5606

MAP2K3

mitogen-activated protein kinase kinase 3;MAP2K3;mitogen-activated protein kinase kinase 3

MAP kinase kinase 3|MAPK/ERK kinase 3|MAPKK 3|MEK 3|SAPK kinase 2|dual specificity mitogen-activated protein kinase kinase 3|stress-activated protein kinase kinase 2

17q11.2

protein-coding

In primary mammalian cells, oncogenic ras induces premature senescence, depending on an active MEK-extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase (MAPK) pathway. It has been unclear how activation of the mitogenic MEK-ERK pathway by ras can confer growth inhibition. In this study, we have found that the stress-activated MAPK, p38, is also activated during the onset of ras-induced senescence in primary human fibroblasts. Constitutive activation of p38 by active MKK3 or MKK6 induces senescence. Oncogenic ras fails to provoke senescence when p38 activity is inhibited, suggesting that p38 activation is essential for ras-induced senescence. Furthermore, we have demonstrated that p38 activity is stimulated by ras as a result of an activated MEK-ERK pathway. Following activation of MEK and ERK, expression of oncogenic ras leads to the accumulation of active MKK3/6 and p38 activation in a MEK-dependent fashion and subsequently induces senescence. Active MEK1 induces the same set of changes and provokes senescence relying on active p38. Therefore, oncogenic ras provokes premature senescence by sequentially activating the MEK-ERK and MKK3/6-p38 pathways in normal, primary cells. These studies have defined the molecular events within the ras signaling cascade that lead to premature senescence and, thus, have provided new insights into how ras confers oncogenic transformation in primary cells.

PURPOSE: immortalization is one of the first changes in cells undergoing carcinogenic transformation. Proteome profiling of the immortalization-senescence transition is expected to provide insights into the molecular mechanisms of early tumorigenesis. EXPERIMENTAL DESIGN: 2-DE and MALDI-MS were used to identify proteins in primary human breast epithelial cells, relevant to the immortalization-senescence transition. cell and molecular biology and immunohistochemistry were used to validate involvement of mitogen-activated protein kinase kinase 3 (MAP2K3) in the immortalization-senescence transition. RESULTS: we identified 71 proteins whose expression changed upon induction of senescence. The identified proteins include regulators of cell growth, death, cell assembly and organization. Analysis of the network formed by the identified proteins suggested that the immortalization-to-senescence transition could affect regulators of the cell cycle, protein synthesis, transport, post-translational modifications, DNA recombination and repair, and lipid and amino acid metabolism. We observed that MAP2K3 was downregulated in immortal human breast epithelial cells and that upregulation of MAP2K3 expression promoted cell senescence. Decreased expression of MAP2K3 was observed in human breast infiltrating ductal carcinomas, as compared to non-cancerous human breast tissues. CONCLUSION AND CLINICAL RELEVANCE: we described a proteome profile of the immortalization-to-senescence transition for human breast epithelial cells, and identified MAP2K3 as a protein that promotes senescence in these cells.