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Abstract Mechanosensitive mechanisms are often used to sense damage to tissue structure, stimulating matrix synthesis and repair. While this kind of mechanoregulatory process is well recognized in eukaryotic systems, it is not known whether such a process occurs in bacteria. InVibrio cholerae, antibiotic-induced damage to the load-bearing cell wall promotes increased signaling by the two-component system VxrAB, which stimulates cell wall synthesis. Here we show that changes in mechanical stress within the cell envelope are sufficient to stimulate VxrAB signaling in the absence of antibiotics. We applied mechanical forces to individual bacteria using three distinct loading modalities: extrusion loading within a microfluidic device, direct compression and hydrostatic pressure. In all cases, VxrAB signaling, as indicated by a fluorescent protein reporter, was increased in cells submitted to greater magnitudes of mechanical loading, hence diverse forms of mechanical stimuli activate VxrAB signaling. Reduction in cell envelope stiffness following removal of the endopeptidase ShyA led to large increases in cell envelope deformation and substantially increased VxrAB response, further supporting the responsiveness of VxrAB. Our findings demonstrate a mechanosensitive gene regulatory system in bacteria and suggest that mechanical signals may contribute to the regulation of cell wall homeostasis. 

Bacteria experience substantial physical forces in their natural environment including forces caused by osmotic pressure, growth in constrained spaces, and fluid shear. The cell envelope is the primary load-carrying structure of bacteria, but the mechanical properties of the cell envelope are poorly understood; reports of Young’s modulus of the cell envelope of E. coli are widely range from 2 MPa to 18 MPa. We have developed a microfluidic system to apply mechanical loads to hundreds of bacteria at once and demonstrated the utility of the approach for evaluating whole-cell stiffness. Here we extend this technique to determine Young’s modulus of the cell envelope of E. coli and of the pathogens V. cholerae and S. aureus. An optimization-based inverse finite element analysis was used to determine the cell envelope Young’s modulus from observed deformations. The Young’s modulus of the cell envelope was 2.06±0.04 MPa for E. coli, 0.84±0.02 MPa for E. coli treated with a chemical known to reduce cell stiffness, 0.12±0.03 MPa for V. cholerae, and 1.52±0.06 MPa for S. aureus (mean ± SD). The microfluidic approach allows examining hundreds of cells at once and is readily applied to Gram-negative and Gram-positive organisms as well as rod-shaped and cocci cells, allowing further examination of the structural causes of differences in cell envelope Young's modulus among bacteria species and strains. 

Engineered living materials (ELMs) are an emerging class of materials that are synthesized and/or populated by living cells to achieve novel functionalities including self‐healing and sensing. Providing nutrients to living cells within an ELM over prolonged periods remains a major technical challenge that limits the service life of ELMs. Bone maintains living cells for decades by delivering nutrients through a network of nanoscale channels punctuated by microscale pores. Nutrient transfer in bone is enabled by mechanical loading experienced by the material during regular use. Herein, the geometric traits of the network of channels and pores that can be used in ELMs to allow mechanical loading to enable nutrient delivery to resident cell populations are identified in a manner seen in bone. Transport occurs when deformation in the microscale pore network exceeds the volume of the connecting channels. Computational models show that transport is enhanced at greater loading magnitudes and lower loading frequencies. The computational results are confirmed using experiments with microfluidic systems. In the findings, quantitative design principles are provided for channel‐pore networks capable of sustained delivery of nutrients to living cells within materials.