Spatio-temporal images of growth-factor-induced activation of Ras and Rap1

Naoki Mochizuki, Shigeko Yamashita, Kazuo Kurokawa, Yusuke Ohba, Takeharu Nagai, Atsushi Miyawaki and Michiyuki Matsuda ()
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Naoki Mochizuki: National Cardiovascular Center Research Institute
Shigeko Yamashita: Research Institute, International Medical Center of Japan
Kazuo Kurokawa: Institute for Microbial Diseases, Osaka University
Yusuke Ohba: Institute for Microbial Diseases, Osaka University
Takeharu Nagai: Laboratory for Cell Function and Dynamics, Brain Science Institute, RIKEN
Atsushi Miyawaki: Laboratory for Cell Function and Dynamics, Brain Science Institute, RIKEN
Michiyuki Matsuda: Institute for Microbial Diseases, Osaka University

Nature, 2001, vol. 411, issue 6841, 1065-1068

Abstract: Abstract G proteins of the Ras family function as molecular switches in many signalling cascades1,2,3; however, little is known about where they become activated in living cells. Here we use FRET (fluorescent resonance energy transfer)-based sensors to report on the spatio-temporal images of growth-factor-induced activation of Ras and Rap1. Epidermal growth factor activated Ras at the peripheral plasma membrane and Rap1 at the intracellular perinuclear region of COS-1 cells. In PC12 cells, nerve growth factor-induced activation of Ras was initiated at the plasma membrane and transmitted to the whole cell body. After three hours, high Ras activity was observed at the extending neurites. By using the FRAP (fluorescence recovery after photobleaching) technique, we found that Ras at the neurites turned over rapidly; therefore, the sustained Ras activity at neurites was due to high GTP/GDP exchange rate and/or low GTPase activity, but not to the retention of the active Ras. These observations may resolve long-standing questions as to how Ras and Rap1 induce different cellular responses4 and how the signals for differentiation and survival are distinguished by neuronal cells5.

Date: 2001
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DOI: 10.1038/35082594

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