BACKGROUND:​

 

A growing body of evidence shows that cells respond to both biochemical signals and mechanical signals. However, how cells integrate information from both types of cues remains unclear. During my PhD, I investigated how developing neurons balance information from both biochemical and mechanical guidance cues while connecting to the right place at the right time, a process called axon pathfinding. Specifically, I focused on the role of tissue stiffness, a mechanical signal, in the regulation of Eph/ephrin signalling, a biochemical signalling pathway, in the African clawed frog (Xenopus laevis) embryonic optic pathway. I found that environmental stiffness is a critical regulator of contact-dependent cell–cell communication via EphB/ephrinB signalling.

 

These findings challenge the current dogma in the field that guidance cue expression is exclusively regulated by chemical signals. Instead, the expression of the biochemical signals responsible for correct axon pathfinding in the X. laevis optic pathway is modulated by the stiffness of the tissues surrounding the developing neurons. Eph/ephrin signalling is a highly conserved pathway involved in the formation of topographic maps, tissue boundaries, and epithelial-to-mesenchymal transitions. As its mechanical regulation is currently poorly understood, this close link between tissue mechanics and Eph/ephrin signalling is also likely to improve our understanding of the development of many organ systems as well as cancer progression.

 

The preprint is available on bioRxiv: Sipkova, J. & Franze, K. (2024) Eph/ephrin signalling in the developing brain is regulated by tissue stiffness. doi: 10.1101/2024.02.15.580461

​

 

METHODOLOGY:​

 

I used methods from classical neuroscience (growth cone collapse assay), embryology (retinal explant culture; fluorescence in situ hybridisation: HCR RNA-FISH), and materials science (in vivo atomic force microscopy; 2D and 3D soft hydrogel fabrication) to simultaneously study chemical and mechanical signalling. 

 

RESULTS:​

 

My findings can be summarised in three main points:

 

• The response of neurons to guidance molecule ephrinB1 depends on the mechanical properties of the environment in vitro.

• The patterns of tissue stiffness and expression of some EphBs and ephrinBs highly correlate during development in vivo. Here I show the mean tissue stiffness and mRNA levels of EphB2 across the optic tectum, the visual area of the frog brain, as it is innervated by retinal neurons. They are highly positively correlated.

​

​

​

​

​

​

​

​

​

​

​

​

​

​

​

 

​

• Local tissue stiffening triggers the ectopic expression of EphB in the developing brain in vivo. Since brain tissue stiffens under compression, I could locally and precisely stiffen the softer anterior optic tectum using AFM, fix the embryos, and then quantify EphB2 mRNA expression levels in the optic tectum using HCR RNA-FISH.​​​​​​​

​

​

​

​

​

​

​

​

Taken together, these results provide evidence for instructive mechanochemical coupling in vivo.

​​

​

​​​​​​​​​​​​FUTURE WORK: 

​

If I had another PhD to spend on this research question, I would focus on the molecular underlying how tissue stiffness regulates Eph/ephrin signalling. Is it through regulation of upstream pathways, mechanosensitive ion channels, or secondary messenger levels? Or perhaps, since both Ephs and ephrins are transmembrane proteins, they can directly sense changes in the mechanical properties of the environment through, for example, the physical state of the plasma membrane?

​

​

​

FUNDED BY:

​​​​

Wellcome Trust Developmental Mechanisms PhD programme 

​

​

​

​

​

​

​

CONTACT: 

​

​