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The ability of eukaryotic cells to differentiate surface stiffness is fundamental for many processes like stem cell development. Bacteria were previously known to sense the presence of surfaces, but the extent to which they could differentiate stiffnesses remained unclear. Here we establish that the human pathogen Pseudomonas aeruginosa actively measures surface stiffness using type IV pili (TFP). Stiffness sensing is nonlinear, as induction of the virulence factor regulator is peaked with stiffness in a physiologically important range between 0.1 kPa (similar to mucus) and 1,000 kPa (similar to cartilage). Experiments on surfaces with distinct material properties establish that stiffness is the specific biophysical parameter important for this sensing. Traction force measurements reveal that the retraction of TFP is capable of deforming even stiff substrates. We show how slow diffusion of the pilin PilA in the inner membrane yields local concentration changes at the base of TFP during extension and retraction that change with substrate stiffness. We develop a quantitative biomechanical model that explains the transcriptional response to stiffness. A competition between PilA diffusion in the inner membrane and a loss/gain of monomers during TFP extension/retraction produces substrate stiffness-dependent dynamics of the local PilA concentration. We validated this model by manipulating the ATPase activity of the TFP motors to change TFP extension and retraction velocities and PilA concentration dynamics, altering the stiffness response in a predictable manner. Our results highlight stiffness sensing as a shared behavior across biological kingdoms, revealing generalizable principles of environmental sensing across small and large cells.
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Competitive binding of independent extension and retraction motors explains the quantitative dynamics of type IV pili
Type IV pili (TFP) function through cycles of extension and retraction. The coordination of these cycles remains mysterious due to a lack of quantitative measurements of multiple features of TFP dynamics. Here, we fluorescently label TFP in the pathogenPseudomonas aeruginosaand track full extension and retraction cycles of individual filaments. Polymerization and depolymerization dynamics are stochastic; TFP are made at random times and extend, pause, and retract for random lengths of time. TFP can also pause for extended periods between two extension or two retraction events in both wild-type cells and a slowly retracting PilT mutant. We developed a biophysical model based on the stochastic binding of two dedicated extension and retraction motors to the same pilus machine that predicts the observed features of the data with no free parameters. We show that only a model in which both motors stochastically bind and unbind to the pilus machine independent of the piliation state of the machine quantitatively explains the experimentally observed pilus production rate. In experimental support of this model, we show that the abundance of the retraction motor dictates the pilus production rate and that PilT is bound to pilus machines even in their unpiliated state. Together, the strong quantitative agreement of our model with a variety of experiments suggests that the entire repetitive cycle of pilus extension and retraction is coordinated by the competition of stochastic motor binding to the pilus machine, and that the retraction motor is the major throttle for pilus production.
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- Award ID(s):
- 1734030
- PAR ID:
- 10214219
- Publisher / Repository:
- Proceedings of the National Academy of Sciences
- Date Published:
- Journal Name:
- Proceedings of the National Academy of Sciences
- Volume:
- 118
- Issue:
- 8
- ISSN:
- 0027-8424
- Page Range / eLocation ID:
- Article No. e2014926118
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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