Detection of presenilin-1 homodimer formation in intact cells using fluorescent lifetime imaging microscopy
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- 作者：Lauren Herl ; Alberto Lleo ; Anne V. Thomas ; Andrew C. Nyborg ; Karen Jansen ; Todd E. Golde ; Bradley T. Hyman and Oksana Berezovska
- 关键词：Presenilin ; γ ; -Secretase ; FLIM ; Protein– ; protein interactions ; Dimerization ; Alzheimer’ ; s disease
- 刊名：Biochemical and Biophysical Research Communications
- 出版年：2006
- 出版时间：10 February 2006
- 年：2006
- 卷：340
- 期：2
- 页码：668-674
- 全文大小：247 K
文摘
Presenilin-1 (PS1) is a multipass transmembrane domain protein, which is believed to be the catalytic component of the γ-secretase complex. The complex is comprised of four major components: PS1, nicastrin, Aph-1, and Pen-2. The exact stoichiometric relationship between the four components remains unclear. It has been shown that γ-secretase exists as high molecular weight complexes, suggesting the possibility of dimer/multimer formation. We combined a biochemical approach with a novel morphological microscopy assay to analyze PS1 dimer formation and subcellular distribution in situ, in intact mammalian cells. Both coimmunoprecipitation and fluorescent lifetime imaging microscopy approaches showed that wildtype PS1 molecules form dimers. Moreover, PS1 holoproteins containing the D257A mutation also come into close enough proximity to form a dimer, suggesting that cleavage within the loop is not necessary for dimer formation. Taken together these data suggest that PS1 dimerization occurs during normal PS1 function.