Sphingosine Inhibits the Synthesis of RNA Primers by Primase in Vitro

Cynthia Marie G. Simbulan, Keiko Tamiya-Koizumi, Motoshi Suzuki, Mami Shoji, Shonen Yoshida, Takao Taki

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

We have previously shown the presence of sphingomyelin and sphingomyelinase in cell nuclei, suggesting that they may play a role in the intranuclear production of sphingosine, a potent bioactive molecule modulating diverse cellular functions. In the present study, the direct effects of sphingosine (Ci8:1) on the activity of DNA replication/repair polymerases were studied in vitro. Sphingosine had no effect on DNA polymerases a and β and slightly inhibited DNA polymerases γ, δ, and ϵ. In contrast, sphingosine strongly inhibited the activity of primase in a dose-dependent manner. On the other hand, dihydrosphingosine (Cisio), glycolipids, sphingomyelin, and ceramide had no effect on primase activity. Sphingosine equally inhibited the activity of primase complexed with DNA polymerase a, as well as its free form, with a Ki value of 4 µM. A gel-retardation analysis showed that the binding of primase with 32P-labeled template DNA was suppressed by sphingosine. Inhibition by sphingosine was competitive with the DNA template, but not with the substrate NTPs. After product analysis, a dose-dependent decrease in the amount of RNA primer products, consisting mainly of 10- and 11-mers, was observed in the presence of sphingosine, indicating that it inhibits the synthesis of RNA primers by primase. Sphingosine, however, had no effect on T7 RNA polymerase.

Original languageEnglish
Pages (from-to)9007-9012
Number of pages6
JournalBiochemistry
Volume33
Issue number30
DOIs
Publication statusPublished - 01-08-1994
Externally publishedYes

Fingerprint

DNA Primase
Sphingosine
DNA-Directed DNA Polymerase
Sphingomyelins
DNA
Sphingomyelin Phosphodiesterase
RNA primers
In Vitro Techniques
Ceramides
Glycolipids
Cell Nucleus
DNA Replication
DNA Repair
Repair
Gels
Cells

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

Simbulan, C. M. G., Tamiya-Koizumi, K., Suzuki, M., Shoji, M., Yoshida, S., & Taki, T. (1994). Sphingosine Inhibits the Synthesis of RNA Primers by Primase in Vitro. Biochemistry, 33(30), 9007-9012. https://doi.org/10.1021/bi00196a019
Simbulan, Cynthia Marie G. ; Tamiya-Koizumi, Keiko ; Suzuki, Motoshi ; Shoji, Mami ; Yoshida, Shonen ; Taki, Takao. / Sphingosine Inhibits the Synthesis of RNA Primers by Primase in Vitro. In: Biochemistry. 1994 ; Vol. 33, No. 30. pp. 9007-9012.
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Simbulan, CMG, Tamiya-Koizumi, K, Suzuki, M, Shoji, M, Yoshida, S & Taki, T 1994, 'Sphingosine Inhibits the Synthesis of RNA Primers by Primase in Vitro', Biochemistry, vol. 33, no. 30, pp. 9007-9012. https://doi.org/10.1021/bi00196a019

Sphingosine Inhibits the Synthesis of RNA Primers by Primase in Vitro. / Simbulan, Cynthia Marie G.; Tamiya-Koizumi, Keiko; Suzuki, Motoshi; Shoji, Mami; Yoshida, Shonen; Taki, Takao.

In: Biochemistry, Vol. 33, No. 30, 01.08.1994, p. 9007-9012.

Research output: Contribution to journalArticle

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Simbulan CMG, Tamiya-Koizumi K, Suzuki M, Shoji M, Yoshida S, Taki T. Sphingosine Inhibits the Synthesis of RNA Primers by Primase in Vitro. Biochemistry. 1994 Aug 1;33(30):9007-9012. https://doi.org/10.1021/bi00196a019