- Research Article
- Open Access
Survivin Expression in Tuberous Sclerosis Complex Cells
© Feinstein Institute for Medical Research 2007
- Received: 25 October 2006
- Accepted: 4 January 2007
- Published: 1 March 2007
Tuberous Sclerosis Complex (TSC) is a tumor suppressor gene disorder with mutations of TSC1/TSC2 genes. This leads to the development of hamartomas that most frequently affect central nervous system, kidney, and skin. Angiomyolipomas are abdominal masses made up of muscle vessels and adipose tissues that grow mostly in proximity to kidneys and liver. Bleeding and kidney failure are the major justification for surgery. This study shows that angiomyolipoma-derived human smooth muscle TSC2−/− cells express the apoptosis inhibitor protein survivin when exposed to IGF-1. Survivin expression is also triggered whenever culture conditions perturb normal TSC2−/− cell function, such as the omission of EGF from the growth medium, the supplementation of anti-EGFR, blockade of PI3K and ERK, or inhibition of mTOR. Interestingly, single or simultaneous inhibition of PI3K by LY294002 and ERK by PD98059 does not prevent IGF-1-mediated survivin expression. Apoptogenic Smac/DIABLO, which is constitutively expressed by TSC2−/−A+ cells, is down-regulated by IGF-1 even in the presence of LY294002 and PD98059. These cells release IGF-1 by means of a negative feedback-regulated mechanism that is overrun when they are exposed to antibodies to IGF-1R, which increases the released amount by more than 400%. The autocrine release of IGF-1 may therefore be a powerful mechanism of survival of the tightly packed cells in the thick-walled vessels of TSC angiomyolipoma and in lymphangioleiomyomatosis (LAM) nodules. Future experimental therapies for TSC and LAM may result from the targeted inhibition of survivin, which may enhance sensitivity to TSC2 therapy.
Tuberous Sclerosis Complex (TSC) is an autosomal dominant syndrome with 95% penetrance, and a birth incidence of 1 in 11,000 (1). It is characterized by the multi-organ development of benign and occasionally malignant tumors that most frequently affect the central nervous system, abdomen, and skin (2). In particular, the kidney tumors include angiomyolipomas and renal cell carcinomas. Angiomyolipomas consist of smooth muscle cells, adipose tissue, and disorganized thick-walled vascular channels (3). Their abnormal vasculature may lead to spontaneous life-threatening bleeding (4). Angiomyolipomas increase in size and/or number in about 60% of children affected by TSC, highlighting the need for close surveillance of the kidneys and the entire abdomen (5). TSC can occur in association with pulmonary lymphangioleiomyomatosis (LAM), a progressive and often fatal interstitial lung disease characterized by the diffuse proliferation of abnormal smooth muscle cells and cystic degeneration of lung parenchyma. The TSC2 smooth muscle cells in angiomyolipomas are very similar to those of pulmonary LAM, and genetic data suggest that LAM may be the result of benign cell metastases, a highly unusual disease mechanism (6,7).
TSC is characterized by mutations in the tumor suppressor genes TSC1 on chromosome 9q34 or TSC2 on chromosome 16p13, which seem to act together as a complex of the encoded hamartin (TSC1) and tuberin (TSC2) proteins (8). These mutations lead to similar clinical phenotypes, which are more severe in the case of the TSC2 subtype (9). The TSC1/TSC2 complex negatively regulates cell size and proliferation (10,11) is a direct target of Akt, a PI3K-regulated effector that promotes cell growth and survival by means of a mammalian target of rapamycin (mTOR)-dependent mechanism (12). Most of the extracellular and intracellular signaling pathways involved in the regulation of growth factor- and nutrient-mediated cell growth are integrated by mTOR (13). Rapamycin is a microbial product that counteracts these effects by inhibiting mTOR, and Akt is a pro-survival and pro-oncogenic protein that is phosphorylated following the activation of growth factor receptors (13). Akt phosphorylation decreases the ability of TSC2 to inhibit the phosphorylation of the mTOR substrates S6 kinase and eukaryotic initiation factor 4E binding protein-1 (13–15), and excessive Akt, mTOR, and S6K activation causes various types of tumor, including hamartomas (2).
The inhibitors of apoptosis (IAP) gene family (16) is one of the most pursued targets among the regulators of apoptosis. Survivin is a structurally unique IAP protein (17) that is organized as a stable dimer (18) and contains a single baculovirus IAP repeat (BIR) and a COOH terminus coiled-coil domain (19). It has been suggested recently that survivin has dual function, as it is involved in both mitosis and apoptosis (20,21). Survivin expression is transcriptionally controlled in a cell cycle-dependent manner, triggered in the G2/M phase, and localized near the microtubules of centrosomes and mitotic spindle (22), and its down-regulation leads to multiple cell division defects with supernumerary centrosomes, multipolar mitotic spindles, and multinucleation (23–25).
