The cw15 sta6 mutant strain of C. reinhardtii produces lipid bodies in both the cytoplasm and the chloroplast in response to nitrogen starvation in 20 mM acetate
Ursula Goodenough1, John Heuser2, Robyn Roth2, Luis Mario Aguirre Palma3, and Sabine Waffenschmidt3
1) Dept. Biology, Washington University, St. Louis MO 63130
2) Dept. Cell Biology, Washington University School of Medicine, St. Louis MO 63110
3) Dept. Biochemistry, University of Cologne, Cologne 4750674 Germany
The cw15 sta6 mutant strain, blocked in starch biosynthesis and hence affording clear views of the chloroplast stroma, was subjected to quick-freeze deep-etch electron microscopy ( ) following 30 hr of nitrogen starvation in 20 mM acetate. As expected, numerous lipid bodies (LB) were detected in the cytoplasm, many showing punctate connections with the endoplasmic reticulum (ER) where the outer leaflet of the ER membrane appears continuous with a monolayer surrounding the LB. Unexpectedly, numerous LBs of indistinguishable morphology were also detected in the chloroplast stroma. To confirm this identification, intact N-starved cw15 sta6 chloroplasts were isolated as described by Klein et al. (Plant Physiol. 72:481, 1983) and shown to contain abundant Nile-red-fluorescing bodes, a LB hallmark. Triacylglycerol, fatty-acid, and glycerollipid profiles of the cp-LBs will be presented and compared with the profiles of cyt-LBs that have been previ ously reported (Wang et al., Eukaryotic Cell 8: 1856-1868, 2009). We will also assess whether the cp-LB pathway is unique to the cw15 sta6 mutant or is also employed by the wild-type parental strain. Discovery of this second mode of LB formation in C. reinhardtii opens up an additional avenue for biochemical analysis and genetic manipulation of its TAG-producing potential.
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