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Originally published In Press as doi:10.1074/jbc.M009878200 on December 19, 2000

J. Biol. Chem., Vol. 276, Issue 11, 8492-8499, March 16, 2001
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Highly Saturated Endonuclear Phosphatidylcholine Is Synthesized in Situ and Colocated with CDP-choline Pathway Enzymes*

Alan N. HuntDagger §, Graeme T. ClarkDagger , George S. Attard, and Anthony D. PostleDagger

From the Dagger  Department of Child Health, University of Southampton, Southampton General Hospital, Southampton SO16 6YD, United Kingdom and the  Department of Chemistry, University of Southampton, Southampton SO17 1BJ, United Kingdom

Chromatin-associated phospholipids are well recognized. A report that catalytically active endonuclear CTP:choline-phosphate cytidylyltransferase alpha  is necessary for cell survival questions whether endonuclear, CDP-choline pathway phosphatidylcholine synthesis may occur in situ. We report that chromatin from human IMR-32 neuroblastoma cells possesses such a biosynthetic pathway. First, membrane-free nuclei retain all three CDP-choline pathway enzymes in proportions comparable with the content of chromatin-associated phosphatidylcholine. Second, following supplementation of cells with deuterated choline and using electrospray ionization mass spectrometry, both the time course and molecular species labeling pattern of newly synthesized endonuclear and whole cell phosphatidylcholine revealed the operation of spatially separate, compositionally distinct biosynthetic routes. Specifically, endogenous and newly synthesized endonuclear phosphatidylcholine species are both characterized by a high degree of diacyl/alkylacyl chain saturation. This unusual species content and synthetic pattern (evident within 10 min of supplementation) are maintained through cell growth arrest by serum depletion and when proliferation is restored, suggesting that endonuclear disaturated phosphatidylcholine enrichment is essential and closely regulated. We propose that endonuclear phosphatidylcholine synthesis may regulate periodic nuclear accumulations of phosphatidylcholine-derived lipid second messengers. Furthermore, our estimates of saturated phosphatidylcholine nuclear volume occupancy of around 10% may imply a significant additional role in regulating chromatin structure.

* This work was supported by The Wellcome Trust (Grants 055490 and 457405).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

§ To whom correspondence should be addressed: Dept. of Child Health (803), Allergy and Inflammation Sciences Division, University of Southampton, Level G, Centre Block, Southampton General Hospital, Tremona Rd., Southampton SO16 6YD, UK. Tel.: 44 23 8079 4178; Fax: 44 23 8079 6378; E-mail: anh@soton.ac.uk.

Copyright © 2001 by The American Society for Biochemistry and Molecular Biology, Inc.
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