Survivin also is involved in the suppression of cell death triggered by extrinsic or intrinsic apoptotic stimuli (26,27). When survivin expression is enhanced by genetic manipulation in transgenic mice, phenotypes consistent with its cytoprotective activity and enhanced cell viability are observed (28), whereas interference with its expression/function by means of antisense oligonucleotides, dominant negative mutants, ribozymes, or siRNA oligonucleotides triggers caspase-dependent apoptosis and increases apoptotic stimuli (24,29–32). Its antiapoptotic mechanism of action is not completely understood, but it has been suggested recently that it may affect the upstream initiation of mitochondrial-mediated apoptosis (33). Furthermore, its phosphorylation at threonine 34 allows the formation of a complex with caspase-9 and thus leads to the inhibition of apoptosis (34). In addition to preventing caspase-9 activation within a functional apoptosome (35), survivin binds the apoptogenic Smac/DIABLO released by mitochondrial membranes (36).
One of the most significant features of survivin is its expression in cancers and embryonic and fetal tissues (37), whereas its expression is very low or undetectable in normal differentiated tissues (38). Survivin up-regulation occurs in response to growth factor signaling (39,40), STAT activation (41), oncogene activation (42), loss of p53 (43,44), and PI3 kinase/Akt signaling (45).
We have isolated and characterized two cell populations from an angiomyolipoma in a patient affected by TSC2: one consists of β actin-positive TSC2 LOH smooth muscle-like cells (A+) and the other of keratin 8/18 positive TSC2 non-LOH epithelial-like cells (R+). Both cell types are HMB45 positive and require EGF for growth and proliferation (46). We report here that an angiomyolipoma in a TSC2 patient includes a small population of survivin-expressing cells, and that the same expression is also present in purified TSC2+/− epithelioid cells (R+). Survivin is not detectable in TSC2−/− smooth muscle-like cells (A+), but the addition of IGF-1 or removal of EGF from the growth medium, or the exposure of A+ cells to specific EGFR antibodies (Merck Biosciences, Darmstadt, Germany), promote survivin expression. A+ cells release IGF-1, and agents such as anti-EGFR antibodies increase IGF-1 release by up to about 13 ng/1.5 × 105 cells/24 h.
The SDS-PAGE materials came from Biorad (Hercules, CA, USA); LY294002, PD98059, rapamycin, staurosporine-A, wortmannin, cycloheximide (CHX), and recombinant human EGF (rhuEGF) came from Sigma-Aldrich (St. Louis, MO, USA); the fetal bovine serum and media were purchased from Euroclone (Paignton, UK); recombinant human IGF-1 (rhuIGF-1) was provided by Peprotech EC Ltd.; the polyclonal antibodies against survivin (FL-142), caspase-3 (H-277), RhoA, and β-actin came from Santa-Cruz Biotechnology (Santa-Cruz, CA, USA); the polyclonal antibody against β-actin came from Sigma-Aldrich, that against keratin 8/18 from Neomarkers (Freemont, CA, USA), and those against Smac/DIABLO and phospho-S6 from Cell Signaling (Beverly, MA, USA); the mouse monoclonal antibody against EGFR came from Merck Biosciences (Darmstadt, Germany), and that against IGF-1R from Calbiochem (Darmstadt, Germany). All of the other reagents were of high analytical grade and were purchased from Sigma-Aldrich.
The primary cultures were isolated from the angiomyolipoma of a 42-year-old female TSC2 patient previously described by Lesma et al. (46). The study was approved by the Institutional Review Board of Milan’s San Paolo Hospital. The β actin-positive TSC2 LOH smooth muscle-like cells (A+) and keratin 8/18 positive TSC2 non-LOH epitheliallike cells (R+) were kept in culture medium containing a 50/50 mixture of DMEM/Ham F12, supplemented with hydrocortisone 2 × 10−7 mol/L, rhuEGF 10 ng/mL, sodium selenite 5 × 10−8 mol/L (Sigma), insulin 25 µg/mL (Sigma), transferrin 10 µg/mL (Sigma), ferrous sulfate 1.6 × 10−6 (Si gma), and 15% heat-inactivated fetal bovine serum (FBS). C2C12 (mouse muscle myoblast) cells, CT/G vascular smooth muscle cells from human aorta (VSMCs), and A549 (human lung adenocarcinoma) cells were obtained from the American Type Culture Collection (ATCC, Rockville, MD, USA) and maintained in accordance with the manufacturer’s instructions: C2C12 in DMEM medium, A549 in RPMI 1640 medium, and VSMCs in F12 medium. FBS was added to all of the media at a concentration of 10%. All of the cell lines were kept at 37°C in a humidified atmosphere (5% CO2: 95% air). The cell cultures (except that of C2C12) were routinely sub-cultivated by means of trypsinisation to maintain long-phase growth. For the comparative studies of survivin expression, the cells were subconfluently grown as previously described.
