Separating the Anti-apoptotic and Mitotic Roles of Survivin

Abstract

Survivin is a bifunctional protein that acts as a suppressor of apoptosis and has an essential role in mitosis. To date whether these two functions can be divorced has not been addressed. Here we show that the linker region between the BIR (baculovirus inhibitor of apoptosis repeat) domain of survivin and COOH-terminal ␣ helix may be the key to separating its roles. When overexpressed survivin is present in interphase cells and shuttles between the cytoplasm and nucleus. Here we identify a rev-like nuclear exportation signal (NES) in the central domain of survivin and demonstrate that point mutations within this region cause accumulation of survivin in the nucleus. Interestingly cells expressing NES mutants exhibit reduced survival after X-irradiation. Moreover, cells expressing survivin L98A -green fluorescent protein (GFP) showed increased poly(ADPribose) polymerase-cleavage and caspase-3 activity after tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) treatment compared with cells expressing full-length survivingreen fluorescent protein. These data suggest a direct link between the interphase localization of survivin and cellular responsiveness to apoptotic stimuli. Using a cell proliferation assay, we also found that ectopic expression of NES mutants can complement for depletion of endogenous survivin, indicating that they can execute the mitotic duties of survivin. Thus we demonstrate for the first time that 1) survivin has a functional NES; 2) nuclear accumulation of overexpressed survivin correlates with increased sensitivity of cells to ionising radiation; and 3) the anti-apoptotic and mitotic roles of survivin can be separated through mutation of its NES. Separating these two functions of survivin could open up new possibilities for therapeutic strategies aimed at eliminating cancer cells yet preserving normal cell viability.

Phosphorylation of survivin at threonine 34 inhibits its mitotic function and enhances its cytoprotective activity

“…Cells were seeded at 2 x 10 4 per well (24 well plate) in anti-biotic free media then transfected with 60 pmol of control or survivin specific siRNA (Ambion) using HyperFect (Qiagen), as described in. 2 Constructs were rendered resistant to RNAi-targetting by introduction of a silent mutation, G54C. 20,21 Immunoblotting.…”

“…20,21 Immunoblotting. Immunoblot analysis was performed as described in Colnaghi et al (2006). Antibodies used: anti-survivin (1:750, rabbit, Ab469, Abcam) and anti-tubulin (B512, 1:2000, Sigma).…”

“…When overexpressed in tumors, however, it is present throughout the cell cycle. Survivin expression confers resistance to irradiation and chemotherapy, [1][2][3] and correlates with refractory tumors in the clinic. 4,5 In addition to its role in mitosis, survivin is a member of the inhibitor of apoptosis family, IAP.…”

XPO1 Inhibition Enhances Radiation Response in Preclinical Models of Rectal Cancer

“…XPO1 plays an essential role in the function of survivin, as its cytoplasmic localization is required for antiapoptotic and tumor-promoting functions of survivin (27,28). Studies using mutants with export-deficient survivin showed that tumor cells were more sensitive to radiotherapy-induced apoptosis (28,31,32). On the basis of these studies, we hypothesize that blocking survivin nuclear export using XPO1 inhibitors could prevent radiation resistance.…”

Functional analysis of a survivin-like gene in Bombyx mori

“…It has been demonstrated that svv is a member of the chromosomal passenger complex (CPC) proteins and interacts with Aurora-B kinase, Borealin and inner centromere protein (INCENP) to play a significant role in cell division (Lens et al 2006;Ruchaud et al 2007). In addition, svv contains a Crm1-dependent nuclear export signal (NES), which is essential for the anti-apoptotic and regulatory role of svv during cell division and its correct localization (Stauber et al 2006;Habtemichael et al 2010;Knauer et al 2007a, b;Colnaghi et al 2006). This gene is highly expressed in cancers including neuroblastoma, gastric cancer, colorectal cancer, and bladder cancer (Adida et al 1998;Lu et al 1998;Kawasaki et al 1998;Swana et al 1999).…”