NDLI: In vivo hyperspectral confocal fluorescence imaging to determine pigment localization and distribution in cyanobacterial cells

Content Provider	PubMed Central
Author	Vermaas, Wim F. J. Timlin, Jerilyn A. Jones, Howland D. T. Sinclair, Michael B. Nieman, Linda T. Hamad, Sawsan W. Melgaard, David K. Haaland, David M.
Abstract	Hyperspectral confocal fluorescence imaging provides the opportunity to obtain individual fluorescence emission spectra in small (≈0.03-μm3) volumes. Using multivariate curve resolution, individual fluorescence components can be resolved, and their intensities can be calculated. Here we localize, in vivo, photosynthesis-related pigments (chlorophylls, phycobilins, and carotenoids) in wild-type and mutant cells of the cyanobacterium Synechocystis sp. PCC 6803. Cells were excited at 488 nm, exciting primarily phycobilins and carotenoids. Fluorescence from phycocyanin, allophycocyanin, allophycocyanin-B/terminal emitter, and chlorophyll a was resolved. Moreover, resonance-enhanced Raman signals and very weak fluorescence from carotenoids were observed. Phycobilin emission was most intense along the periphery of the cell whereas chlorophyll fluorescence was distributed more evenly throughout the cell, suggesting that fluorescing phycobilisomes are more prevalent along the outer thylakoids. Carotenoids were prevalent in the cell wall and also were present in thylakoids. Two chlorophyll fluorescence components were resolved: the short-wavelength component originates primarily from photosystem II and is most intense near the periphery of the cell; and the long-wavelength component that is attributed to photosystem I because it disappears in mutants lacking this photosystem is of higher relative intensity toward the inner rings of the thylakoids. Together, the results suggest compositional heterogeneity between thylakoid rings, with the inner thylakoids enriched in photosystem I. In cells depleted in chlorophyll, the amount of both chlorophyll emission components was decreased, confirming the accuracy of the spectral assignments. These results show that hyperspectral fluorescence imaging can provide unique information regarding pigment organization and localization in the cell.
Related Links	http://dx.doi.org/10.1073/pnas.0708090105
Starting Page	4050
File Format	PDF
ISSN	10916490
e-ISSN	10916490
Journal	Proceedings of the National Academy of Sciences of the United States of America
Issue Number	10
Volume Number	105
Language	English
Publisher	National Academy of Sciences
Publisher Date	2008-03-11
Access Restriction	Open
Rights Holder	National Academy of Sciences
Subject Keyword	Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

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