Cell proliferation was evaluated in the presence of various agents added to the culture media by counting the cells in Neubauer chamber.
At the end of the treatments, the cell plates were placed on ice, the medium was removed, and the cells were lyzed in SDS-containing buffer (2% SDS, 0.1 M sodium acetate, 0.5 mM EDTA, 1 mM phenylmethylsulphonylfluoride, 1 mM aprotinin, 1 mM leupeptin, 2 mM sodium orthovanadate, 2 mM sodium fluoride). Angiomyolipoma was homogenized under isotonic condition (25 mM TrisHCl, PH 7.4, 250 mM sucrose, 1% Triton X-100, 1% NP-40, 5 mM EDTA, 1 mM EGTA, 1 mM phenylmethylsulphonylfluoride, 1 mM aprotinin, 1 mM leupetin, 2mM sodium orthovanadate, 2 mM sodium fluoride) using an Ultra-Turrax T25. The homogenates were centrifuged at 1500g for 20 min at 4°C, and the supernatant was collected and used for Western blot analysis.
Protein concentrations were determined using the Lowry Ciocalteau method with BSA as standard. Equal amounts (75 µg) of solubilized protein were dissolved in Laemmli sample buffer (Biorad, Hercules, CA, USA), separated by SDS-PAGE in reducing conditions, and electroblotted onto nitrocellulose membranes. The blots were blocked in 5% non-fat dry milk, 0.05% Tween-20 in TBS, pH 7.5, and probed with specific antibodies overnight at 4°C. The membranes were probed with horseradish peroxidase-conjugated goat anti-rabbit antibody (1:10,000) (Chemicon, Temecula, CA) at room temperature for 1 h. The protein was visualized by means of the enhanced chemioluminescence detection system (ECL; Pierce, Rockford, IL, USA), and the autoradiograms were analyzed using Kodak 1D 3.6 Software after acquisition on a Kodak image station 440 CF (Kodak).
Paraffin-embedded tissue sections of angiomyolipoma were stained with the anti-survivin polyclonal antibody raised against the full length of the protein (Santa Cruz). Briefly, the slides were de-waxed and rehydrated, treated with 3% hydrogen peroxide in distilled water to inhibit endogenous peroxidase activity and, after extensive washing in TBS/Triton X-100, incubated overnight at 4°C with a 1:200 dilution of the anti-survivin antibody. Sections were then rinsed and incubated with biotinylated goat anti-rabbit antibody (Pierce), and visualized using a standard ImmunoPure Ultrasensitive ABC Detection Kit (Pierce) with 3,3′-diaminobenzidine (DAB) as the chromogenic substrate. Positive and negative controls were included.
The cells were seeded and maintained in culture for 24 h before staining for survivin. Endogenous peroxidase activity was blocked with 3% hydrogen peroxide for 1 min at room temperature. Nonspecific background was eliminated by incubating the cells with 1.5% BSA in 0.2% Triton X-110 in PBS for 90 min, after which they were incubated overnight at 4°C in a humidified chamber with polyclonal antibody against survivin in 0.1% BSA. They were then rinsed and incubated with biotinylated goat anti-rabbit antibody (Chemicon) for 90 min at room temperature, and visualized using a standard ImmunoPure Ultrasensitive ABC Detection Kit (Pierce) with 3,3′-diaminobenzidine (DAB) as chromogen.
Cell Immunofluoroscence Microscopy
The cells were cultured on glass slides, permeabilized with 70% methanol for 10 min, and dried in air, and the primary antibodies against β-actin (1:100; Sigma), RhoA (1:100; Santa Cruz) and keratin 8/18 (1:100; Neomarkers) were applied overnight at 4°C. The samples were incubated for 3 h at room temperature with fluorescein isothiocyanate-conjugated rabbit anti-mouse antibody (Chemicon) for β-actin and keratin 8/18, and fluorescein-conjugated donkey anti-goat antibody (Chemicon) for RhoA. After washing, the slides were mounted in 50% glycerol with 1 µg/mL 4,6-diamono-2-phenylindole.
