Abstract
Phosphatidylserine (PtdSer) synthesis in Chinese hamster ovary (CHO) cells occurs through the exchange of L-serine with the base moiety of phosphatidylcholine or phosphatidylethanolamine. The synthesis is depressed on the addition of PtdSer to the culture medium. A CHO cell mutant named mutant 29, whose PtdSer biosynthesis is highly resistant to this depression by exogenous PtdSer, has been isolated from CHO-K1 cells. In the present study, the PtdSer-resistant PtdSer biosynthesis in the mutant was traced to a point mutation in the PtdSer synthase I gene, pssA, resulting in the replacement of Arg-95 of the synthase by lysine. Introduction of the mutant pssA cDNA, but not the wild-type pssA cDNA, into CHO-K1 cells induced the PtdSer-resistant PtdSer biosynthesis. In a cell-free system, the serine base- exchange activity of the wild-type pssA-transfected cells was inhibited by PtdSer, but that of the mutant pssA-transfected cells was resistant to the inhibition. Like the mutant 29 cells, the mutant pssA-transfected cells grown without exogenous PtdSer exhibited an ≃2-fold increase in the cellular PtdSer level compared with that in CHO-K1 cells, although the wild-type pssA- transfected cells did not exhibit such a significant increase. These results indicated that the inhibition of PtdSer synthase I by PtdSer is essential for the maintenance of a normal PtdSer level in CHO-K1 cells and that Arg-95 of the synthase is a crucial residue for the inhibition.
| Original language | English |
|---|---|
| Pages (from-to) | 4199-4203 |
| Number of pages | 5 |
| Journal | Proceedings of the National Academy of Sciences of the United States of America |
| Volume | 95 |
| Issue number | 8 |
| DOIs | |
| Publication status | Published - Apr 14 1998 |
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All Science Journal Classification (ASJC) codes
- Genetics
- General
Cite this
Control of phosphatidylserine biosynthesis through phosphatidylserine- mediated inhibition of phosphatidylserine synthase I in Chinese hamster ovary cells. / Kuge, Osamu; Hasegawa, K.; Saito, K.; Nishijima, M.
In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 95, No. 8, 14.04.1998, p. 4199-4203.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Control of phosphatidylserine biosynthesis through phosphatidylserine- mediated inhibition of phosphatidylserine synthase I in Chinese hamster ovary cells
AU - Kuge, Osamu
AU - Hasegawa, K.
AU - Saito, K.
AU - Nishijima, M.
PY - 1998/4/14
Y1 - 1998/4/14
N2 - Phosphatidylserine (PtdSer) synthesis in Chinese hamster ovary (CHO) cells occurs through the exchange of L-serine with the base moiety of phosphatidylcholine or phosphatidylethanolamine. The synthesis is depressed on the addition of PtdSer to the culture medium. A CHO cell mutant named mutant 29, whose PtdSer biosynthesis is highly resistant to this depression by exogenous PtdSer, has been isolated from CHO-K1 cells. In the present study, the PtdSer-resistant PtdSer biosynthesis in the mutant was traced to a point mutation in the PtdSer synthase I gene, pssA, resulting in the replacement of Arg-95 of the synthase by lysine. Introduction of the mutant pssA cDNA, but not the wild-type pssA cDNA, into CHO-K1 cells induced the PtdSer-resistant PtdSer biosynthesis. In a cell-free system, the serine base- exchange activity of the wild-type pssA-transfected cells was inhibited by PtdSer, but that of the mutant pssA-transfected cells was resistant to the inhibition. Like the mutant 29 cells, the mutant pssA-transfected cells grown without exogenous PtdSer exhibited an ≃2-fold increase in the cellular PtdSer level compared with that in CHO-K1 cells, although the wild-type pssA- transfected cells did not exhibit such a significant increase. These results indicated that the inhibition of PtdSer synthase I by PtdSer is essential for the maintenance of a normal PtdSer level in CHO-K1 cells and that Arg-95 of the synthase is a crucial residue for the inhibition.
AB - Phosphatidylserine (PtdSer) synthesis in Chinese hamster ovary (CHO) cells occurs through the exchange of L-serine with the base moiety of phosphatidylcholine or phosphatidylethanolamine. The synthesis is depressed on the addition of PtdSer to the culture medium. A CHO cell mutant named mutant 29, whose PtdSer biosynthesis is highly resistant to this depression by exogenous PtdSer, has been isolated from CHO-K1 cells. In the present study, the PtdSer-resistant PtdSer biosynthesis in the mutant was traced to a point mutation in the PtdSer synthase I gene, pssA, resulting in the replacement of Arg-95 of the synthase by lysine. Introduction of the mutant pssA cDNA, but not the wild-type pssA cDNA, into CHO-K1 cells induced the PtdSer-resistant PtdSer biosynthesis. In a cell-free system, the serine base- exchange activity of the wild-type pssA-transfected cells was inhibited by PtdSer, but that of the mutant pssA-transfected cells was resistant to the inhibition. Like the mutant 29 cells, the mutant pssA-transfected cells grown without exogenous PtdSer exhibited an ≃2-fold increase in the cellular PtdSer level compared with that in CHO-K1 cells, although the wild-type pssA- transfected cells did not exhibit such a significant increase. These results indicated that the inhibition of PtdSer synthase I by PtdSer is essential for the maintenance of a normal PtdSer level in CHO-K1 cells and that Arg-95 of the synthase is a crucial residue for the inhibition.
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U2 - 10.1073/pnas.95.8.4199
DO - 10.1073/pnas.95.8.4199
M3 - Article
C2 - 9539713
AN - SCOPUS:0032516053
VL - 95
SP - 4199
EP - 4203
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
SN - 0027-8424
IS - 8
ER -