Chapter 14: Centrosome Dynamics in Living Cells

Abstract

Publisher Summary Green fluorescent protein (GFP) is a bioluminescent protein encoded by a gene from the jellyfish, Aequorea victoria. The excitation and emission profile of GFP is similar to that of fluorescein, rendering a chimeric protein visible by standard epi-fluorescence microscopy; filter sets optimized for GFP spectra can be used to image proteins at low light levels. The ability to produce fluorescent chimeric proteins in cells simply by molecular cloning techniques has revolutionized the study of proteins and cellular processes in vivo . This chapter focuses on GFP-based technology to study centrosomes in live cells. Centrosomes are small organelles located at the focus of microtubules where they play a key role in the organization of microtubule arrays for a number of cellular functions including cellular organization, morphogenesis, and mitosis. Through their ability to nucleate the growth of microtubules, centrosomes control the geometry, polarity, and number of cellular microtubules, and to a large extent, the microtubule distribution in the cell. The chapter also describes how GFP-pericentrin can be used in combination with a recently developed system for image capture and a new algorithm for image restoration, to obtain high-resolution three-dimensional images of centrosomes in living cells.