IGF-1 Release Assay
The IGF-1 released in the culture media was measured by means of an enzyme-linked immunosorbent assay (ELISA) for human IGF-1 (Biosource International) according to the manufacturer’s instructions. Briefly, the cells were seeded in 5 cm Petri dishes at a dilution of 1.5 × 105 cells/mL and, before testing, maintained in culture for different times (1, 5, 14 or 21 days) or treated as stated in the Results. Fresh media were collected and immediately stored at −80°C.
The data are expressed as mean values ± SEM, and were statistically analyzed using Student t-test or ANOVA followed by Tukey’s test; significance is indicated for P values of * < 0.05. All of the analyses were made using GraphPad Prism 4.0 software.
Survivin Expression in TSC2 Angiomyolipoma
The expression of survivin in human angiomyolipoma was also assayed by means of the Western blotting of four different samples (Figure 1B), with A549 cells being used as positive control (47). As survivin clearly plays an anti-apoptotic role (48), we also investigated cas-pase-3 by means of an antibody recognizing both the pro-peptide and the active form, and found that it was abundantly expressed in all of the angiomyolipoma samples (Figure 1B).
Survivin Expression in Isolated TSC2 Cells
Pharmacological Regulation of Survivin Expression in Isolated TSC2 Cells
The induction of survivin expression in A+ cells also was triggered when IGF-1 was added to the standard culture growth medium containing EGF (Figure 5B). IGF-1-mediated survivin expression in A+ cells was accompanied by the down-regulation of Smac/DIABLO. This was not observed when IGF-1 was added to the growth medium without EGF, and the increased survivin level was accompanied by the expression of Smac/DIABLO (Figure 5B). The absence of both growth factors led to the same survivin and Smac/DIABLO results. It is well known that survivin-mediated apoptosis suppression may occur via a direct interaction with the caspase activator Smac/DIABLO (36). None of the experimental conditions affected the expression of phospho-S6 and caspase-3. The pharmacological induction of survivin in TSC2 cells was quite rapid and did not require serum: it was expressed as little as 2 h after the addition of IGF-1 or LY to serum-deprived standard medium, and the inhibition of PI3K by LY did not antagonize the effect of IGF-1 on survivin expression (Figure 5C).
Cell Proliferation and Survivin Expression
Only rapamycin and anti-EGFR affected the proliferation rate during 48 h exposure, which suggests that the increased expression of survivin may be unrelated to proliferation in this instance (Figure 6C).
The results of this study show that angiomyolipoma cells from a TSC2 patient express the apoptosis inhibitor survivin. Two pure homogenous cell populations were isolated from the angiomyolipoma and characterized: TSC2−/− A+ smooth muscle cells and TSC2+/− R+ epithelioid cells. The latter show detectable levels of survivin, whereas the former express the protein when the growth medium is supplemented with IGF-1. Furthermore, survivin is expressed when the cells are exposed to conditions that perturb normal TSC2−/− cell function, such as the omission of EGF from the growth medium, the addition of anti-EGFR, the blockade of PI3K and ERK, or the inhibition of mTOR.
Interestingly, single or dual inhibition of PI3K by LY and ERK by PD does not prevent the triggering of survivin expression in TSC2−/− A+ cell exposed to IGF-1, therefore IGF-1 may play a role in the survival of these cells via a pathway that does not require PI3K and ERK function. Such a role is also supported by the IGF-1-mediated down-regulation of apoptogenic Smac/DIABLO, which is constitutively expressed by TSC2−/− A+ cells. Once again, the inhibition of PI3K by LY and ERK by PD does not prevent the marked reduction of Smac/DIABLO expression caused by IGF-1 exposure but, unlike that of survivin, its expression is not affected by LY or PD applied alone.
We found that the amounts of IGF-1 released by TSC2−/− A+ cells into the culture medium become progressively larger over time, being much higher on day 21 than on day one or five. This is probably an auto-regulated process with a feedback mechanism activated by the accumulation of IGF-1 in the growth medium because, during the first 10 h of 24 h collection, the amounts of IGF-1 released remained quite constant and were larger than later. However, this feedback regulation is overcome if TSC2−/− A+ cells are exposed to antibodies to IGF-1R, which increases the released amount by more than 400%. We have reported previously that TSC2−/− A+ cells exposed to anti-IGF-1R gradually die in 12–14 days (46), and so the autocrine release of IGF-1 seems to be a powerful survival mechanism activated by TSC2−/− smooth muscle cells which are tightly packed in the thick-walled vessels of TSC angiomyolipoma and lymphangioleiomyomatosis (LAM) nodules.
