Abstract
Recent technical advances have broadened our understanding of processes that govern mammalian cell migration in health and disease but many of the molecular and morphological alterations that precede and accompany movement of cells - in particular in three-dimensional (3D) environments - are still incompletely understood. In this manuscript, using high-resolution and time-lapse microscopy imaging approaches, we describe morphodynamic processes during rounded/amoeboid cell invasion and molecules associated with the cellular invasion structures. We used macrophages infected with the intracellular protozoan parasite Theileria annulata, which causes Tropical Theileriosis in susceptible ruminants such as domestic cattle. T. annulata transforms its host cell that, as a result, acquires many characteristics of human cancer cells including a markedly increased potential to migrate, disseminate and expand in the body of the host animal. Hence, virulence of the disease is associated with the capability of infected cells to disseminate inside the host. Using T. annulata-transformed macrophages as a model system, we described a novel mode of rounded/amoeboid macrophage migration. We show that filopodia-like membrane extensions at the leading edge lead the way and further evolve in blebbing membrane protrusions to promote progressive expansion of the matrix. Associated with focal invasion structures we detected ezrin, radixin, moesin-family proteins and their regulatory kinase MAP4K4. Furthermore, we linked Rho-kinase activity to contractile force generation, which is essential for infected cell motility. Thus, the motility mode of these parasite-transformed macrophages contrasts with those described so far in human macrophages such as the tunneling or mesenchymal modes, which require engulfment, compaction and ingestion of matrix or proteolytic matrix degradation, respectively. Together, our data reveal protrusion dynamics at the leading edge of invading cells in 3D at unprecedented temporal and spatial resolution and suggest a novel mode of rounded/amoeboid invasive cell motility that exploits actin-driven filopodia formation in combination with pressure-driven membrane blebs.