PLD1 Regulates mTOR Signaling and Mediates Cdc42 Activation of S6K1

Yimin Fang, In Hyun Park, Ai Luen Wu, Guangwei Du, Ping Huang, Michael A. Frohman, Stephanie J. Walker, H. Alex Brown, Jie Chen

Research output: Contribution to journalArticle

Abstract

Background: The mammalian target of rapamycin (mTOR) regulates cell growth and proliferation via the downstream targets ribosomal S6 kinase 1 (S6K1) and eukaryotic translation initiation factor 4E binding protein 1 (4E-BP1). We have identified phosphatidic acid (PA) as a mediator of mitogenic activation of mTOR signaling. In this study, we set out to test the hypotheses that phospholipase D 1 (PLD1) is an upstream regulator of mTOR and that the previously reported S6K1 activation by Cdc42 is mediated by PLD1. Results: Overexpression of wild-type PLD1 increased S6K1 activity in serum-stimulated cells, whereas a catalytically inactive PLD1 exerted a dominant-negative effect on S6K1. More importantly, eliminating endogenous PLD1 by RNAi led to drastic inhibition of serum-stimulated S6K1 activation and 4E-BP1 hyperphosphorylation in both HEK293 and COS-7 cells. Knockdown of PLD1 also resulted in reduced cell size, suggesting a critical role for PLD1 in cell growth control. Using a rapamycin-resistant S6K1 mutant, Cdc42's action was demonstrated to be through the mTOR pathway. When Cdc42 was mutated in a region specifically required for PLD1 activation, its ability to activate S6K1 in the presence of serum was hindered. However, when exogenous PA was used as a stimulus, the PLD1-inactive Cdc42 mutant behaved similarly to the wild-type protein. Conclusions: Our observations reveal the involvement of PLD1 in mTOR signaling and cell size control, and provide a molecular mechanism for Cdc42 activation of S6K1. A new cascade is proposed to connect mitogenic signals to mTOR through Cdc42, PLD1, and PA.

Original languageEnglish (US)
Pages (from-to)2037-2044
Number of pages8
JournalCurrent Biology
Volume13
Issue number23
DOIs
StatePublished - Dec 2 2003

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Ribosomal Protein S6 Kinases
Phospholipase D
phospholipase D
Sirolimus
phosphotransferases (kinases)
Chemical activation
Phosphatidic Acids
Cell growth
Cell Size
binding proteins
cell growth
acids
Carrier Proteins
Serum
Eukaryotic Initiation Factor-4E
cells
Eukaryotic Initiation Factors
mutants
COS Cells
Cell proliferation

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)

Cite this

Fang, Y., Park, I. H., Wu, A. L., Du, G., Huang, P., Frohman, M. A., ... Chen, J. (2003). PLD1 Regulates mTOR Signaling and Mediates Cdc42 Activation of S6K1. Current Biology, 13(23), 2037-2044. https://doi.org/10.1016/j.cub.2003.11.021

PLD1 Regulates mTOR Signaling and Mediates Cdc42 Activation of S6K1. / Fang, Yimin; Park, In Hyun; Wu, Ai Luen; Du, Guangwei; Huang, Ping; Frohman, Michael A.; Walker, Stephanie J.; Brown, H. Alex; Chen, Jie.

In: Current Biology, Vol. 13, No. 23, 02.12.2003, p. 2037-2044.

Research output: Contribution to journalArticle

Fang, Y, Park, IH, Wu, AL, Du, G, Huang, P, Frohman, MA, Walker, SJ, Brown, HA & Chen, J 2003, 'PLD1 Regulates mTOR Signaling and Mediates Cdc42 Activation of S6K1', Current Biology, vol. 13, no. 23, pp. 2037-2044. https://doi.org/10.1016/j.cub.2003.11.021
Fang Y, Park IH, Wu AL, Du G, Huang P, Frohman MA et al. PLD1 Regulates mTOR Signaling and Mediates Cdc42 Activation of S6K1. Current Biology. 2003 Dec 2;13(23):2037-2044. https://doi.org/10.1016/j.cub.2003.11.021
Fang, Yimin ; Park, In Hyun ; Wu, Ai Luen ; Du, Guangwei ; Huang, Ping ; Frohman, Michael A. ; Walker, Stephanie J. ; Brown, H. Alex ; Chen, Jie. / PLD1 Regulates mTOR Signaling and Mediates Cdc42 Activation of S6K1. In: Current Biology. 2003 ; Vol. 13, No. 23. pp. 2037-2044.
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AU - Frohman, Michael A.

AU - Walker, Stephanie J.

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AB - Background: The mammalian target of rapamycin (mTOR) regulates cell growth and proliferation via the downstream targets ribosomal S6 kinase 1 (S6K1) and eukaryotic translation initiation factor 4E binding protein 1 (4E-BP1). We have identified phosphatidic acid (PA) as a mediator of mitogenic activation of mTOR signaling. In this study, we set out to test the hypotheses that phospholipase D 1 (PLD1) is an upstream regulator of mTOR and that the previously reported S6K1 activation by Cdc42 is mediated by PLD1. Results: Overexpression of wild-type PLD1 increased S6K1 activity in serum-stimulated cells, whereas a catalytically inactive PLD1 exerted a dominant-negative effect on S6K1. More importantly, eliminating endogenous PLD1 by RNAi led to drastic inhibition of serum-stimulated S6K1 activation and 4E-BP1 hyperphosphorylation in both HEK293 and COS-7 cells. Knockdown of PLD1 also resulted in reduced cell size, suggesting a critical role for PLD1 in cell growth control. Using a rapamycin-resistant S6K1 mutant, Cdc42's action was demonstrated to be through the mTOR pathway. When Cdc42 was mutated in a region specifically required for PLD1 activation, its ability to activate S6K1 in the presence of serum was hindered. However, when exogenous PA was used as a stimulus, the PLD1-inactive Cdc42 mutant behaved similarly to the wild-type protein. Conclusions: Our observations reveal the involvement of PLD1 in mTOR signaling and cell size control, and provide a molecular mechanism for Cdc42 activation of S6K1. A new cascade is proposed to connect mitogenic signals to mTOR through Cdc42, PLD1, and PA.

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