Induction of G1 Arrest and Apoptosis in Human Jurkat T Cells by Pentagalloylglucose through Inhibiting Proteasome Activity and Elevating p27Kip1, p21Cip1/WAF1, and Bax Proteins

Abstract

Pentagalloylglucose, which is found in many medicinal plants, can arrest the cell cycle at G 1 phase through down-regulation of cyclin-dependent kinases 2 and 4 and up-regulation of the cyclin-dependent kinase inhibitors p27 Kip1 and p21 Cip1/WAF1 in human breast cancer cells. Pentagalloylglucose also induces apoptosis in human leukemic cells. However, the mechanisms by which pentagalloylglucose induces these effects is unclear. We now show that pentagalloylglucose inhibits the activities of purified 20 and 26 S proteasomes in vitro, the 26 S proteasome in Jurkat T cell lysates, and chymotrypsinlike activity of the 26 S proteasome in intact Jurkat T cells. The turnover of p27 Kip1 and p21 Cip1/WAF1 , which is necessary for cell cycle progression mediated by proteasome degradation, was disrupted by treatment of human Jurkat T cells with pentagalloylglucose. This was shown by cycloheximide treatment and in vivo pulsechase labeling experiments, and this effect correlated with the arrest of proliferation of Jurkat T cells at G 1 . Inhibition of the proteasome by pentagalloylglucose and by the proteasome inhibitor MG132 caused accumulation of ubiquitin-tagged proteins in Jurkat T cells. The addition of pentagalloylglucose to Jurkat T cells enhanced the stability of the proteasome substrate Bax and increased cytochrome c release and apoptosis. Our findings suggest a mechanism for the effect of pentagalloylglucose on the cell cycle in human leukemic cells: that pentagalloylglucose down-regulates proteasomemediated pathways because it is a proteasome inhibitor.In eukaryotic cells, there are two distinct proteolytic mechanisms essential for regulating levels of cellular proteins. One is lysosomal mediated degradation, which is involved in the breakdown of intracellular proteins under conditions of cellular stress, membrane-associated proteins, and proteins that have entered the cell by endocytosis. The ubiquitin-proteasome pathway is the other mechanism responsible for the turnover of intracellular proteins. This second system works by marking specific substrates with ubiquitins and degrading the marked proteins by the 26 S proteasome in an ATP-dependent manner.

Gentamicin Causes Apoptosis at Low Concentrations in Renal LLC-PK 1 Cells Subjected to Electroporation

“…Bax is degraded by the proteasome (8,35) but, like other proteins, needs to be ubiquitinated before it can be processed (specific signaling [21]). Inhibition of the proteasome by specific inhibitors is known to cause an increase in ubiquitinated Bax, and this was shown to slow the rate at which Bax itself can be ubiquitinated (9,30,35,50). Our observations are consistent with such a process but do not identify the nature of the proteasome inhibitor.…”

Phosphorylation of the Cyclin-Dependent Kinase Inhibitor p21Cip1 on Serine 130 Is Essential for Viral Cyclin-Mediated Bypass of a p21Cip1-Imposed G1 Arrest

“…Cip1 has been intimately linked to turnover of this cdk inhibitor (for a review, see reference 5), although recent studies have questioned this view (8). Whatever the mechanism of p21 Cip1 proteolysis, our data indicate that protein degradation is not a major cause for K cyclin-mediated release of a p21…”

The Proteasome Inhibitor Bortezomib Acts Independently of p53 and Induces Cell Death via Apoptosis and Mitotic Catastrophe in B-Cell Lymphoma Cell Lines

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