Leucine-Rich Repeat Kinase 2 (LRRK2) phosphorylates p53 and induces p21WAF1/CIP1 expression

Dong Hwan Ho, Hyejung Kim, Jisun Kim, Hyuna Sim, Hyunjun Ahn, Janghwan Kim, Hyemyung Seo, Kwang Chul Chung, Bum Joon Park, Ilhong Son, Wongi Seol

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Abstract

Background: Leucine-rich repeat kinase 2 (LRRK2) is a gene in which a mutation causes Parkinson's disease (PD), and p53 is a prototype tumor suppressor. In addition, activation of p53 in patient with PD has been reported by several studies. Because phosphorylation of p53 is critical for regulating its activity and LRRK2 is a kinase, we tested whether p53 is phosphorylated by LRRK2. Results: LRRK2 phosphorylates threonine (Thr) at TXR sites in an in vitro kinase assay, and the T304 and T377 were identified as putative phosphorylated residues. An increase of phospho-Thr in the p53 TXR motif was confirmed in the cells overexpressing G2019S, and human induced pluripotent stem (iPS) cells of a G2019S carrier. Interactions between LRRK2 and p53 were confirmed by co-immunoprecipitation of lysates of differentiated SH-SY5Y cells. LRRK2 mediated p53 phosphorylation translocalizes p53 predominantly to nucleus and increases p21WAF1/CIP1 expression in SH-SY5Y cells based on reverse transcription-polymerase chain reaction and Western blot assay results. The luciferase assay using the p21WAF1/CIP1 promoter-reporter also confirmed that LRRK2 kinase activity increases p21 expression. Exogenous expression of G2019S and the phosphomimetic p53 T304/377D mutants increased expression of p21WAF1/CIP1 and cleaved PARP, and cytotoxicity in the same cells. We also observed increase of p21 expression in rat primary neuron cells after transient expression of p53 T304/377D mutants and the mid-brain lysates of the G2019S transgenic mice. Conclusion: p53 is a LRRK2 kinase substrate. Phosphorylation of p53 by LRRK2 induces p21WAF1/CIP1 expression and apoptosis in differentiated SH-SY5Y cells and rat primary neurons.

Original languageEnglish
Article number145
JournalMolecular Brain
Volume8
Issue number1
DOIs
Publication statusPublished - 2015 Sep 18

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Leucine
Phosphotransferases
Phosphorylation
Threonine
Parkinson Disease
Neurons
Induced Pluripotent Stem Cells
Luciferases
Immunoprecipitation
Transgenic Mice
Reverse Transcription
Western Blotting
Apoptosis
Polymerase Chain Reaction

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Cellular and Molecular Neuroscience

Cite this

Ho, Dong Hwan ; Kim, Hyejung ; Kim, Jisun ; Sim, Hyuna ; Ahn, Hyunjun ; Kim, Janghwan ; Seo, Hyemyung ; Chung, Kwang Chul ; Park, Bum Joon ; Son, Ilhong ; Seol, Wongi. / Leucine-Rich Repeat Kinase 2 (LRRK2) phosphorylates p53 and induces p21WAF1/CIP1 expression. In: Molecular Brain. 2015 ; Vol. 8, No. 1.
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title = "Leucine-Rich Repeat Kinase 2 (LRRK2) phosphorylates p53 and induces p21WAF1/CIP1 expression",
abstract = "Background: Leucine-rich repeat kinase 2 (LRRK2) is a gene in which a mutation causes Parkinson's disease (PD), and p53 is a prototype tumor suppressor. In addition, activation of p53 in patient with PD has been reported by several studies. Because phosphorylation of p53 is critical for regulating its activity and LRRK2 is a kinase, we tested whether p53 is phosphorylated by LRRK2. Results: LRRK2 phosphorylates threonine (Thr) at TXR sites in an in vitro kinase assay, and the T304 and T377 were identified as putative phosphorylated residues. An increase of phospho-Thr in the p53 TXR motif was confirmed in the cells overexpressing G2019S, and human induced pluripotent stem (iPS) cells of a G2019S carrier. Interactions between LRRK2 and p53 were confirmed by co-immunoprecipitation of lysates of differentiated SH-SY5Y cells. LRRK2 mediated p53 phosphorylation translocalizes p53 predominantly to nucleus and increases p21WAF1/CIP1 expression in SH-SY5Y cells based on reverse transcription-polymerase chain reaction and Western blot assay results. The luciferase assay using the p21WAF1/CIP1 promoter-reporter also confirmed that LRRK2 kinase activity increases p21 expression. Exogenous expression of G2019S and the phosphomimetic p53 T304/377D mutants increased expression of p21WAF1/CIP1 and cleaved PARP, and cytotoxicity in the same cells. We also observed increase of p21 expression in rat primary neuron cells after transient expression of p53 T304/377D mutants and the mid-brain lysates of the G2019S transgenic mice. Conclusion: p53 is a LRRK2 kinase substrate. Phosphorylation of p53 by LRRK2 induces p21WAF1/CIP1 expression and apoptosis in differentiated SH-SY5Y cells and rat primary neurons.",
author = "Ho, {Dong Hwan} and Hyejung Kim and Jisun Kim and Hyuna Sim and Hyunjun Ahn and Janghwan Kim and Hyemyung Seo and Chung, {Kwang Chul} and Park, {Bum Joon} and Ilhong Son and Wongi Seol",
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Ho, DH, Kim, H, Kim, J, Sim, H, Ahn, H, Kim, J, Seo, H, Chung, KC, Park, BJ, Son, I & Seol, W 2015, 'Leucine-Rich Repeat Kinase 2 (LRRK2) phosphorylates p53 and induces p21WAF1/CIP1 expression', Molecular Brain, vol. 8, no. 1, 145. https://doi.org/10.1186/s13041-015-0145-7

