Dynamic filopodia are required for chemokine-dependent intracellular polarization during guided cell migration in vivo.

Cell migration and polarization is controlled by signals in the environment. Migrating cells typically form filopodia that extend from the cell surface, but the precise function of these structures in cell polarization and guided migration is poorly understood. Using the in vivo model of zebrafish primordial germ cells for studying chemokine-directed single cell migration, we show that filopodia distribution and their dynamics are dictated by the gradient of the chemokine Cxcl12a. By specifically interfering with filopodia formation, we demonstrate for the first time that these protrusions play an important role in cell polarization by Cxcl12a, as manifested by elevation of intracellular pH and Rac1 activity at the cell front. The establishment of this polarity is at the basis of effective cell migration towards the target. Together, we show that filopodia allow the interpretation of the chemotactic gradient in vivo by directing single-cell polarization in response to the guidance cue.

Cxcl12 evolution--subfunctionalization of a ligand through altered interaction with the chemokine receptor.

The active migration of primordial germ cells (PGCs) from their site of specification towards their target is a valuable model for investigating directed cell migration within the complex environment of the developing embryo. In several vertebrates, PGC migration is guided by Cxcl12, a member of the chemokine superfamily. Interestingly, two distinct Cxcl12 paralogs are expressed in zebrafish embryos and contribute to the chemotattractive landscape. Although this offers versatility in the use of chemokine signals, it also requires a mechanism through which migrating cells prioritize the relevant cues that they encounter. Here, we show that PGCs respond preferentially to one of the paralogs and define the molecular basis for this biased behavior. We find that a single amino acid exchange switches the relative affinity of the Cxcl12 ligands for one of the duplicated Cxcr4 receptors, thereby determining the functional specialization of each chemokine that elicits a distinct function in a distinct process. This scenario represents an example of protein subfunctionalization--the specialization of two gene copies to perform complementary functions following gene duplication--which in this case is based on receptor-ligand interaction. Such specialization increases the complexity and flexibility of chemokine signaling in controlling concurrent developmental processes.

A role for Rho GTPases and cell-cell adhesion in single-cell motility in vivo.

Cell migration is central to embryonic development, homeostasis and disease, processes in which cells move as part of a group or individually. Whereas the mechanisms controlling single-cell migration in vitro are relatively well understood, less is known about the mechanisms promoting the motility of individual cells in vivo. In particular, it is not clear how cells that form blebs in their migration use those protrusions to bring about movement in the context of the three-dimensional cellular environment. Here we show that the motility of chemokine-guided germ cells within the zebrafish embryo requires the function of the small Rho GTPases Rac1 and RhoA, as well as E-cadherin-mediated cell-cell adhesion. Using fluorescence resonance energy transfer we demonstrate that Rac1 and RhoA are activated in the cell front. At this location, Rac1 is responsible for the formation of actin-rich structures, and RhoA promotes retrograde actin flow. We propose that these actin-rich structures undergoing retrograde flow are essential for the generation of E-cadherin-mediated traction forces between the germ cells and the surrounding tissue and are therefore crucial for cell motility in vivo.