Probing the in situ volumes of Arabidopsis leaf plastids using three‐dimensional confocal and scanning electron microscopy
- School of Biological Sciences Washington State University P.O. Box 644236 Pullman Washington 99164‐4236 USA
- Institute of Plant Biology and Biotechnology University of Münster Schlossplatz 7 48149 Münster Germany
- School of Biological Sciences Washington State University P.O. Box 644236 Pullman Washington 99164‐4236 USA, LMU Munich, Plant Biochemistry Großhadernerstr. 2‐4 82152 Planegg‐Martinsried Germany
Leaf plastids harbor a plethora of biochemical reactions including photosynthesis, one of the most important metabolic pathways on Earth. Scientists are eager to unveil the physiological processes within the organelle but also their interconnection with the rest of the plant cell. An increasingly important feature of this venture is to use experimental data in the design of metabolic models. A remaining obstacle has been the limited in situ volume information of plastids and other cell organelles. To fill this gap for chloroplasts, we established three microscopy protocols delivering in situ volumes based on: (i) chlorophyll fluorescence emerging from the thylakoid membrane, (ii) a CFP marker embedded in the envelope, and (iii) calculations from serial block‐face scanning electron microscopy (SBFSEM). The obtained data were corroborated by comparing wild‐type data with two mutant lines affected in the plastid division machinery known to produce small and large mesophyll chloroplasts, respectively. Furthermore, we also determined the volume of the much smaller guard cell plastids. Interestingly, their volume is not governed by the same components of the division machinery which defines mesophyll plastid size. Based on our three approaches, the average volume of a mature Col‐0 wild‐type mesophyll chloroplasts is 93 μm 3 . Wild‐type guard cell plastids are approximately 18 μm 3 . Lastly, our comparative analysis shows that the chlorophyll fluorescence analysis can accurately determine chloroplast volumes, providing an important tool to research groups without access to transgenic marker lines expressing genetically encoded fluorescence proteins or costly SBFSEM equipment.
- Sponsoring Organization:
- USDOE
- Grant/Contract Number:
- SC0018277
- OSTI ID:
- 2217386
- Alternate ID(s):
- OSTI ID: 2283117
- Journal Information:
- The Plant Journal, Journal Name: The Plant Journal Journal Issue: 2 Vol. 117; ISSN 0960-7412
- Publisher:
- Wiley-BlackwellCopyright Statement
- Country of Publication:
- United Kingdom
- Language:
- English
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