RESEARCH
Our Current Focus
Our Research
We are conducting a range of research projects to characterise molecular interactions regulating gene expression and pre-mRNA splicing, aimed primarily at identifying human disease mechanisms and investigating opportunities for developing new therapeutic approaches. This includes biochemical analyses of the human splicing machinery and functional characterization of compartments and bodies in mammalian cell nuclei. Our projects involve mechanistic studies at the molecular and cellular levels, spanning biochemical, microscopy and MS-based proteomics technologies, including studies on libraries of human induced pluripotent stem cells. We also have a strong interest in using state of the art, quantitative mass spectrometry-based approaches to perform detailed analyses of protein expression and protein interactions in cells and tissues.
Modulating Alternative Splicing (AS) with Small Molecules
The information contained in human genes is ‘split’, which allows a single gene to code for multiple different proteins. Alternative Splicing (AS) of mRNA precursors is the RNA processing mechanism that enables the production of different messenger RNAs and proteins from a single gene. Defects in AS are known to be involved in a wide range of human disease mechanisms. We are studying the mechanisms in human cells that regulate splice site selection by the splicing machinery and how this can be modulated by small molecules. Our aim is to explore how specifically we can modulate targeted AS events in cells using small molecules, which has great potential for developing novel therapeutics to treat a range of inherited neurodegenerative disorders and other diseases of unmet need. Our work on AS has been funded by grants from the BBSRC and includes participation in the ‘UNLEASH’ project, which is a close collaboration with the groups of David Gray in the University of Dundee Drug Discovery Center and with Juan Valcarcel at the CRG Institute in Barcelona and Michael Sattler at the Helmholtz Institute in Munich. The UNLEASH project is supported by a Synergy Grant with funding provided by both the European Research Council (ERC) and UK Research and Innovation (UKRI).
Characterising Splice Site Modulation Approaches for Huntington’s Disease Therapeutics
We are studying the role of pre-mRNA splicing mechanisms in Huntington’s disease and investigating opportunities for targeting splicing modulation as a future therapeutic strategy. The project is conducted in collaboration with Gillian Bates and colleagues at UCL. A major focus of this project is to apply the biochemical and proteomics technologies and assay systems we have developed to study the composition of splicing complexes that recognise splice sites in Huntington’s gene (HTT) transcripts. This involves a combination of biochemical and cellular assays (mass spectrometry-based proteomics, fluorescence microscopy and FACS, protein complex purification, biochemical analyses and RNAseq), using both mouse brain tissue and cultured cell models. We are also analysing the specificity of the small molecule drugs branaplam and risdiplam for their ability to modulate selection of splice sites relevant to Huntington’s disease, using both human cells and mouse models. The project aims to illuminate how alternative splice site recognition and the pre-mRNA splicing machinery is affected by branaplam & risdiplam, which can modulate HTT intron49 splicing and by mutations in mouse models of Huntington’s disease. We also plan to extend these studies to an analysis of the impact of HTT
mutations on rates of protein turnover in mouse brain regions, using in vivo SILAC-MS.
Functional Analyses of Protein Modification by Proline Hydroxylation
In collaboration with the groups of Sonia Rocha at the University of Liverpool and Jason Swedlow at the University of Dundee School of Life Sciences, we are studying how site-specific hydroxylation of proline residues on proteins can affect cellular regulatory mechanisms. With previous funding from the Wellcome Trust, we have developed novel MS-based methods to reliably identify protein targets for enzymatic hydroxylation of proline residues by Prolyl Hydroxylases (PHDs). Using this methodology, we have identified hundreds of novel hydroxylation sites on proteins and in parallel cell assays shown that some of these hydroxylated proteins are involved in regulating cell cycle progression. This includes the protein Repo-Man (also called CDCA2), a cell cycle regulated protein we discovered previously as a regulatory subunit for protein phosphatase PP1γ, with important roles in mitotic progression and cell viability. We find that Pro604 on Repo-Man is hydroxylated by PHD1 and that mutation of proline 604 to alanine (P604A) reduces the interaction between RepoMan and phosphatase PP2A-B56g and results in defects in chromosome alignment, segregation (chromosomes bridges) and cell death.
Deciphering the role of adipose tissue in common metabolic disease via adipose tissue proteomics
This project is a collaboration with the group of Kerrin Small at King’s College London (KCL), supported by a grant from the MRC. The project leverages the TwinsUK resource at KCL and has the overarching aim to characterize the adipose proteome in healthy ageing individuals at unprecedented depth and resolution. By comparing adipose proteomes in 1,000 twins, we can identify its variance across individuals, genetics and phenotypic traits. Adipose tissue is a critical endocrine organ which plays a central role in the regulation of metabolism, inflammation, synthesis and secretion of hormones and serves as a buffering system for lipid energy balance. Adipose tissue dysfunction is known to be involved in monogenic forms of metabolic disease and GWAS studies of common genetic variants have demonstrated that adipose tissue is a key mediator of genetic risk for metabolic traits. This project will quantify the breadth of proteins, peptides and post-translational modifications present in human adipose tissue and these novel adipose
proteome data will be integrated with existing TwinsUK data. By profiling adipose proteomics in the data-rich Twins-UK population we can enable identification of novel protein regulatory variants and determine whether they mediate disease-associated loci. Integration with phenotypic data will identify proteins, specific protein isoforms & PTMs and protein networks associated to clinical traits, providing important insights for designing future therapeutic strategies.