Survivin originally was isolated as a member of IAP family, and it was suggested that it plays a fundamental role in inhibiting apoptosis and regulating the mitotic process and cell proliferation (19,20). Its expression profile is indicative of this role as it is detectable in embryonic and fetal tissues, but poorly or not expressed in normal adult tissues (37). However, it is expressed in cancer cells, and higher levels of expression have been correlated with a poor prognosis (38); its expression also has been correlated with resistance to chemotherapeutic agents, thus suggesting that lower expression may indicate greater tumor sensitivity to chemotherapy (52,53). Survivin affects the initiation of mitochondrial-mediated apoptosis by binding the Smac/DIABLO released from mitochondria membranes, and then inhibiting caspase-9 activation (33–36).
Epithelioid TSC2+/− R+ cells constitutively express survivin, but it is undetectable in smooth muscle TSC2−/− A+ cells both in vivo and in vitro. However, its expression increases dramatically when the cells are exposed to agents that affect their function, such as LY, PD, and the mTOR inhibitor rapamycin. At a dose of 5 ng/mL, rapamycin affects the proliferation of smooth muscle TSC2−/− A+ cells (46), but LY and PD do not; the addition of anti-EGFR to the growth medium affects the rate of proliferation and promotes the expression of survivin. Both rapamycin and anti-EGFR increase survivin levels without affecting the constitutively expressed Smac/DIABLO, but their effects are time related: after five days’ treatment, the survivin-promoting effects of anti-EGFR or rapamycin have almost vanished, whereas Smac/DIABLO is still expressed at normal levels. We have reported previously that anti-EGFR-induced TSC2−/− A+ cell death begins after five and seven days’ exposure, and so it is possible that the progressive down-regulation of survivin may be one of the mechanisms triggered by anti-EGFR in killing these cells. In addition, anti-EGFR is probably a highly unfavorable signal for cells that require EGF to grow and proliferate (46): the 24-hour application of anti-EGFR enhances the autocrine release of IGF-1, which is comparable to that observed when the IGF-1 receptor is blocked by means of the specific antibody.
Epithelioid TSC2+/− R+ cells are very sensitive to EGF and its omission from the culture medium leads to a quick death, whereas smooth muscle TSC2−/− A+ cells stop proliferating but do not die when EGF is not added (46). Furthermore, R+ cells secrete significantly smaller amounts of IGF-1, about 10% of that released by A+ cells (data not shown). Both TSC2 cell types are therefore sensitive to EGF, but the extent of their dependence is rather different and R+ cells have a less developed ability to release IGF-1. Smooth muscle TSC2−/− A+ cells have developed a survival strategy that links the self-secretion of IGF-1 to the expression of anti-apoptotic survivin and the down-regulation of Smac/DIA-BLO. This survival mechanism seems to be quite specific as TSC2−/− A+ cells also show the constitutive expression of p53, which is genetically normal and without any mutation (unpublished data), and remains unaffected by IGF-1 or anti-EGFR exposure, and more generally by all of the experimental procedures used in this study (unpublished data).
Furthermore, IGF-1 supplementation promotes the proliferation of VSMCs and inhibits that of A+ cells. The role of IGF-1 has thus changed from inducing proliferation in normal smooth muscle to auto-secreted survival activity with no proliferative ability in smooth muscle TSC2−/− A+ cells (46).
The mechanisms developed by TSC2−/− A+ cells in terms of IGF-1 action and survivin expression are thus rather different from those described in cancer, in which the inhibition of PI3K leads to the down-regulation of survivin expression and increased sensitivity to chemotherapy (54–57), and CD34+ cells, in which the selective inhibition of PI3K and MAPK blocks the up-regulation of survivin by growth factors (58).
In conclusion, Tuberous Sclerosis Complex (TSC) is a tumor suppressor gene disorder characterized by mutations in the TSC1 or TSC2 genes that lead to the development of benign tumors involving smooth muscle cells, and also cause life-threatening LAMs. TSC2 is the form most often correlated with a poor clinical outcome. We have isolated and grown in vitro a pure colony of TSC2−/− smooth muscle cells from angiomyolipoma (christened A+) that require EGF to grow, and die in 12–14 days after exposure to EGFR and IGF-1R antibodies (46). These cells secrete much IGF-1 that is greatly increased when they are subjected to killing insults, such as exposure to EGFR and IGF-1R antibodies. Pharmacological treatments capable of interfering with the function of key enzymes such as PI3K, ERK, and mTOR, or with vital EGFR functions, such as the omission of EGF from growth medium or exposure to anti-EGFR, trigger the expression of survivin by TSC2−/− smooth muscle cells. The autocrine release of IGF-1 seems to be a powerful survival mechanism activated by TSC2−/− smooth muscle cells, which are tightly packed in the thick-walled vessels of TSC angiomyolipomas and LAM nodules, and in which IGF-1 concentrations may increase to reach effective anti-apoptotic concentrations.
