Role of Rho GTPases During Primordial Germ Cell Migration in Zebrafish

Abstract

Cell migration is an essential biological process during development, adult homeostasis, and disease. Actin cytoskeleton plays crucial role in generating forces required for cell migration. Rho GTPases are key regulators of actin cytoskeleton remodeling during cell migration. To understand the function of Rho GTPases it is important to have the information regarding their spatial-temporal activation pattern in migrating cell. Accessibility of cells migrating in vivo has proven to be a major challenge in studying molecular mechanisms underlying cell migration in their natural milieu. Primordial germ cells (PGCs) in zebrafish arise at distinct positions of the developing embryo and migrate towards their target guided by the chemokine SDF-1a. PGC migration in zebrafish is an excellent in vivo model system to study cell migration processes in vertebrates. It was shown that PGC migrate by forming blebs at their leading edge. The present work focuses at elucidating the role of Rho GTPases in actin cytoskeleton remodeling during PGC migration in zebrafish. To this end I studied actin cytoskeleton dynamics in the presence of mutant forms for three important Rho GTPases: Rac, Cdc42 and RhoA. To study the spatial-temporal activation pattern for Rho GTPases in migrating PGCs, I optimized ratio imaging with FRET biosensors in zebrafish embryo. Utilizing FRET biosensors I showed that Rac and RhoA activities are elevated at the front of the migrating PGC, while Cdc42 is active in the nucleus. Imaging actin cytoskeleton in PGCs revealed that depending on Rac activity, actin accumulates at the front of the cell forming a dense network, which then flows backwards in the process that requires RhoA activity. I propose that in migrating PGCs, Rac function promotes actin accumulation at the front of the cell, while RhoA mediates the actin backward flow thereby generating forces essential for cell locomotion.