Jeong Ae Park

Ginsenoside-Rh2 blocks the cell cycle of SK-HEP-1 cells at the G1/S boundary by selectively inducing the protein expression of p27kip1

The mechanism of action by which ginsenoside-Rh2 (G-Rh2) suppresses the proliferation of SK-HEP-1 cells is reported. The results from flow cytometric analyses show that G-Rh2 arrested the cell cycle at the G1/S transition phase. The cyclin E-dependent kinase activity which had been immunoprecipitated with cyclin E-specific antibody was down-regulated in the cells in response to G-Rh2. The IC50 value required to down-regulate the kinase activity by 50% was approximately 0.75 microM. Immunoblotting analyses show that G-Rh2 selectively induced the expression of p27kip1 in a dose-dependent manner whereas it had no effect on the levels of cyclin E, cdk2, and p21WAF1. In addition, our data show that G-Rh2 reduced the protein levels of cdc25A at doses higher than 10 microM. Collectively, these data suggest that ginsenoside-Rh2 arrests the cell cycle at the G1/S transition phase by selectively inducing protein expression of p27Kip1 and, as a consequence, down-regulating cyclin E-dependent kinase activity.

Negative regulation of hypoxia inducible factor-1α by necdin

Hypoxia‐inducible factor 1 (HIF‐1) is a master transcription factor that mediates cellular and systemic homeostatic responses to reduce O2 availability, such as erythropoiesis, angiogenesis, and glycolysis. Using the yeast two‐hybrid screening system, we found that the oxygen dependent degradation (ODD) domain of HIF‐1α interacts with necdin, a growth suppressor. The interaction of necdin with HIF‐1α was confirmed using coimmunoprecipitation with the overexpressed HIF‐1α. Biological effect of necdin on HIF‐1α showed that necdin reduces the transcriptional activity of HIF‐1 under hypoxia. Moreover, necdin decreased the level of the HIF‐1α protein, but not that of mRNA, implying a possibility of necdin‐mediated HIF‐1α degradation. Furthermore, necdin has an anti‐angiogenic activity in the tube formation assay and CAM assay, which might be due to the downregulation of HIF‐1α. Collectively, these results suggest that necdin can be a novel negative regulator of HIF‐1α stability via the direct interaction.

Arrest Defective 1 Autoacetylation Is a Critical Step in Its Ability to Stimulate Cancer Cell Proliferation

The N-acetyltransferase arrest defective 1 (ARD1) is an important regulator of cell growth and differentiation that has emerged recently as a critical molecule in cancer progression. However, the regulation of the enzymatic and biological activities of human ARD1 (hARD1) in cancer is presently poorly understood. Here, we report that hARD1 undergoes autoacetylation and that this modification is essential for its functional activation. Using liquid chromatography-tandem mass spectrometry and site-directed mutational analyses, we identified K136 residue as an autoacetylation target site. K136R mutation abolished the ability of hARD1 to promote cancer cell growth in vitro and tumor xenograft growth in vivo. Mechanistic investigations revealed that hARD1 autoacetylation stimulated cyclin D1 expression through activation of the transcription factors beta-catenin and activator protein-1. Our results show that hARD1 autoacetylation is critical for its activation and its ability to stimulate cancer cell proliferation and tumorigenesis.

Blood-brain barrier interfaces and brain tumors

In the developing brain, capillaries are differentiated and matured into the blood-brain barrier (BBB), which is composed of cerebral endothelial cells, astrocyte end-feet, and pericytes. Since the BBB regulates the homeostasis of central nervous system (CNS), the maintenance of the BBB is important for CNS function. The disruption of the BBB may result in many brain disorders including brain tumors. However, the molecular mechanism of BBB formation and maintenance is poorly understood. Here, we summarize recent advances in the role of oxygen tension and growth factors on BBB development and maintenance, and in BBB dysfunction related with brain tumors.

Receptor Activator of Nuclear Factor κB Ligand Is a Novel Inducer of Tissue Factor in Macrophages

Rationale: Although recent studies have suggested a role for the receptor activator of nuclear factor &kgr;B ligand (RANKL) in the late stages of atherosclerosis (eg, plaque destabilization and rupture), the underlying mechanisms and subsequent events are unclear. Objective: Because blood clotting is common after plaque rupture, we hypothesized that RANKL influenced tissue factor (TF) expression and activity to initiate the coagulation cascade. Methods and Results: RANKL increased the TF mRNA level and procoagulant activity in macrophages, as determined by semiquantitative reverse transcription polymerase chain reaction (semiquantitative RT-PCR) and a chromogenic assay. TF promoter analysis revealed that AP-1 and Egr-1 are responsible for RANKL-induced TF transcription. In addition, RANKL increased phosphorylation of c-Jun NH2-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK)1/2. RANKL-induced TF expression was attenuated by JNK- and MEK1-specific inhibitors and by small interfering RNA knockdown of c-Jun and Egr-1. Conclusion: Our results indicate that RANKL induces TF in macrophages mainly through the cooperative action of AP-1 and Egr-1 via JNK and ERK1/2 pathways. These findings provide strong mechanistic support for the role of RANKL in the thrombogenicity of atherosclerotic plaques.