The emergence of complex structural organisation in skeletal muscle

During development, an organism goes from a single fertilised cell through to an adult with multiple organs. These organs are precisely shaped and the internal cellular structure tightly defined to ensure efficient function. For example, the lungs generate a large-scale branching network to enable rapid oxygen transport into the bloodstream.

In the skeletal muscle, muscle cells form elongated, and typically multinucleated, fibres that enable the generation of significant force. How complex organ shape emerges during development has been a long standing question in biology, going back to before even D'Arcy Thompson. Yet, understanding the underlying processes has long proven challenging, partially due to the difficulty of imaging the cellular processes at appropriate spatial and temporal resolution.

Recent years have seen three major advances that are helping us to tackle this challenge. First, biophysical models have proven to be very powerful in describing the structural changes in the material properties of biological materials. Developing tissues can undergo transitions, such as between fluid-like (i.e. rapid cell rearrangements) and solid-like (i.e. rigid internal structure).

Second, imaging advances mean we can record with subcellular resolution the dynamics of cell morphogenesis in living embryos. Third, there have been major steps forward in our ability to segment and quantify complex biological imaging data using machine learning and more traditional approaches. Analysis of muscle fibre formation is especially challenging due to the speed with which the cell structure changes, including cell fusion.

By combining these advances with the powerful genetics and optical accessibility of the zebrafish embryo, here we aim to dissect how the internal structural organisation of skeletal muscle emerges.

Through the following Aims, we explore if the developing skeletal muscle undergoes a material change in its properties and how cellular processes drive cell and tissue shaping:

1) Provide the first dissection of the dynamics for every cell within an internal vertebrate organ as they reach their final position and morphology. 2) Uncover the mechanisms happening within the cells that drive cell and nucleus reshaping and positioning. 3) Use suitable mutants to perturb the cellular environment to test our models of cell and tissue shaping.

In Aim 1, we will develop imaging and image analysis techniques to allow us to access the cellular behaviour throughout initial skeletal muscle development. We will track the position and morphology of every cell within each selected future muscle segment as they go from round cells through to the highly elongated and tightly structured muscle. The quantitative data is a key input into our analysis of the tissue structural order, to test if there are hallmarks of transitions in the material properties.

In Aim 2, we dissect some of the subcellular mechanisms driving the changes in cell and tissue shape. We focus on microtubules due to their importance in a range of cellular processes associated with muscle formation. We will utilise lattice light-sheet microscopy - which enables very fast imaging at high spatial resolution - to record the dynamics of microtubules and their associated motor proteins during skeletal muscle formation.

We will combine this with suitable drug and light-tuneable perturbations to dissect the role of microtubules in guiding muscle morphogenesis.

In Aim 3, we utilise a range of mutants to further explore the mechanisms driving tissue organisation. We focus on perturbing muscle cell fate specification and inhibiting muscle fusion. These perturbations allow us to access the role of both biochemical and biomechanical inputs in driving skeletal muscle formation.

Around 40% of human body mass is skeletal muscle. Using zebrafish development. we will dissect the fundamental mechanisms ensuring this tissue is precisely structured.