Fluorescence Fluctuation Analysis of AtSERK and BRI1 Oligomerization

¹ Max Planck Institute for Physiology
² Harvard Medical School
³ Ghent University
⁴ Wageningen University

^* To whom correspondence should be addressed. E-mail: ton.visser{at}wur.nl.

Submitted on May 3, 2007
Revised on June 8, 2007
Accepted on 31 August 2007

	Abstract

Receptor kinases play a key role in the cellular perception of signals. To verify models for receptor activation through dimerization, an experimental system is required to determine the precise oligomerization status of proteins within living cells. Here we show that photon counting histogram analysis and dual-color fluorescence cross-correlation spectroscopy is able to monitor fluorescently labeled proteins at the single-molecule detection level in living plant cells. In-frame fusion proteins of the Brassinosteroid Insensitive 1 (BRI1) receptor and the Arabidopsis thaliana Somatic Embryogenesis Receptor-like Kinases 1 and 3 (AtSERK1 & 3) to the enhanced cyan or yellow fluorescent protein were transiently expressed in plant cells. Although no oligomeric structures were detected for AtSERK3, fifteen (AtSERK1) to twenty (BRI1) percent of the labeled proteins in the plasma membrane was found to be present as homodimers, while no evidence was found for higher oligomeric complexes.

Key Words: FCCS, FCS, Oligomerization, PCH, Receptor-like kinases

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