Alternative splicing enables a single gene to produce multiple mRNA isoforms, and potentially proteins with distinct functions. It is especially important in the brain, where transcript diversity supports development and ageing. Studies have implicated disrupted RNA splicing in neurodevelopmental, neuropsychiatric and neurodegenerative disorders. Yet detecting a novel mRNA isoform does not prove that it is translated into a stable, functional protein. Establishing which transcripts reach the proteome is a major challenge in functional genomics. Long-read sequencing allows complete RNA molecules to be characterised, and studies have uncovered extensive unannotated transcript diversity in the human brain.
If transcripts are predicted to have coding potential, then these novel transcripts contain uncharacterised coding sequences (CDS). Our research has identified over 20,000 novel transcripts in the brain with CDS. These are a result of alternative splicing events such as exon skipping, alternative transcriptional start/end sites and fusion transcripts. Preliminary analyses suggest that genetic variation affecting these sequences is associated with neurodevelopmental and neuropsychiatric conditions. The key question is ‘which novel brain mRNA isoforms are translated into proteins, and what are their functional and disease-relevant consequences?’
Objective 1 is to identify and prioritise novel CDS predicted to generate new protein isoforms, alter known proteins, or change the balance between functionally distinct isoforms. The initial focus will be on genes involved in brain development, ageing and disorders.
Objective 2 is to determine whether prioritised transcripts are translated. The student will integrate transcriptomic and proteomic data and validate selected candidates using mass spectrometry and Quantum-Si single-molecule protein sequencing. These approaches will also investigate protein isoforms and post-translational modifications.
Objective 3 is to establish how selected isoforms affect protein expression, localisation, interactions, and function using cell models. This interdisciplinary project combines bioinformatics, transcriptomics, proteomics and molecular biology. It is a combined bioinformatics and wet laboratory project. The focus will be on brain and blood, not any neuropsychiatric disease or disorder – this is open for the student to take a lead on genes of interest (candidate genes are available). This project can be steered towards where the student’s talent meets interest. The project involves bioinformatic analysis - prioritising the best targets to take forward for proteomics and then functional validation.
The student will analyse long-read datasets and can incorporate single-nucleus, spatial transcriptomic and public data to select candidates for experimental validation. Full computational and laboratory training will be provided, with opportunities to work with international collaborators developing methods for long-read transcriptome–proteome analysis and integration. The student will prepare proteomics samples from postmortem human brain for mass spectrometry and Quantum-si single protein molecule sequencing. Key splicing events will be investigated in cell models using protein extraction, co-immunoprecipitation and Western blotting to determine whether the resulting proteins have distinct biological effects. The expected outcome will be a comprehensive analysis and validation of predicted novel CDS in the brain, with a better understanding of the functional consequences of these downstream and their involvement in neuropsychiatric disease and disorder.
The student will join an internationally recognised environment in brain genomics, long-read sequencing and functional genomics, with extensive datasets, advanced bioinformatics and high performance computing. The student will have access to all necessary laboratory facilities and equipment to support the project.
Funded by: EPSRC Engineering and Physical Sciences Research Council