Future experimental therapies for TSC and LAM will need to overcome this obstacle to achieve their goal, and a novel approach may be the targeted inhibition of survivin, which may enhance sensitivity to TSC2 therapy.
The authors acknowledge financial support to Alfredo Gorio from the Italian Ministry of Education and Research program, FIRB 2003.
- O’Callaghan FJ, Shiell AW, Osborne JP, Martyn CN (1998) Prevalence of tuberous sclerosis estimated by capture-recapture analysis. Lancet 351:1490.View ArticleGoogle Scholar
- Young J, Povey S (1998) The genetic basis of tuberous sclerosis. Mol. Med. Today. 4:313–9.View ArticleGoogle Scholar
- Karbowniczek M, Yu J, Henske EP (2003) Renal angiomyolipomas from patients with sporadic lymphangiomyomatosis contain both neoplastic and non-neoplastic vascular structures. Am. J. Pathol. 162:491–500.View ArticleGoogle Scholar
- Neumann HP, Schwarzkopf G, Henske EP (1998) Renal angiomyolipomas, cysts, and cancer in tuberous sclerosis complex. Semin. Pediatr. Neurol. 5:269–75.View ArticleGoogle Scholar
- Henske EP (2005) Tuberous sclerosis and the kidney: from mesenchyme to epithelium, and beyond. Pediatr. Nephrol. 20:854–7.View ArticleGoogle Scholar
- Henske EP (2003) Metastasis of benign tumor cells in tuberous sclerosis complex. Genes Chromosom. Cancer. 38:376–81.View ArticleGoogle Scholar
- Carsillo T, Astrinidis A, Henske EP (2000) Mutations in the tuberous sclerosis complex gene TSC2 are a cause of sporadic pulmonary lymphangioleiomyomatosis. Proc. Natl. Acad. Sci. USA. 97:6085–90.View ArticleGoogle Scholar
- van Slegtenhorst M et al. (1998) Interaction between hamartin and tuberin, the TSC1 and TSC2 gene products. Hum. Mol. Genet. 7:1053–7.View ArticleGoogle Scholar
- Dabora SL et al. (2001) Mutational analysis in a cohort of 224 tuberous sclerosis patients indicates increased severity of TSC2, compared with TSC1, disease in multiple organs. Am. J. Hum. Genet. 68:64–80.View ArticleGoogle Scholar
- Tapon N, Ito N, Dickson BJ, Treisman JE, Hariharan IK (2001) The drosophila tuberous sclerosis complex gene homologs restrict cell growth and cell proliferation. Cell 105:345–55.View ArticleGoogle Scholar
- Hengstschlager M et al. (2001) Tuberous sclerosis gene products in proliferation control. Mutat. Res. 488:233–9.View ArticleGoogle Scholar
- Edinger AL, Thompson CB (2002) Akt maintains cell size and survival by increasing mTOR-dependent nutrient uptake. Mol. Biol. Cell. 13:2276–88.View ArticleGoogle Scholar
- Inoki K, Li Y, Zhu T, Wu J, Guan KL (2002) TSC2 is phosphorylated and inhibited by Akt and suppresses mTOR signaling. Nat. Cell. Biol. 4:648–57.View ArticleGoogle Scholar
- Dan HC et al. (2002) PI3K/AKT pathway regulates TSC tumor suppressor complex by phosphorylation of tuberin. J. Biol. Chem. 277:35364–70.View ArticleGoogle Scholar
- Manning BD, Tee AR, Logsdon MN, Blenis J, Cantley LC (2002) Identification of the tuberous sclerosis complex-2 tumor suppressor gene product tuberin as a target of the phosphoinositide 3-kinase/akt pathway. Mol. Cell. 10:151–62.View ArticleGoogle Scholar
- Deveraux QL, Reed JC (1999) IAP family proteins—suppressors of apoptosis. Genes Dev. 13:239–52.View ArticleGoogle Scholar
- Salvesen GS, Duckett CS (2002) IAP proteins: blocking the road to death’s door. Nat. Rev. Mol. Cell. Biol. 3:401–10.View ArticleGoogle Scholar
- Verdecia MA et al. (2000) Structure of the human anti-apoptotic protein survivin reveals a dimeric arrangement. Nat. Struct. Biol. 7:602–8.View ArticleGoogle Scholar
- Ambrosini G, Adida C, Altieri DC (1997) Anovel anti-apoptosis gene, survivin, expressed in cancer and lymphoma. Nat. Med. 3:917–21.View ArticleGoogle Scholar
