Live-cell imaging studies of collective cell migration and tissue elongation using Drosophila egg chamber.

摘要

Drosophila oogenesis is a powerful model for the study of fundamental questions in cell and developmental biology including stem cell, collective cell migration, mRNA transportation, and epithelium morphogenesis. In addition to its longstanding value as a genetically tractable model, it has recently emerged as an excellent system for live imaging. My graduate studies used both genetic and microscopic techniques to study cellular activities in living tissue, and includes two parts: the first is the study of collective guidance of border cell migration using a photoactivatable form of the small GTPase Rac and a Rac FRET biosensor; secondly we used live-cell imaging and genetic manipulation to discover and characterize basal actomyosin oscillations that represent a novel mechanism of tissue elongation.;Collective cell migration is an important process both physiology and pathological conditions. The small GTPase Rac is an important regulator of actin polymerization and is required for most cell migrations. However, global activation or inhibition of Rac activity blocks collective border cell migration suggesting that temporal and/or spatial regulation of Rac activity is essential. Using a recently developed photoactivatable Rac (PA-Rac), which allows rapid, focal, and reversible activation or inactivation of Rac using light, we found local activation of Rac was sufficient to redirect the whole border cell cluster and rescue the loss-of-guidance phenotype. While inactivation of Rac in one cell of the cluster caused a dramatic response in the other cells, suggesting that the cells sense migratory direction as a group based on the relative levels of Rac activity. Using a Rac FRET biosensor reporter, Rac activity was found to be higher in the leading cell than the followers. Taken together, these studies show that border cells sense direction collectively based on relative levels of Rac activity. Understanding how molecular dynamics lead to cellular behaviors that ultimately sculpt organs and tissues is a major challenge not only in basic developmental biology but also in tissue engineering and regenerative medicine. Using live imaging, we found that the basal surfaces of Drosophila follicle cells undergo a series of directional, oscillating contractions driven by periodic myosin accumulation on a polarized actin network. Inhibition of the actomyosin contractions or their coupling to extracellular matrix (ECM) blocked elongation of the whole tissue, whereas enhancement of the contractions exaggerated it. This Myosin contraction was regulated by the small GTPase Rho, ROCK and cytosolic calcium. Disrupting the link between the actin cytoskeleton and the ECM decreased, while enhancing cell-ECM adhesion increased, the amplitude and period of the contractions. In contrast, disrupting cell-cell adhesions resulted in loss of the actin network. Our findings reveal a novel mechanism controlling organ shape and a new model for the study of the effects of oscillatory actomyosin activity within a coherent cell sheet.