Real-time centrosome reorientation during fibroblast migration.
The centrosome is positioned between the nucleus and the leading edge of many types of migrating cells. Cdc42 regulates this centrosome reorientation through its effectors Par6 and MRCK. Using time-lapse microscopy of live cells, the mechanisms and kinetics of centrosome reorientation can be studied. In this chapter, we describe a modification in the standard wound healing assay that allows the study of signaling pathways involved in centrosome reorientation and other polarization events that occur before cell migration. We also describe a method for visualization of centrosome reorientation by time-lapse microscopy using NIH 3T3 fibroblasts stably transfected with GFP-tubulin.
Department of Anatomy & Cell Biology, Columbia University, New York, NY, USA.
Edgar R
ER
Gomes
Gregg G
GG
Gundersen
D000818:Animals; D002465:Cell Movement / Q000502:physiology*; D018385:Centrosome / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D049452:Green Fluorescent Proteins / Q000235:genetics; D008055:Lipids / Q000494:pharmacology; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods; D041681:NIH 3T3 Cells; D014162:Transfection; D014404:Tubulin / Q000235:genetics; D014945:Wound Healing / Q000502:physiology*
D008055:Lipids; C086724:Lipofectamine; D014404:Tubulin; D049452:Green Fluorescent Proteins