Disruption of PKB signaling restores polarity to cells lacking tumor suppressor PTEN

By limiting phosphotidylinositol 3,4,5-triphosphate (PIP(3)) levels, tumor suppressor PTEN not only controls cell growth but also maintains cell polarity required for cytokinesis and chemotaxis. To identify the critical targets of PIP(3) that link it to the cytoskeleton, we deleted secondary genes to reverse the deficiencies of pten- cells in Dictyostelium. The polarity defects in pten- cells correlate with elevated phosphorylations of PKB substrates. Deletion of AKT orthologue, PkbA, or a subunit of its activator TORC2, reduced the phosphorylations and suppressed the cytokinesis and chemotaxis defects in pten- cells. In these double mutants, the excessive PIP(3) levels and, presumably, activation of other PIP(3)-binding proteins had little or no effect on the cytoskeleton. In bands with increased phosphorylation in pten- cells, we found PKB substrates, PI5K, GefS, GacG, and PakA. Disruption of PakA in pten- cells restored a large fraction of the cells to normal behavior. Consistently, expression of phosphomimetic PakA in pten- cells exacerbated the defects but nonphosphorylatable PakA had no effect. Thus, among many putative PTEN- and PIP(3)-dependent events, phosphorylation of PKB substrates is the key downstream regulator of cell polarity.     

The tumor suppressor PTEN regulates T cell survival and antigen receptor signaling by acting as a phosphatidylinositol 3-phosphatase

The tumor suppressor gene PTEN encodes a 55-kDa enzyme that hydrolyzes both protein phosphotyrosyl and 3-phosphorylated inositol phospholipids in vitro. We have found that the latter activity is physiologically relevant in intact T cells. Expression of active PTEN lead to a 50% loss of transfected cells due to increased apoptosis, which was completely prevented by coexpression of a constitutively active, membrane-bound form of protein kinase B. A mutant of PTEN selectively lacking lipid phosphatase activity, but retaining protein phosphatase activity, had no effects on cell number. Active (but not mutant) PTEN also decreased TCR-induced activation of the mitogen-activated protein kinase ERK2 (extracellular signal-related kinase 2), as seen after inhibition of phosphatidylinositol 3-kinase. Our data indicate that PTEN is a phosphatidylinositol 3-phosphatase in T cells, and we suggest that PTEN may play a role in the regulation of T cell survival and TCR signaling by directly opposing phosphatidylinositol 3-kinase.     

PTEN: life as a tumor suppressor

PTEN, a tumor suppressor located at chromosome 10q23, is mutated in a variety of sporadic cancers and in two autosomal dominant hamartoma syndromes. PTEN is a phosphatase which dephosphorylates phosphatidylinositol (3,4,5)-triphosphate (PtdIns-3,4,5-P3), an important intracellular second messenger, lowering its level within the cell. By dephosphorylating PtdIns-3,4,5-P3, PTEN acts in opposition to phosphatidylinositol 3-kinase (PI3K), which has a pivotal role in the creation of PtdIns-3,4,5-P3. PtdIns-3,4,5-P3 is necessary for the activation of Akt, a serine/threonine kinase involved in cell growth and survival. By blocking the activation of Akt, PTEN regulates cellular processes such as cell cycling, translation, and apoptosis. In this review, we will discuss the identification of PTEN, its mutational status in cancer, its role as a regulator of PI3K, and its domain structure.     

The tumor suppressor PTEN negatively regulates insulin signaling in 3T3-L1 adipocytes

PTEN is a tumor suppressor with sequence homology to protein-tyrosine phosphatases and the cytoskeleton protein tensin. PTEN is capable of dephosphorylating phosphatidylinositol 3,4, 5-trisphosphate in vitro and down-regulating its levels in insulin-stimulated 293 cells. To study the role of PTEN in insulin signaling, we overexpressed PTEN in 3T3-L1 adipocytes approximately 30-fold above uninfected or control virus (green fluorescent protein)-infected cells, using an adenovirus gene transfer system. PTEN overexpression inhibited insulin-induced 2-deoxy-glucose uptake by 36%, GLUT4 translocation by 35%, and membrane ruffling by 50%, all of which are phosphatidylinositol 3-kinase-dependent processes, compared with uninfected cells or cells infected with control virus. Microinjection of an anti-PTEN antibody increased basal and insulin stimulated GLUT4 translocation, suggesting that inhibition of endogenous PTEN function led to an increase in intracellular phosphatidylinositol 3,4,5-trisphosphate levels, which stimulates GLUT4 translocation. Further, insulin-induced phosphorylation of downstream targets Akt and p70S6 kinase were also inhibited significantly by overexpression of PTEN, whereas tyrosine phosphorylation of the insulin receptor and IRS-1 or the phosphorylation of mitogen-activated protein kinase were not affected, suggesting that the Ras/mitogen-activated protein kinase pathway remains fully functional. Thus, we conclude that PTEN may regulate phosphatidylinositol 3-kinase-dependent insulin signaling pathways in 3T3-L1 adipocytes.     