Leucine-Rich Repeat Kinase 2 (LRRK2) phosphorylates p53 and induces p21WAF1/CIP1 expression. / Ho, Dong Hwan; Kim, Hyejung; Kim, Jisun; Sim, Hyuna; Ahn, Hyunjun; Kim, Janghwan; Seo, Hyemyung; Chung, Kwang Chul; Park, Bum Joon; Son, Ilhong; Seol, Wongi.

In: Molecular Brain, Vol. 8, No. 1, 145, 18.09.2015.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Leucine-Rich Repeat Kinase 2 (LRRK2) phosphorylates p53 and induces p21WAF1/CIP1 expression

AU - Ho, Dong Hwan

AU - Kim, Hyejung

AU - Kim, Jisun

AU - Sim, Hyuna

AU - Ahn, Hyunjun

AU - Kim, Janghwan

AU - Seo, Hyemyung

AU - Chung, Kwang Chul

AU - Park, Bum Joon

AU - Son, Ilhong

AU - Seol, Wongi

PY - 2015/9/18

Y1 - 2015/9/18

N2 - Background: Leucine-rich repeat kinase 2 (LRRK2) is a gene in which a mutation causes Parkinson's disease (PD), and p53 is a prototype tumor suppressor. In addition, activation of p53 in patient with PD has been reported by several studies. Because phosphorylation of p53 is critical for regulating its activity and LRRK2 is a kinase, we tested whether p53 is phosphorylated by LRRK2. Results: LRRK2 phosphorylates threonine (Thr) at TXR sites in an in vitro kinase assay, and the T304 and T377 were identified as putative phosphorylated residues. An increase of phospho-Thr in the p53 TXR motif was confirmed in the cells overexpressing G2019S, and human induced pluripotent stem (iPS) cells of a G2019S carrier. Interactions between LRRK2 and p53 were confirmed by co-immunoprecipitation of lysates of differentiated SH-SY5Y cells. LRRK2 mediated p53 phosphorylation translocalizes p53 predominantly to nucleus and increases p21WAF1/CIP1 expression in SH-SY5Y cells based on reverse transcription-polymerase chain reaction and Western blot assay results. The luciferase assay using the p21WAF1/CIP1 promoter-reporter also confirmed that LRRK2 kinase activity increases p21 expression. Exogenous expression of G2019S and the phosphomimetic p53 T304/377D mutants increased expression of p21WAF1/CIP1 and cleaved PARP, and cytotoxicity in the same cells. We also observed increase of p21 expression in rat primary neuron cells after transient expression of p53 T304/377D mutants and the mid-brain lysates of the G2019S transgenic mice. Conclusion: p53 is a LRRK2 kinase substrate. Phosphorylation of p53 by LRRK2 induces p21WAF1/CIP1 expression and apoptosis in differentiated SH-SY5Y cells and rat primary neurons.

AB - Background: Leucine-rich repeat kinase 2 (LRRK2) is a gene in which a mutation causes Parkinson's disease (PD), and p53 is a prototype tumor suppressor. In addition, activation of p53 in patient with PD has been reported by several studies. Because phosphorylation of p53 is critical for regulating its activity and LRRK2 is a kinase, we tested whether p53 is phosphorylated by LRRK2. Results: LRRK2 phosphorylates threonine (Thr) at TXR sites in an in vitro kinase assay, and the T304 and T377 were identified as putative phosphorylated residues. An increase of phospho-Thr in the p53 TXR motif was confirmed in the cells overexpressing G2019S, and human induced pluripotent stem (iPS) cells of a G2019S carrier. Interactions between LRRK2 and p53 were confirmed by co-immunoprecipitation of lysates of differentiated SH-SY5Y cells. LRRK2 mediated p53 phosphorylation translocalizes p53 predominantly to nucleus and increases p21WAF1/CIP1 expression in SH-SY5Y cells based on reverse transcription-polymerase chain reaction and Western blot assay results. The luciferase assay using the p21WAF1/CIP1 promoter-reporter also confirmed that LRRK2 kinase activity increases p21 expression. Exogenous expression of G2019S and the phosphomimetic p53 T304/377D mutants increased expression of p21WAF1/CIP1 and cleaved PARP, and cytotoxicity in the same cells. We also observed increase of p21 expression in rat primary neuron cells after transient expression of p53 T304/377D mutants and the mid-brain lysates of the G2019S transgenic mice. Conclusion: p53 is a LRRK2 kinase substrate. Phosphorylation of p53 by LRRK2 induces p21WAF1/CIP1 expression and apoptosis in differentiated SH-SY5Y cells and rat primary neurons.

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