Primary Supervisor: Dr Timothy Saunders, WMS
Secondary supervisor: Dr Sascha Ott
PhD project title: Systems approach to dissecting the processes underlying skeletal muscle fibre formation
University of Registration: University of Warwick
Background: How does complex biological form first emerge? Organs have specific shapes that are crucial for efficient function. Understanding the mechanisms driving organ formation and maintenance has potential for wide impact, including in regeneration, wound healing and ageing.
Vertebrate skeletal muscle development is a highly conserved process. Each new segment (myotome) of muscle is generated in a clock-like fashion.1 Muscle fibres in the myotome can form multinucleated fast-twitch fibres (through cell fusion, whereby two fast fibres combine their membranes) or mononucleated slow-twitch fibres. These fibres have different physiological properties. Recently, the signalling molecules and networks that define these different cell populations have been identified.2
Muscle fibres undergo remarkable morphological changes during development. They elongate in a highly-polarised manner and cell fusion leads to highly coordinated cell and tissue transformations. Yet, how such large-scale morphological processes are dynamically regulated remains largely unknown. In particular, (1) what are the genetic and mechanical interactions that ensure proteins for cell elongation and fusion are expressed in the correct cells at the right time; and (2) can we relate the cell genetic state with its mechanical state during muscle formation?
Model system: Tackling these problems requires an in vivo model, as the whole system is important. We use zebrafish, which is easy to image during development and amenable to drug and genetic perturbations. This contrasts with mouse, where the developing embryo cannot be imaged in vivo. Zebrafish lay eggs in large numbers, facilitating -omic approaches.
Project: We will combine the power of single cell sequencing4 with live imaging to tackle the above questions. We will collaborate with Sascha Ott at the single cell sequencing platform at Warwick to dissect the transcriptional state of the developing myotome. Recent advances in single cell sequencing enable accurate sequencing with relatively few samples (~ 1000 cells). An advantage of the developing myotome is that it effectively has its own internal clock; each new myotome segment is specified 30 minutes (in zebrafish) after the preceding segment. Therefore, from sequencing neighbouring myotome segments we can extract temporal information about how the transcriptional state of the myotome evolves; these maps will have greater temporal resolution than those derived from typical single cell transcriptomic analysis. We will also complement this with ATAC-seq, to explore the changes in chromatin accessibility during myotome development.
Using the sequencing results, we will generate maps of cellular states in both space and time. We will focus on cellular expression of fusion-related proteins and ones playing a role in cell shape. We will complement analysis of wild-type embryos with suitable -omic analysis of mutants that alter cell fate.2
In the second part, we will link the maps of cell fate to single cell tracking from live imaging. We will follow the position and morphology of cells as they develop within the myotome. We will quantify the “morphospace” of the muscle fibres. By combining single-cell transcriptomics with tracking and quantitative cell shape information, we will generate spatial and temporal predictions for potential targets that are crucial in the formation of different muscle fibre morphologies. We will then perform targeted perturbations combined with live imaging to test these predictions, leading to novel insights into how skeletal muscle is generated.
Overall, this is an exciting interdisciplinary project combining systems biology and live imaging, which is suitable for a student looking to apply the latest systems biology approaches to important questions in development.
1 Soroldoni et al., Science 345, 222 (2014)
2 Jackson & Ingham, Mech Dev 130, 447 (2013)
3 Millay et al., Nature 499, 301 (2013)
3 Wagner & Klein, Nat Rev Gen 21, 410 (2020)
BBSRC Strategic Research Priority: Understanding the Rules of Life: Systems Biology
Techniques that will be undertaken during the project:
- Single cell transcriptomics
- Computational systems biology
- Live microscopy imaging
- Image analysis and quantification
- Zebrafish husbandry and genetics
Contact: Dr Timothy Saunders, University of Warwick