Journal: Cell Death & Disease
Article Title: FGF1 protects neuroblastoma SH-SY5Y cells from p53-dependent apoptosis through an intracrine pathway regulated by FGF1 phosphorylation
Figure Lengend Snippet: Role of extra- and intracellular FGF1 in neuronal SH-SY5Y, N2a and PC12 cells. ( a ) Both extra- and intra-cellular FGF1 protect human SH-SY5Y and rat PC12 cells from p53-dependent apoptosis (left). Endogenous fgf1 expression is increased by rFGF1 addition (top left). In contrast, extra- and intra-cellular FGF1 show no anti-apoptotic activity, and rFGF1 addition does not increase endogenous fgf1 expression in murine N2a cells (right). ( b ) Overexpression of intracellular FGF1 inhibits etoposide-induced apoptosis in human SH-SY5Y and rat PC12 cells by decreasing PUMA transactivation, mitochondrial membrane depolarization and permeabilization, and activation of caspase-9, caspase-3 as assessed by the cleavage of its substrate PARP (left). On the contrary, FGF1 overexpression does not protect N2a cells from etoposide-induced apoptosis, and modifies neither p53 activation, mitochondrial depolarization and permeabilization, nor cleavage of caspase-3 (right). ( c ) In human SH-SY5Y and rat PC12 cells, overexpression of wild-type FGF1 or non-phosphorylable FGF1 S130A inhibits p53-dependent apoptosis, while overexpression of the phosphomimetic FGF1 S130D or the FGF1 K132E mutant does not. Therefore, phosphorylation of FGF1 inhibits its anti-apoptotic activity in SH-SY5Y and PC12 cells. This figure compiles the results of the present study performed in neuroblastoma SH-SY5Y and N2a cells and of previous studies performed in pheochromocytoma PC12 cells 14 , 15 , 32
Article Snippet: The primary antibodies used in this study were: anti-FGF1 (AB-32-NA, R & D Systems), anti-His tag (A00186, GenScript, Piscataway, NJ, USA), anti-P-p53 (Ser-15) (9284S, Cell Signaling, Danvers, MA, USA), anti-PUMAα (N-19, Santa Cruz, Dallas, TX, USA), anti-Caspase-9 (5B4, Abcam, Cambridge, UK), anti-cleaved Caspase-3 (Asp175, Cell Signaling), anti-PARP (9542S, Cell Signaling) and anti-Actin (A2066, Sigma-Aldrich).
Techniques: Expressing, Activity Assay, Over Expression, Activation Assay, Mutagenesis