The role of dual-specificity phosphatase 1 and protein phosphatase 1 in (2)-adrenergic receptor-mediated inhibition of extracellular signal regulated kinase 1/2 in triple negative breast cancer cell lines

摘要

Triple negative breast cancer cell lines express high levels of (2)-adrenergic receptor, which have a significant influence on the activity of extracellular signal-regulated kinase (ERK)1/2. Therefore, it is important to understand the link between (2)-adrenergic receptor signaling and ERK1/2 activity in terms of cancer cell regulation and cancer progression. Although the molecular mechanisms are not completely clarified, (2)-adrenergic receptor stimulation appears to reduce the basal levels of phosphorylated (p)ERK1/2 in MDA-MB-231 breast cancer cells. The aim of the current study was to determine the mechanism of (2)-adrenergic receptor-mediated ERK1/2 dephosphorylation by investigating the role of dual-specificity phosphatase (DUSP)1/6 and protein phosphatase (PP)1/2, which are established regulators of ERK1/2 phosphorylation, in MDA-MB-231 and MDA-MB-468 breast cancer cell lines. (E)-2-benzylidene-3-(cyclohexyl amino)-2,3-dihydro-1H-inden-1-one (BCI) and calyculin A were employed as DUSP1/6 and PP1/PP2 inhibitors, respectively. Subsequently, the protein levels of DUSP1, PP1, pPP1, ERK1/2 and pERK1/2 were measured by western blot analysis. Cells were transfected with DUSP1 small interfering (si)RNA or PP1 siRNA to inhibit their expression. The results demonstrated that (2)-adrenergic receptor agonists led to the dephosphorylation of basal pERK1/2 in MDA-MB-231 and MDA-MB-468 cells. The DUSP1/6 inhibitor, BCI, and the PP1/PP2 inhibitor, calyculin A, antagonized the (2)-adrenergic receptor-mediated dephosphorylation of ERK1/2. Furthermore, (2)-adrenergic receptor stimulation increased the protein expression level of DUSP1, with no effects on DUSP6, PP1 and PP2 expression, and enhanced the expression of the active form of PP1. Downregulation of the expression of DUSP1 or PP1 led to a decline in the (2)-adrenergic receptor-mediated dephosphorylation of ERK1/2. The results of the present study indicate that (2)-adrenergic receptor-mediated dephosphorylation of ERK1/2 may be associated with the activity of DUSP1 and PP1 in MDA-MB-231 and MDA-MB-468 triple negative breast cancer cell lines. The clinical importance of (2)-adrenergic receptor-mediated inactivation of ERK1/2 as well as the activation of DUSP1 and PP1 should be carefully evaluated in future studies, particularly when (2)-adrenergic blockers are used in patients with triple negative breast cancer.