14-3-3epsilon protein increases matrix metalloproteinase-2 gene expression via p38 MAPK signaling in NIH3T3 fibroblast cells

One of the 14-3-3 protein isoforms, 14-3-3epsilon, was previously shown to be increased during skin aging. We suggest here a possible role for the 14-3-3epsilon protein in skin aging by providing evidence that 14-3-3epsilon increases the expression of the matrix-metalloproteinase (MMP)-2 gene in NIH3T3 fibroblast cells. Expression of the 14-3-3epsilon gene in NIH3T3 cells primarily up-regulated the expression of the MMP-2 gene at the transcriptional level by inducing specific DNA binding proteins bound to an upstream region of the MMP-2 promoter from -1,629 to -1,612. Inhibition of endogenous 14-3-3epsilon gene expression by RNA interference also decreased endogenous MMP-2 gene expression. Furthermore, up-regulation of the MMP-2 gene by 14-3-3epsilon was suppressed by expression of a dominant-negative mutant of p38 MAP kinase. These findings strongly suggest that increased expression of 14-3-3epsilon contributes to remodeling of extracellular matrix in skin through increasing MMP-2 gene expression via p38 MAP kinase signaling.

Etoposide-induced Smad6 expression is required for the G1 to S phase transition of the cell cycle in CMT-93 mouse intestinal epithelial cells

The inhibitory Smad6 and Smad7 are responsible for cross-talk between TGF-beta/bone morphogenic protein (BMP) signaling and other cellular signaling pathways, as well as negative feedback on their own signaling functions. Although inhibitory Smads are induced by various stimuli, little is known about the stimuli that increase Smad6 transcription, in contrast to Smad7. Here we demonstrate that etoposide, which induces double strand breaks during DNA replication, significantly up-regulates the transcription of the Smad6 gene in CMT-93 mouse intestinal cells by increasing specific DNA binding proteins. In addition, endogenous inhibition of the Smad6 gene by RNAi interference led to transient accumulation of G1 phase cells and reduction in incorporation of bromodeoxyuridine (BrdU). These findings strongly suggest that Smad6 plays a distinct role in the signaling of etoposide-induced DNA damage.

Changes in expression of cell cycle regulators after G1 progression upon repetitive thioacetamide treatment in rat liver

Repetitive low dose thioacetamide (TA) treatment of hepatocytes was found to induce cells in G2 arrest. In the present study, an attempt was made to investigate alterations in expression of cell cycle regulators after G1 progression in the same repetitive low dose TA treated hepatocytes system and to define the determinators involved in G2 arrest. TA was daily administered intraperitoneally, with a dose of 50 mg/kg for 7 days. Expression levels of cyclin E and CDK2 were similar, increased at day 1 and reached a peak at day 2. And they recycled from day 3 reaching a second peak at day 5. Expression level of cyclin A was similar to p27(Kip1) and p57(Kip2) but not to CDK2 and increased to a peak level at day 2. Expression levels of cyclin B1 and cdc2 were similar although the cyclin B1 level was generally low, decreased from day 1 to basal levels at day 3 and persisted at a low level till day 7. The expression level of cyclin G1 was similar to p53 that peaked at day 3 and again at day 6 elevated over basal level. BrdU-labeled hepatocytic nuclei increased from 12 h, reached a peak at day 2, then decreased, and were not detectable from day 6. The number of PCNA-labeled nuclei increased immediately, peaked at day 2, and maintained till day 7. These results suggest that G2 arrest induced by repeated TA treatment might be p53-dependent, via activation of cyclin G1, rather than inhibition of cyclin B1- cdc2 complex, and inhibitors holding S phase progression might be p27(Kip1) and p57(Kip2).