- Altieri DC (2004) Molecular circuits of apoptosis regulation and cell division control: the survivin paradigm. J. Cell. Biochem. 92:656–63.View ArticleGoogle Scholar
- Li F, Ling X (2006) Survivin study: an update of “what is next wave”? J. Cell. Physiol 208:476–86.View ArticleGoogle Scholar
- Li F et al. (1998) Control of apoptosis and mitotic spindle checkpoint by survivin. Nature. 396:580–4.View ArticleGoogle Scholar
- Chen J et al. (2000) Down-regulation of survivin by antisense oligonucleotides increases apoptosis, inhibits cytokinesis and anchorage-independent growth. Neoplasia. 2:235–41.View ArticleGoogle Scholar
- Li F et al. (1999) Pleiotropic cell-division defects and apoptosis induced by interference with survivin function. Nat. Cell. Biol. 1:461–6.View ArticleGoogle Scholar
- Olie RA et al. (2000) A novel antisense oligonucleotide targeting survivin expression induces apoptosis and sensitizes lung cancer cells to chemotherapy. Cancer Res. 60:2805–9.PubMedGoogle Scholar
- Reed JC, Bischoff JR (2000) BIRinging chromosomes through cell division—and survivin’ the experience. Cell. 102:545–8.View ArticleGoogle Scholar
- Grossman D et al. (2001). Transgenic expression of survivin in keratinocytes counteracts UVB-in-duced apoptosis and cooperates with loss of p53. J. Clin. Invest. 108:991–9.View ArticleGoogle Scholar
- Grossman D, Kim PJ, Schechner JS, Altieri DC (2001). Inhibition of melanoma tumor growth in vivo by survivin targeting. Proc. Natl. Acad. Sci. USA. 98:635–40.View ArticleGoogle Scholar
- Kanwar JR, Shen WP, Kanwar RK, Berg RW, Krissansen GW (2001) Effects of survivin antagonists on growth of established tumors and B7-1 immunogene therapy. J. Natl. Cancer Inst. 93:1541–52.View ArticleGoogle Scholar
- Yamamoto T, Manome Y, Nakamura M, Tanigawa N (2002) Downregulation of survivin expression by induction of the effector cell protease receptor-1 reduces tumor growth potential and results in an increased sensitivity to anticancer agents in human colon cancer. Eur. J. Cancer 38:2316–24.View ArticleGoogle Scholar
- Zhou M et al. (2002) DNA damage induces a novel p53-survivin signaling pathway regulating cell cycle and apoptosis in acute lymphoblastic leukemia cells. J. Pharmacol. Exp. Ther. 303:124–31.View ArticleGoogle Scholar
- Beltrami E, Plescia J, Wilkinson JC, Duckett CS, Altieri DC (2004) Acute ablation of survivin uncovers p53-dependent mitotic checkpoint functions and control of mitochondrial apoptosis. J. Biol. Chem. 279:2077–84.View ArticleGoogle Scholar
- McNeish IA et al. (2005) Survivin interacts with Smac/DIABLO in ovarian carcinoma cells but is redundant in Smac-mediated apoptosis. Exp. Cell Res. 302:69–82.View ArticleGoogle Scholar
- O’Connor DS et al. (2000) Regulation of apoptosis at cell division by p34cdc2 phosphorylation of survivin. Proc. Natl. Acad. Sci. USA. 97:13103–7.View ArticleGoogle Scholar
- Marusawa H et al. (2003) HBXIP functions as a cofactor of survivin in apoptosis suppression. EMBO J. 2:2729–40.View ArticleGoogle Scholar
- Song Z, Yao X, Wu M (2003) Direct interaction between survivin and Smac/DIABLO is essential for the anti-apoptotic activity of survivin during taxol-induced apoptosis. J. Biol. Chem. 278:23130–40.View ArticleGoogle Scholar
- Adida C, et al. (1998) Anti-apoptosis gene, survivin, and prognosis of neuroblastoma. Lancet 351:882–3.View ArticleGoogle Scholar
- Altieri DC (2001) The molecular basis and potential role of survivin in cancer diagnosis and therapy. Trends Mol. Med. 7:542–7.View ArticleGoogle Scholar
- Tran J et al. (1999) Marked induction of the IAP family antiapoptotic proteins survivin and XIAP by VEGF in vascular endothelial cells. Biochem. Biophys. Res. Commun. 64:781–8.View ArticleGoogle Scholar
