment and give you a highly sought-after skill set to enhance your future career prospects.

Please contact Dr Niall Kenneth () for more details and informal enquires.

1. Ivanova et al. PERK/eIF2α signaling inhibits HIF-induced gene expression during the unfolded protein response via YB1-dependent regulation of HIF1α translation. NAR. 46,(2018).

2. Hardman et al. Strong anion exchange‐mediated phosphoproteomics reveals extensive human non‐canonical phosphorylation. EMBO J. (2019)

HOW TO APPLY

Applications should be made by emailing  with a CV and a covering letter, including whatever additional information you feel is pertinent to your application; you may wish to indicate, for example, why you are particularly interested in the selected project/s and at the selected University. Applications not meeting these criteria will be rejected. We will also require electronic copies of your degree certificates and transcripts.

In addition to the CV and covering letter, please email a completed copy of the NLD BBSRC DTP Studentship Application Details Form (Word

document) to , noting the additional details that are required for your application which are listed in this form. A blank copy of this form can be found at: https://www.nld-dtp.org.uk/how-apply.

Availability

Open to students worldwide

Funding information

Funded studentship

Studentships are funded by the Biotechnology and Biological Sciences Research Council (BBSRC) for 4 years. Funding will cover tuition fees at the UK rate only, a Research Training and Support Grant (RTSG) and stipend. We aim to support the most outstanding applicants from outside the UK and are able to offer a limited number of bursaries that will enable full studentships to be awarded to international applicants. These full studentships will only be awarded to exceptional quality candidates, due to the competitive nature of this scheme.

Supervisors

References

1. (2018) PERK/eIF2α signaling inhibits HIF-induced gene expression during the unfolded protein response via YB1-dependent regulation of HIF1α translation. Nucleic Acids Research. May 4;46(8):3878-3890
2. (2019) Translating the Hypoxic Response-the Role of HIF Protein Translation in the Cellular Response to Low Oxygen. Cells. Feb 1;8(2).
3. (2017) XIAP upregulates expression of HIF target genes by targeting HIF1α for Lys63-linked polyubiquitination. Nucleic Acids Research. Sep 19;45(16):9336-9347
4. (2020) Kinase inhibition profiles as a tool to identify kinases for specific phosphorylation sites. Nature Communications. 11:1684.
5. (2010) Histone H3 Thr-3 Phosphorylation by Haspin Positions Aurora B at Centromeres in Mitosis. Science 330,231-5.
6. (2019) CDK1-mediated phosphorylation at H2B serine 6 is required for mitotic chromosome segregation.. J. Cell Biol. 2019 218:1164-81.
7. (2019) Strong anion exchange‐mediated phosphoproteomics reveals extensive human non‐canonical phosphorylation. EMBO J 38:e100847 doi.org/10.15252/embj.2018100847.
8. (2017) Evaluation of parameters for confident phosphorylation site localization using an orbitrap fusion tribrid mass spectrometer. J Proteome Research 16:9 3448-3459
9. (2016) Dynamic phosphorylation of RelA on Ser42 and Ser45 in response to TNFα stimulation regulates DNA binding and transcription. Open Biology DOI: 10.1098/rsob.160055