Regulation of the tumor suppressor PTEN by SUMO

The crucial function of the PTEN tumor suppressor in multiple cellular processes suggests that its activity must be tightly controlled. Both, membrane association and a variety of post-translational modifications, such as acetylation, phosphorylation, and mono- and polyubiquitination, have been reported to regulate PTEN activity. Here, we demonstrated that PTEN is also post-translationally modified by the small ubiquitin-like proteins, small ubiquitin-related modifier 1 (SUMO1) and SUMO2. We identified lysine residue 266 and the major monoubiquitination site 289, both located within the C2 domain required for PTEN membrane association, as SUMO acceptors in PTEN. We demonstrated the existence of a crosstalk between PTEN SUMOylation and ubiquitination, with PTEN-SUMO1 showing a reduced capacity to form covalent interactions with monoubiquitin and accumulation of PTEN-SUMO2 conjugates after inhibition of the proteasome. Moreover, we found that virus infection induces PTEN SUMOylation and favors PTEN localization at the cell membrane. Finally, we demonstrated that SUMOylation contributes to the control of virus infection by PTEN.     

Targeting the PDGF signaling pathway in tumor treatment

Platelet-derived growth factor (PDGF) isoforms and PDGF receptors have important functions in the regulation of growth and survival of certain cell types during embryonal development and e.g. tissue repair in the adult. Overactivity of PDGF receptor signaling, by overexpression or mutational events, may drive tumor cell growth. In addition, pericytes of the vasculature and fibroblasts and myofibroblasts of the stroma of solid tumors express PDGF receptors, and PDGF stimulation of such cells promotes tumorigenesis. Inhibition of PDGF receptor signaling has proven to useful for the treatment of patients with certain rare tumors. Whether treatment with PDGF/PDGF receptor antagonists will be beneficial for more common malignancies is the subject for ongoing studies.     

PTEN: a default gate-keeping tumor suppressor with a versatile tail

The tumor suppressor PTEN controls a variety of biological processes including cell proliferation, growth, migration, and death. As a master cellular regulator, PTEN itself is also subjected to deliberated regulation to ensure its proper function. Defects in PTEN regulation have a profound impact on carcinogenesis. In this review, we briefly discuss recent advances concerning PTEN regulation and how such knowledge facilitates our understanding and further exploration of PTEN biology. The carboxyl-tail of PTEN, which appears to be associated with multiple types of posttranslational regulation, will be under detailed scrutiny. Further, a comparative analysis of PTEN and p53 suggests while p53 needs to be activated to suppress tumorigenesis （a dormant gatekeeper）, PTEN is probably a constitutive surveillant against cancer development, thus a default gatekeeper.     

Suppressor of cytokine signaling 6 associates with KIT and regulates KIT receptor signaling

Suppressor of cytokine signaling (SOCS) proteins are a family of Src homology 2-containing adaptor proteins. Cytokine-inducible Src homology domain 2-containing protein, SOCS1, SOCS2, and SOCS3 have been implicated in the down-regulation of cytokine signaling. The function of SOCS4, 5, 6, and 7 are not known. KIT receptor signaling is regulated by protein tyrosine phosphatases and adaptor proteins. We previously reported that SOCS1 inhibited cell proliferation in response to stem cell factor (SCF). By screening the other members of SOCS family, we identified SOCS6 as a KIT-binding protein. Using KIT mutants and peptides, we demonstrated that SOCS6 bound directly to KIT tyrosine 567 in the juxtamembrane domain. To investigate the function of this interaction, we constitutively expressed SOCS6 in cell lines. Ectopic expression of SOCS6 in Ba/F3-KIT cell line decreased cell proliferation in response to SCF but not SCF-induced chemotaxis. SOCS6 reduced SCF-induced activation of ERK1/2 and p38 but not activation of AKT or STATs in Ba/F3, murine embryonic fibroblast (MEF), or COS-7 cells. SOCS6 did not impair ERK and p38 activation by other stimuli. These results indicate that SOCS6 binds to KIT juxtamembrane region, which affects upstream signaling components leading to MAPK activation. Our results indicate that KIT signaling is regulated by several SOCS proteins and suggest a putative function for SOCS6 as a negative regulator of receptor tyrosine kinases.     