- O’Connor DS et al. (2000) Control of apoptosis during angiogenesis by survivin expression in endothelial cells. Am. J. Pathol. 156:393–8.View ArticleGoogle Scholar
- Aoki Y, Feldman GM, Tosato G (2003) Inhibition of STAT3 signaling induces apoptosis and decreases survivin expression in primary effusion lymphoma. Blood. 101:1535–42.View ArticleGoogle Scholar
- Sommer KW et al. (2003) Inhibitor of apoptosis protein (IAP) survivin is upregulated by oncogenic c-H-Ras. Oncogene. 22:4266–80.View ArticleGoogle Scholar
- Mirza A et al. (2002) Human survivin is negatively regulated by wild-type p53 and participates in p53-dependent apoptotic pathway. Oncogene. 21:2613–22.View ArticleGoogle Scholar
- Hoffman WH et al. (2002) Transcriptional repression of the anti-apoptotic survivin gene by wild type p53. J. Biol. Chem. 277:3247–57.View ArticleGoogle Scholar
- Dan HC et al. (2004) Phosphatidylinositol-3-OH kinase/AKT and survivin pathways as critical targets for geranylgeranyltransferase I inhibitor-induced apoptosis. Oncogene. 23:706–15.View ArticleGoogle Scholar
- Lesma E et al. (2005) Isolation and growth of smooth muscle-like cells derived from tuberous sclerosis complex-2 human renal angiomyolipoma: EGF is the required growth factor. Am. J. Path. 167:1093–103.View ArticleGoogle Scholar
- Xu ZX et al. (2004) Promyelocytic leukemia protein 4 induces apoptosis by inhibition of survivin expression. J. Biol. Chem. 279:1838–44.View ArticleGoogle Scholar
- Altieri DC (2003) Validating survivin as a cancer therapeutic target. Nat. Rev. Cancer. 3:46–54.View ArticleGoogle Scholar
- Bonetti F et al. (1994) Clear cell (“sugar”) tumor of the lung is a lesion strictly related to angiomyolipoma—the concept of a family of lesions characterized by the presence of the perivascular epithelioid cells (PEC). Pathology. 26:230–6.View ArticleGoogle Scholar
- Bjornsson J et al. (1996) Tuberous sclerosis-associated renal cell carcinoma. Clinical, pathological, and genetic features. Am. J. Pathol. 149:1201–8.PubMedPubMed CentralGoogle Scholar
- Hengartner MO. (2000) The biochemistry of apoptosis. Nature. 407:770–6.View ArticleGoogle Scholar
- Ambrosini G, Adida C, Sirugo G, Altieri DC (1998) Induction of apoptosis and inhibition of cell proliferation by survivin gene targeting. J. Biol. Chem. 273:11177–82.View ArticleGoogle Scholar
- Kim PJ, Plescia J, Clevers H, Fearon ER, Altieri DC (2003) Survivin and molecular pathogenesis of colorectal cancer. Lancet. 362:205–9.View ArticleGoogle Scholar
- Asanuma H et al. (2005) Survivin expression is regulated by co-expression of human epidermal growth factor receptor 2 and epidermal growth factor via phosphatiylinositol 3-kinase/AKT signaling pathway in breast cancer cells. Cancer Res. 65:11018–25.View ArticleGoogle Scholar
- Ling X, Bernacki RJ, Brattain MG, Li F (2004) Induction of survivin expression by taxol (Paclitaxel) is an early event, which is independent of taxol-mediated G2/M arrest. J. Biol. Chem. 279:15196–203.View ArticleGoogle Scholar
- Zhang M, Latham DE, Delaney MA, Chakravarti A (2005) Survivin mediates resistance to antiandrogen therapy in prostate cancer. Oncogene. 24:2474–82.View ArticleGoogle Scholar
- Testa JR, Tsichilis PN (2005) AKT signaling in normal and malignant cells. Oncogene. 24:7391–3.View ArticleGoogle Scholar
- Fukuda S, Foster RG, Porter SB, Pelus LM (2002) The antiapoptosis protein survivin is associated with cell cycle entry of normal cord blood CD34+ cells and modulates cell cycle and proliferation of mouse hematopoietic progenitor cells. Blood 100:2463–71.View ArticleGoogle Scholar