An actin length threshold regulates adhesion maturation at the lamellipodium/lamellum interface.

The mechanical coupling between adherent cells and their substrates is a major driver of downstream behavior. This coupling relies on the formation of adhesion sites and actin bundles. How cells generate these elements remains only partly understood. A potentially important mechanism, the length threshold maturation (LTM), has previously been proposed to regulate adhesion maturation and actin bundle stabilization tangential to the leading edge. The LTM describes the process by which cells integrate lamellar myosin forces to trigger adhesion maturation. These forces, cumulated over the length of an actin bundle, are balanced at the anchoring focal complexes. When the bundle length exceeds a certain threshold, the distributed lamellar forces become sufficient to trigger the stabilization of the bundle and its adhesions. In this continuing study, we experimentally challenge the LTM for the first time, by seeding cells on micropatterned substrates with various non-adhesive gaps designed to selectively trigger the LTM. While stable actin bundles were observed on all patterns, their lengths were almost exclusively above 3 µm or 4 µm depending on the cell type. Furthermore, the frequency with which gaps were bridged increased nearly as a step function with increasing gap width, indicating a substrate dependent behavioral switch. These combined observations point strongly to LTM with a threshold above 3 µm. We thus experimentally confirm with two cell types our previous theoretical work postulating the existence of a length dependent threshold mechanism that triggers adhesion maturation and actin bundle stabilization.

Numerically bridging lamellipodial and filopodial activity during cell spreading reveals a potentially novel trigger of focal adhesion maturation.

We present a novel approach to modeling cell spreading, and use it to reveal a potentially central mechanism regulating focal adhesion maturation in various cell phenotypes. Actin bundles that span neighboring focal complexes at the lamellipodium-lamellum interface were assumed to be loaded by intracellular forces in proportion to bundle length. We hypothesized that the length of an actin bundle (with the corresponding accumulated force at its adhesions) may thus regulate adhesion maturation to ensure cell mechanical stability and morphological integrity. We developed a model to test this hypothesis, implementing a "top-down" approach to simplify certain cellular processes while explicitly incorporating complexity of other key subcellular mechanisms. Filopodial and lamellipodial activities were treated as modular processes with functional spatiotemporal interactions coordinated by rules regarding focal adhesion turnover and actin bundle dynamics. This theoretical framework was able to robustly predict temporal evolution of cell area and cytoskeletal organization as reported from a wide range of cell spreading experiments using micropatterned substrates. We conclude that a geometric/temporal modeling framework can capture the key functional aspects of the rapid spreading phase and resultant cytoskeletal complexity. Hence the model is used to reveal mechanistic insight into basic cell behavior essential for spreading. It demonstrates that actin bundles spanning nascent focal adhesions such that they are aligned to the leading edge may accumulate centripetal endogenous forces along their length, and could thus trigger focal adhesion maturation in a force-length dependent fashion. We suggest that this mechanism could be a central "integrating" factor that effectively coordinates force-mediated adhesion maturation at the lamellipodium-lamellum interface.