Research Article

Tubular clathrin/AP-2 lattices pinch collagen fibers to support 3D cell migration

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Science  16 Jun 2017:
Vol. 356, Issue 6343, eaal4713
DOI: 10.1126/science.aal4713

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Helping a cell to migrate in 3D space

Clathrin-coated pits are well known to be involved in receptor-mediated endocytosis. Independent of their role in endocytosis, Elkhatib et al. observed that clathrin-coated structures strongly accumulated along collagen fibers in migrating cells. Clathrin-coated structures assembled on and then partially wrapped around and pinched the fibers. In a three-dimensional (3D) network, this mechanism provided multiple anchoring points along cellular protrusions. In the absence of clathrin-coated structures, protrusions were shorter and migration was impaired. This mode of adhesion may cooperate with classical focal adhesions to help cancer cells move in a 3D environment.

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Structured Abstract


Migrating cells need to adhere to their environment in order to pull themselves forward. Adhesion to the extracellular matrix (ECM), including collagen fibers, is achieved by integrins that cluster in focal adhesions (FAs). Integrin clustering is a key process during migration because it increases the cell’s avidity for the ECM. The physical connection between the cellular traction machinery and the ECM that occurs at FAs allows the cell to pull on the substratum, which generates forces that are required for locomotion both in two-dimensional (2D) and 3D environments.


FAs are numerous and prominent when cells migrate on 2D, stiff substrata. However, cells migrating on softer 2D or in 3D environments show fewer and/or seemingly weaker FAs. In addition to FAs, integrins have been shown to accumulate in other structures of the plasma membrane such as clathrin-coated structures (CCSs). CCSs are professional cell surface receptor–clustering machineries that also bend the membrane to progressively pack these receptors into endocytic clathrin-coated vesicles that bud into the cytosol. Although the structure, dynamics, and functions of CCSs have been extensively described in cells seeded on 2D environments, little is known about their features in more physiological soft, 3D conditions. We analyzed the dynamics of CCSs in cells migrating in a 3D environment of collagen fibers.


We observed that CCSs accumulated along collagen fibers in the 3D environment. This was the result of an increased nucleation rate as well as a prolonged lifetime of CCSs at cell–collagen fiber contact sites. Both local plasma membrane bending, resulting from the physical contact with collagen fibers, and local integrin engagement triggered CCS accumulation on fibers. Electron microscopy analyses revealed that the CCSs engaged with collagen fibers adopted a distinct, tubular morphology to wrap around and pinch the fiber. Surprisingly, clathrin was not required for the coat of clathrin-adaptors, which includes adaptor protein 2 (AP-2) and Dab2, to accumulate on and wrap around fibers. We named these particular type of CCSs tubular clathrin/AP-2 lattices (TCALs). Cell adhesion and the cell’s capacity to grab collagen fibers were inhibited by disruption of TCALs or by inhibition of integrin accumulation at TCALs by using AP-2 and Dab2 siRNAs, respectively. However, in agreement with our morphological analysis, clathrin itself was not required for cells to adhere to fibers, demonstrating that this process is independent of CCSs’ role in endocytosis. FAs were mostly found at both extremities of elongated cells migrating in the 3D environment, whereas CCSs were distributed all over the plasma membrane of cellular protrusions. By laser ablating the tip of individual cellular extensions, we demonstrated that TCALs stabilize protrusions and thereby promote cell migration, in an endocytosis-independent manner.


Thus, in soft, 3D environments, CCSs promote cell migration not only through the regulation of endocytosis but also by providing anchoring points to collagen fibers. This is necessary for the cell to cope with the tension exerted at FAs, which allows cells to develop long protrusions and to migrate efficiently. We conclude that TCALs represent a mode of cell adhesion that, in coordination with FAs, supports cell migration in 3D.

Structural and functional analysis of TCALs at cell–collagen fiber contact sites.

Cells migrating in a 3D network of collagen fibers (red) adopt an elongated morphology as they extend protrusions in between fibers in order to move forward. CCSs (green) accumulate on and pinch collagen fibers, thus providing local anchoring points to the environment. This allows the cell to stabilize long extensions that are needed to migrate efficiently.


Migrating cells often use focal adhesions in order to move. Focal adhesions are less prominent in cells migrating in three-dimensional (3D) as compared with 2D environments. We looked for alternative adhesion structures supporting cell migration. We analyzed the dynamics of clathrin-coated pits in cells migrating in a 3D environment of collagen fibers. Both topological cues and local engagement of integrins triggered the accumulation of clathrin-coated structures on fibers. Clathrin/adaptor protein 2 (AP-2) lattices pinched collagen fibers by adopting a tube-like morphology and regulated adhesion to fibers in an endocytosis-independent manner. During migration, tubular clathrin/AP-2 lattices stabilized cellular protrusions by providing anchoring points to collagen fibers. Thus, tubular clathrin/AP-2 lattices promote cell adhesion that, in coordination with focal adhesions, supports cell migration in 3D.

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