Proteasome-dependent Degradation of Cyclin D1 in 1-Methyl-4-phenylpyridinium Ion (MPP+)-induced Cell Cycle Arrest

Abstract

1-Methyl-4-phenylpyridinium ion (MPP ؉), an active metabolite of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, induces cell death and inhibition of cell proliferation in various cells. However, the mechanism whereby MPP ؉ inhibits cell proliferation is still unclear. In this study, we found that MPP ؉ suppressed the proliferation with accumulation in G 1 phase without inducing cell death in p53-deficient MG63 osteosarcoma cells. MPP ؉ induced hypophosphorylation of retinoblastoma protein and rapidly down-regulated the protein but not mRNA levels of cyclin D1 in MG63 cells. The down-regulation of cyclin D1 protein was suppressed by a proteasome inhibitor, MG132. The cyclin D1 down-regulation by MPP ؉ was also observed in p53-positive PC12, HeLa S3, and HeLa 0 cells, which are a subclone of HeLa S3 lacking mitochondrial DNA. Moreover, MPP ؉ dephosphorylated Akt in PC12 cells, which was rescued by the pretreatment with nerve growth factor. In addition, the pretreatment with nerve growth factor or lithium chloride, a glycogen synthase kinase-3␤ inhibitor, suppressed the cyclin D1 down-regulation caused by MPP ؉ . Our results demonstrate that MPP ؉ induces cell cycle arrest independently of its mitochondrial toxicity or the p53 status of the target cells, but rather through the proteasome-and phosphatidylinositol 3-Akt-glycogen synthase kinase-3␤-dependent cyclin D1 degradation.1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) 1 is commonly used in animals to produce an experimental model of Parkinson's disease (1, 2). 1-Methyl-4-phenylpyridinium ion (MPP ϩ ) (3), an active metabolite of MPTP, induces cell death and inhibition of proliferation not only in neuronal cells but also in other types of cell (4 -7). The mechanisms whereby MPP ϩ induces the inhibition of membrane-bound NADH dehydrogenase in mitochondrial complex I (8 -10), ATP depletion (11), and production of the superoxide radicals (12, 13) have been studied extensively. However, the mechanisms involved in the inhibition of cell proliferation by MPP ϩ have rarely been examined.Progression and transition in the cell cycle of mammalian cells are governed by cyclin-dependent kinases (Cdks) whose activities are regulated by the binding of their regulatory subunits (called cyclins) or Cdk inhibitors (14). p21 effectively inhibits Cdk2, Cdk3, Cdk4, and Cdk6, which have a direct role in the G 1 /S transition, but it is a poor inhibitor of other known Cdks (15-17). Cyclin proteolysis is essential for cell cycle progression (18,19). Cyclin D1 regulates G 1 arrest and the phosphorylation status of retinoblastoma protein (Rb). Cyclin D1 degradation is mediated by phosphorylation-triggered, ubiquitin-dependent proteolysis (20). Cyclin D1 proteolysis is regulated by glycogen synthase kinase-3␤ (GSK-3␤) (21), which is inactivated by a pathway that sequentially involves Ras, phosphatidylinositol 3-kinase (PI3K), and Akt (22).The aim of this study was to examine the mechanisms whereby MPP ϩ inhibits cell proliferation. We show here that MPP ϩ initiates an efficient p53-...

Does Thioredoxin-1 Prevent Mitochondria- and Endoplasmic Reticulum-Mediated Neurotoxicity of 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine?

“…PC12 cells were maintained in RPMI1640 medium (Life Technologies) supplemented with 10% heat-inactivated fetal horse serum (HS) and 5% FCS. HeLa-0 cells were described previously (4). PCR confirmed that these cells are subclones of HeLa-S3 cells that have been cultured with ethidium bromide and lack mitochondrial DNA.…”

Commitment of 1-Methyl-4-phenylpyrinidinium Ion-induced Neuronal Cell Death by Proteasome-mediated Degradation of p35 Cyclin-dependent Kinase 5 Activator

“…In contrast, several studies have reported increased proteasome activity as observed here. MPP ϩ induces the rapid proteasomal degradation of short-lived proteins such as Ret protein in PC12 cells (42) and cyclin D1 in MG63 osteosarcoma cells and HeLa cells (43). Proteasome activity, when measured with a peptide substrates, is elevated by exposure to 10 or 100 M MPP ϩ for 2 h (44).…”

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“…The viable cells were also counted using NucleoCounter, as described previously. (17) Flow cytometry. Cell-surface expression of TRAIL-R2 was determined by flow cytometry with an EPICS XL (Beckman Coulter, Miami, FL, USA).…”