Traditionally, a cell’s nucleus has been regarded as a passive organelle that stores and protects the genome, facilitating transcription and replication. However, breakthrough studies have revealed it is a dynamic structure and mechanical sensor for physical cues within the tissue microenvironment.
The overall objective of this project is to image nuclear dynamics for the first time in living, intact hearts to gain insight into understanding how forces that arise from outside the nucleus (extrinsic) and inside the nucleus (intrinsic) contribute to nuclear deformation and function. The importance of understanding this in normal tissue is highlighted by the various diseases that are caused by defects in the nuclear envelope that surrounds the nucleus.
To achieve this objective, live imaging on beating hearts using state-of-the-art 3D light-sheet imaging will be performed. Once established, hearts from different mutant models of nuclear envelope dysfunction will be employed and nuclear dynamics extracted. Furthermore, acute drug addition will be used to modulate muscle contraction and chromatin organization in beating mammalian hearts.
To address the main objective, the project will involve 3 highly inter-disciplinary approaches:
1) Bioengineering/ biophysics: Optimization of light-sheet imaging platform to image beating hearts and nuclear dynamics in real-time and in 3D.(Years 1-2)
2) In vivo and ex vivo approaches: Surgical techniques to extract intact fetal mouse hearts, establishing ex vivo tissue culture and mouse genetics and genotyping.(Years 2-3)
3) Imaging acquisition and post-acquisition software development: Development of code on Matlab/ Fiji software to enable image analysis of nuclear dynamics and automated extraction of nuclear shape/ deformation / strain parameters.(Years 2-4)