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Plant and Cell Physiology Advance Access published online on March 3, 2005
Plant and Cell Physiology, doi:10.1093/pcp/pci070
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Plant and Cell Physiology 2005 © The Japanese Society of Plant Physiologists (JSPP); all rights reserved.
Received December 14, 2004
Accepted February 8, 2005

Regular Paper

Visualization of Plastid Nucleoids in Situ Using the PEND-GFP Fusion Protein

Kimihiro Terasawa 1 and Naoki Sato 2*

1 Department of Molecular Biology, Faculty of Science, Saitama University, Sakura-ku, Saitama, 338-8570 Japan
2 Department of Molecular Biology, Faculty of Science, Saitama University, Sakura-ku, Saitama, 338-8570 Japan; Department of Life Sciences, Graduate School of Arts and Sciences, University of Tokyo, 3-8-1 Komaba, Meguro-ku, Tokyo 153-8902, Japan

* To whom correspondence should be addressed.
Naoki Sato, E-mail: naokisat{at}bio.c.u-tokyo.ac.jp


  Abstract

Plastid DNA is a circular molecule of 120-150 kbp, which is organized into a protein-DNA complex called nucleoid. Although various plastids other than chloroplasts exist, such as etioplasts, amyloplasts and chromoplasts, it is not easy to observe plastid nucleoids within the cells of many non-green tissues. The PEND (plastid envelope DNA-binding) protein is a DNA-binding protein in the inner envelope membrane of developing chloroplast, and a DNA-binding domain called cbZIP is present at its N-terminus. We made various PEND-GFP fusion proteins using the cbZIP domains from various plants, and found that they were localized in the chloroplast nucleoids in transient expression in leaf protoplasts. In stable transformants of Arabidopsis thaliana, PEND-GFP fusion proteins were also localized in the nucleoids of various plastids. We have succeeded in visualizing plastid nucleoids in various intact tissues using this stable transformant. This technique is useful in root, flower and pollen, in which it had been difficult to observe plastid nucleoids. The relative arrangement of nucleoids within a chloroplast was kept unchanged when the chloroplast moved within a cell. During the division of plastid, nucleoids formed a network structure, which made possible equal partition of nucleoids.

Keywords: DNA binding protein; Green fluorescent protein; PEND protein; plastid division; plastid nucleoid; real-time imaging.
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