In preprints: unpicking chemical and mechanical crosstalk for axonal pathfinding Free

To better understand the direct effect of altering either mechanical or chemical signals, systems are required where tissues can be manipulated and specific responses measured in a controlled environment. Here, Pillai, Mukherjee et al. excise and embed embryonic brain tissue in both soft and stiff hydrogels to externally perturb the mechanical environment of the tissue (Pillai et al., 2025 preprint). This isolates the tissue from potential variable mechanical and chemical cues from the rest of the brain and provides a known and controllable external environment. By combining this technique with hybridization chain reaction-fluorescence in situ hybridization (HCR-FISH) to observe the expression of chemical signals in the tissue, the authors show that, ex vivo, increased environmental stiffness is sufficient to induce expression of the chemical guidance cues Sema3A and Slit1 (Pillai et al., 2025 preprint).

This result is corroborated in vivo by locally exploiting compression-stiffening of brain tissue using an atomic force microscopy (AFM) probe. Compression-stiffening has been previously observed in both healthy and cancerous mouse brain tissue, whereby uniaxial compression increases the resistance of the tissue to shear forces (Pogoda et al., 2014). Pogoda et al. suggest from this that stiffness changes in brain tumours could arise from pressure gradients within the tissue (Pogoda et al., 2014). In this preprint, Pillai, Mukherjee et al. exploit this phenomenon by applying a prolonged 30 nN compressive force to stiffen soft Xenopus brain tissue. They show that stiffening tissue in this way is sufficient to induce Sema3A expression in brain regions which do not usually express Sema3A. This requires the presence of Piezo1 to communicate the compression stiffening into a chemical change (Pillai et al., 2025 preprint).

Combining in vivo and ex vivo approaches in this way helps to provide an understanding of how mechanical signals and chemical cues interact in both controlled environments and within the complexity of a living tissue. Ultimately, used in tandem they enable more accurate insights into the mechanisms underlying tissue response and gene expression.

Piezo1 plays a central role in this study. Highlighted initially as the core mediator translating mechanical cues into chemical signals, it is also revealed to be a manipulator of the mechanical environment. Its knockdown in brain tissue affects both the expression of the chemical signals Slit1 and Sema3A (by reducing them) and the stiffness of the tissue (by softening it). Interestingly, the effect of Piezo1 on chemical signals appears to occur through mechanical perturbation. Knockdown of Piezo1 via morpholino injection leads to softer tissues by depleting the cell-cell adhesion molecules NCAM1 and N-Cadherin, while targeted reduction of these molecules specifically reduces tissue stiffness and Sema3A expression. Conversely, tissue stiffening increases the expression of Sema3A, with Piezo1 as a necessary protein for this chemical response (Pillai et al., 2025 preprint). Thus, Piezo1 is thrust into the spotlight as a manipulator of the mechanical environment as well as a mediator linking mechanical changes to chemical signalling.

The work of Pillai, Mukherjee et al. highlights the co-dependence of the mechanical environment and chemical signalling, though their influences are not always reciprocal. The next step to unlocking this dynamic interplay between mechanics and signalling is to assess where and how bidirectional feedback occurs. Overall, this work adds to the literature evidencing the importance of the mechanical environment for cellular processes and shows how local mechanical changes can be translated into much longer-range chemical cues. The ability of a tissue to read (via Piezo1) and respond (via chemical signalling) to its mechanical environment is vital for normal development. Indeed, Piezo1 is required in both the brain tissue and the RGC axons themselves for successful axon pathfinding along the contralateral brain surface (Pillai et al., 2025 preprint). Erroneous axon pathfinding has been linked to numerous congenital human disorders affecting the nervous system and the development of scoliosis (Nugent et al., 2012). Thus, correctly responding to and maintaining the mechanical and chemical environment is crucial for development.

Given that both the Slit and Semaphorin families of proteins have roles in cell proliferation (Blockus and Chédotal, 2016) and motility (Blockus and Chédotal, 2016; Curreli et al., 2016), this study could also provide insights into cancers such as breast cancer, where one of the highest risk factors is changes in the mechanical environment (Yaghjyan et al., 2011). Indeed, Slit overexpression has been shown to suppress tumour growth and its expression is reduced in many types of cancer (Marlow et al., 2008), a role which is similarly adopted by semaphorins (Neufeld et al., 2016). The ability of the mechanical environment to affect the expression of these factors via Piezo1 may therefore provide insights into disease state and progression.