Functional Phosphoproteomics

Our laboratory studies cell signalling to understand the driving forces behind complex human diseases.

Cells rely on a biochemical process called phosphorylation to respond to the environment and to engage in numerous biological functions. Phosphorylation is therefore an important mechanism that underpins almost all cellular behaviour. Over the last decade, powerful new technologies using mass spectrometry have emerged that now make it possible to measure cellular phosphorylation at a genome-wide scale.

In our lab, we use these mass spectrometry technologies to explore how cells behave in both healthy and disease contexts. By studying phosphorylation in this way, we can obtain unique insights into how cells function differently during disease. This information can help uncover how human diseases arise and discover effective new therapies to treat them.

Our projects

Signalling network target deconvolution of a small-molecule library

This project aims to characterise the biological pathways and downstream effector proteins engaged by a small-molecule library. High-throughput phenotypic screens using compound libraries are a fundamental approach for drug discovery pipelines. A major limitation of such screens, however, is in identifying what pathways in the cell are specifically responsible for the observed effects.

Leveraging world-leading mass spectrometry instrumentation and methodologies pioneered by our lab, we will perform an extensive characterisation of a small-molecule library. This unique data resource will support the interpretation of both existing and future drug development efforts. The outcome of this work will be an amplification of the utility of compound libraries, enabling the design and execution of phenotypic screens in complex cellular models using rationally selected subsets of compounds. Finally, this project will enable MCRI researchers to embrace phosphoproteomics technologies more broadly within drug development pipelines, with major implications for the design of future therapeutics.

Mapping the Beneficial Effects of Exercise in Humans using Personalised Phosphoproteomics

Exercise has potent beneficial effects on many degenerative disorders including type II diabetes and cardiovascular disease. We hypothesise that these wide-ranging benefits are largely due to exercise improving the sensitivity of tissues to the hormone insulin. In this project, we aim to characterise the molecular mechanisms responsible for the insulin-sensitising actions of exercise in skeletal muscle.

Despite its potent effects, exercise is not well prescribed due to its complexity both at a molecular level and as an activity, and because not all patients can achieve the level of exercise needed to receive its insulin-sensitising effects. This highlights the need for therapies that mimic the beneficial effects of exercise. This will require deep insights into the mechanistic function of pathways engaged by exercising muscles so they can be selectively targeted. In this proposal, we will exploit technological innovations to uncover this knowledge.

Funding

• National Health and Medical Research Council (NHMRC)
• Novo Nordisk Foundation Center for Stem Cell Medicine, reNEW

Collaborations

• Professor Enzo Porrello (MCRI)
• Associate Professor David Elliott (MCRI)
• Dr Kevin Watt (MCRI)
• Professor David Eisenstat (MCRI)
• Associate Professor Silvia Velasco (MCRI)
• Dr James McNamara (MCRI)
• Associate Professor Richard Mils (MCRI)
• Professor Janni Petersen (Flinders University)
• Professor Misty Jenkins (WEHI)
• Associate Professor Andy Philp (Centenary Institute)
• Professor David James (The University of Sydney)
• Professor Jørgen Wojtaszewski (University of Minnesota Morris)
• Professor James Hudson (QIMR Berghofer)

Featured publications

Fazakerley DJ et al. James DE, Humphrey SJ. Phosphoproteomics reveals rewiring of the insulin signaling network and multi-nodal defects in insulin resistance. Nature Communications (2023) 14:923.

Needham EJ et al., James DE, Wojtaszewski JFP, Humphrey SJ. Personalized phosphoproteomics identifies functional signaling. Nature Biotechnology (2022) 40:576-84.

Needham EJ et al. James DE, Humphrey SJ. Illuminating the dark phosphoproteome. Science Signaling (2019) 12:eaau8645.

Humphrey SJ et al., High-throughput and high-sensitivity phosphoproteomics with the EasyPhos platform. Nature Protocols (2018) 13:1897-916.

Humphrey SJ, Azimifar B, Mann M. High-throughput phosphoproteomics reveals in vivo insulin signaling dynamics. Nature Biotechnology (2015) 33:990-5.