Subcellular localization and oligomerization of the Arabidopsis thaliana somatic embryogenesis receptor kinase 1 protein

The Arabidopsis thaliana somatic embryogenesis receptor kinase 1 (AtSERK1) gene is expressed in developing ovules and early embryos. AtSERK1 is also t ransiently expressed during somatic embryogenesis. The predicted AtSERK1 pr otein contains an extracellular domain with a leucine zipper motif followed by five leucine-rich repeats, a proline-rich region, a single transmembran e region and an intracellular kinase domain. The AtSERK1 cDNA was fused to two different variants of green fluorescent protein (GFP), a yellow-emittin g GFP (YFP) and a cyan-emitting GFP (CFP), and transiently expressed in bot h plant protoplasts and insect cells. Using confocal laser scanning microsc opy it was determined that the AtSERK1-YFP fusion protein is targeted to pl asma membranes in both plant and animal cells. The extracellular leucine-ri ch repeats, and in particular the N-linked oligosaccharides that are presen t on them appear to be essential for correct localization of the AtSERK1-YF P protein. The potential for dimerization of the AtSERK1 protein was invest igated by measuring the YFP/CFP fluorescence emission ratio using fluoresce nce spectral imaging microscopy. This ratio will increase due to fluorescen ce resonance energy transfer if the AtSERK1-CFP and AtSERK1-YFP fusion prot eins interact. LII 15% of the cells the YFP/CFP emission ratio for plasma m embrane localized AtSERK1 proteins was enhanced. Yeast-protein interaction experiments confirmed the possibility for AtSERK1 homodimerization. Elimina tion of the extracellular leucine zipper domain reduced the YFP/ CFP emissi on ratio to control levels indicating that without the leucine zipper domai n AtSERK1 is monomeric. (C) 2001 Academic Press.