Spectro-Microscopy of Living Plant Cells  

作　　者：Klaus Hatter Alfred J. Meixner Frank Schleifenbaum 

机构地区：[1]Center for Plant Molecular Biology （ZMBP）, Plant Physiology and Biophysical Chemistry, University of Tubingen, Auf der Morgenstelle 1, 72076 Tuebingen, Germany [2]Institute for Physical and Theoretical Chemistry （IPTC）, Nano-Optics, University of Tubingen, Auf der Morgenstelle 8, 72076 Tubingen, Germany

出　　处：《Molecular Plant》2012年第1期14-26,共13页分子植物（英文版）

摘　　要：Spectro-microscopy, a combination of fluorescence microscopy with spatially resolved spectroscopic techni- ques, provides new and exciting tools for functional cell biology in living organisms. This review focuses on recent devel- opments in spectro-microscopic applications for the investigation of living plant cells in their native tissue context. The application of spectro-microscopic methods led to the recent discovery of a fast signal response pathway for the brassi- nosteroide receptor BRI1 in the plasma membrane of living plant cells. Moreover, the competence of different plant cell types to respond to environmental or endogenous stimuli was determined in vivo by correlation analysis of different optical and spectroscopic readouts such as fluorescence lifetime （FLT）. Furthermore, a new spectro-microscopic technique, fluorescence intensity decay shape analysis microscopy （FIDSAM）, has been developed. FIDSAM is capable of imaging low- expressed fluorophore-tagged proteins at high spatial resolution and precludes the misinterpretation of autofluorescence artifacts. In addition, FIDSAM provides a very effective and sensitive tool on the basis of F6rster resonance energy transfer （FRET） for the qualitative and quantitative determination of protein-protein interaction. Finally, we report on the quan- titative analysis of the photosystem I and II （PSI/PSII） ratio in the chloroplasts of living Arabidopsis plants at room tem- perature, using high-resolution, spatially resolved fluorescence spectroscopy. With this technique, it was not only possible to measure PSI/PSII ratios, but also to demonstrate the differential competence of wild-type and carbohydrate-deficient plants to adapt the PSI/PSII ratio to different light conditions. In summary, the information content of standard microscopic images is extended by several dimensions by the use of spectro-microscopic approaches. Therefore, novel cell physiolog- ical and molecular topics can be addressed and valuable insights into molecular and subcellulaSpectro-microscopy, a combination of fluorescence microscopy with spatially resolved spectroscopic techni- ques, provides new and exciting tools for functional cell biology in living organisms. This review focuses on recent devel- opments in spectro-microscopic applications for the investigation of living plant cells in their native tissue context. The application of spectro-microscopic methods led to the recent discovery of a fast signal response pathway for the brassi- nosteroide receptor BRI1 in the plasma membrane of living plant cells. Moreover, the competence of different plant cell types to respond to environmental or endogenous stimuli was determined in vivo by correlation analysis of different optical and spectroscopic readouts such as fluorescence lifetime （FLT）. Furthermore, a new spectro-microscopic technique, fluorescence intensity decay shape analysis microscopy （FIDSAM）, has been developed. FIDSAM is capable of imaging low- expressed fluorophore-tagged proteins at high spatial resolution and precludes the misinterpretation of autofluorescence artifacts. In addition, FIDSAM provides a very effective and sensitive tool on the basis of F6rster resonance energy transfer （FRET） for the qualitative and quantitative determination of protein-protein interaction. Finally, we report on the quan- titative analysis of the photosystem I and II （PSI/PSII） ratio in the chloroplasts of living Arabidopsis plants at room tem- perature, using high-resolution, spatially resolved fluorescence spectroscopy. With this technique, it was not only possible to measure PSI/PSII ratios, but also to demonstrate the differential competence of wild-type and carbohydrate-deficient plants to adapt the PSI/PSII ratio to different light conditions. In summary, the information content of standard microscopic images is extended by several dimensions by the use of spectro-microscopic approaches. Therefore, novel cell physiolog- ical and molecular topics can be addressed and valuable insights into molecular and subcellula

关 键 词：FLUORESCENCE MICROSCOPY FLIM FRET PHOTOSYSTEMS protein-protein interaction FIDSAM. 

分 类 号：Q942[生物学—植物学] TH742.65[机械工程—光学工程]