Dissecting a TDP-43 knock-in allelic series to yield diverse ALS drug targets
This is a new project, aimed at studying mechanisms underlying ALS, which we are conducting together with Jemeen Sreedharan and Ravindra Prajapati at King’s College London (KCL) and their collaborators, supported by a grant from the My Name’5 Doddie Foundation. The project will focus on how mutations in TDP43, the most important protein in ALS pathogenesis, contribute to disease mechanisms. The Sreedharan group are establishing a library of human neuronal cell lines expressing TDP43 mutants of known clinical relevance, using a switch that allows swapping of pieces of TDP43. In the Lamond group we will then study the phenotypes resulting from expression of these TDP43 mutations, using state of the art, quantitative mass spectrometry-based proteomics methods to perform unbiased characterisation of protein expression and protein complex formation. The phenotypes of all TDP43 mutant cell lines will be rigorously characterised at the levels of gene expression, protein expression and protein complex profiling, to dissect mechanisms underlying TDP43 dysregulation in ALS.
Analysing Variations in Human Drug Response using Pluripotent Human iPS Cells
This project is a collaboration with Jason Swedlow and Melpi Platani at the University of Dundee School of Life Sciences. Our main goal is to advance our understanding of how human genetic diversity affects fundamental biological mechanisms and cellular pathways and to use this information to help improve drug development pipelines. The current pharmaceutical industry drug discovery pipeline has a very high failure rate (overall only ~10% of drug candidates sent for clinical trials gain regulatory approval for use in patients) and it typically focusses on patients from limited ethnic, geographic and gender groups. We have developed an innovative, lab-based drug screening platform that uses cell painting, machine learning and MS-based proteomics to compare drug responses across libraries of human iPSC cell lines derived from genetically distinct, healthy human donors. We have shown this platform can detect and explain mechanisms causing variable drug responses for clinically approved drugs used to treat a range of diseases. We are continuing to develop this novel technology platform to improve efficiency and to explore how it can be used to improve the delivery of safe and effective new drugs that cater for the needs of diverse human populations. The platform has the potential to identify patient biomarkers for predicting variable response that can inform the construction of more efficient and successful clinical trials and aid translation of existing drugs for use in different geographical regions.
Structure-Function Analyses of Nuclear Bodies
We have a long-standing interest in studying the functional compartmentalization of mammalian cell nuclei and specifically how RNA processing machineries are organised in different types of subnuclear bodies (NBs). This includes analyses of the morphology, composition and function of nucleoli, Cajal bodies and ‘splicing speckles’ (interchromatin granule clusters). With funding from the BBSRC, we have identified novel families of small molecules – ‘Nuclear Body Modulators’ (NBMs) - that alter either the size, morphology and/or composition of specific classes of NBs. We are using Thermal Proteome Profiling MS analysis to help identify the protein targets of the NBMs. In parallel, we are using a range of methods, including fluorescence and electron microscopy, together with biochemical assays, to characterize the impact of the NBMs on nuclear RNA processing mechanisms. This project includes an analysis of nuclear ultrastructure, in collaboration with Gareth Griffiths and Jens Wohlmann at the University of Oslo.
Grant Funding
Wellcome Trust Collaborative Award (206293/Z/17/Z); “The interplay between the oxygen sensors PHDs and the cell cycle”; £1,542,380; (01.07.2018 - 31/08/2024); Co-applicant together with Prof Sonia Rocha, Univ of Liverpool; Prof Jason Swedlow, UoD; Prof Stewart Fleming, UoD
BBSRC Project Grant (BB/V010948/1); “Molecular Analysis of Nuclear Bodies and RNP Trafficking Pathways in the Cell Nucleus”; £1,128,053; (01.05.2021 - 31/04/2025)
ERC Synergy Grant “UNLEASH Project: ‘Harnessing the splicing code for targeted control of gene expression’ (01.06.2023-31.05.2029) Total award – Euros 10,232,728; Lamond group awarded £2,117,288 from UKRI (under ‘guarantee scheme’ while UK status for ERC funding was still in negotiation). Co-applicant together with Professor Juan Valcarcel, CRG, Barcelona, Spain; Prof Michael Sattler, Helmholtz Institute, Munich, Germany; Professor David Gray, UoD.
CHDI Project ‘Characterising Splice Site Modulation Approaches for Huntington’s Disease Therapeutics’ £1,262,082 (01.06.2023-31.05.2025)
BBSRC Pioneer Award (BB/Y513040/1); “Unlocking the Alternative Splicing Code”; £198,277; (01.02.2024 - 31/01/2025)
HFSP Postdoctoral Fellowship (LT000438/2021-L / Angela Harrison) “Protein correlation profiling analysis of the Nipah virus-host interface”; (31.01.2022-31.12.20.24)
We have also been informed that another collaborative grant has just been funded that will start in 2025:
My Name’5 Doddie Foundation Discovery Network Award
Project title: Dissecting a TDP-43 knock-in allelic series to yield diverse MND drug targets
Application number: 2DNFA\100010
This is a collaboration with Jemeen Sreedharan’s group at KCL and their collaborators at the University of Cambridge, as described in the text about our research projects.