Proteome changes induced by expression of tumor suppressor PTEN

The tumor suppressor, PTEN, located at 10q23, is one of the most frequently mutated tumor suppressors in a number of sporadic cancers and in two autosomal dominant harmatomas. It is considered one of the most important tumor suppressors in the post p53 era. To identify the molecules involved in the signal network regulated by PTEN using proteomic tools, a PTEN-inducible expression system was established in NIH 3T3 mouse embryonic fibroblast cells. We compared proteome images of PTEN-induced and non-induced cells by 2-dimensional electrophoresis. Twenty-nine differentially expressed protein spots were identified by MALDI-TOF MS and NSI MS/MS. We conclude that expression of PTEN by itself leads to protein profile changes, and those proteins affected are likely to be directly and/or indirectly involved in the function and physiology of the tumor suppressor.     

Redox regulation of the tumor suppressor PTEN by glutathione

Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) expressed in Saccharomyces cerevisiae was reversibly oxidized by hydrogen peroxide and reduced by cellular reductants. Reduction of hPTEN was delayed in each of S. cerevisiae gsh1Δ and gsh2Δ mutants. Expression of γ-glutamylcysteine synthetase Gsh1 in the gsh1Δ mutant rescued regeneration rate of hPTEN. Oxidized hPTEN was reduced by glutathione in a concentration- and time-dependent manner. Glutathionylated PTEN was detected. Incubation of 293T cells with BSO and knockdown expression of GCLc in HeLa cells by siRNA resulted in the delay of reduction of oxidized PTEN. Also, in HeLa cells transfected with GCLc siRNA, stimulation with epidermal growth factor resulted in the increase of oxidized PTEN and phosphorylation of Akt. These results suggest that the reduction of oxidized hPTEN is mediated by glutathione.     

神经系统中的肿瘤抑制因子PTEN

PTEN是定位于10q23的第一个具有磷酸酶活性的抑癌基因,它能通过PI3K/AKT、FAK和MAPK信号转导通路调节细胞的生长、凋亡、迁移和转化。PTEN在神经系统的神经元中有广泛表达,其在调节神经干细胞的增殖、SVZ前体细胞的迁移及凋亡、PC12细胞的分化和神经突触的建立方面具有重要作用。因此,对PTEN功能的进一步研究将为肿瘤和神经性疾病的治疗提供新的思路。     

Kinase-inactive G-protein-coupled receptor kinases are able to attenuate follicle-stimulating hormone-induced signaling

Homologous desensitization of G-protein-coupled receptors (GPCR) is thought to occur in several steps: binding of G-protein-coupled receptor kinases (GRKs) to receptors, receptor phosphorylation, kinase dissociation, and finally binding of beta-arrestin to phosphorylated receptors and functional uncoupling of the associated Galpha protein. It has recently been reported that GRKs can inhibit Galphaq-mediated signaling in the absence of phosphorylation of some GPCRs. Whether or not comparable phosphorylation-independent effects are also possible with Galphas-coupled receptors remains unclear. In the present study, using the tightly Galphas-coupled FSR receptor (FSH-R) as a model, we observed inhibition of the cAMP-dependent signaling pathway using kinase-inactive mutants of GRK2, 5, and 6. These negative effects occur upstream of adenylyl cyclase activation and are likely independent of GRK interaction with G protein alpha or beta/gamma subunits. Moreover, we demonstrated that, when overexpressed in Cos 7 cells, mutated GRK2 associates with the FSH activated FSH-R. We hypothesize that phosphorylation-independent dampening of the FSH-R-associated signaling could be attributable to physical association between GRKs and the receptor, subsequently inhibiting G protein activation.     

Inhibition of mycobacterial infection by the tumor suppressor PTEN

The tumor suppressor PTEN is a lipid phosphatase that is frequently mutated in various human cancers. PTEN suppresses tumor cell proliferation, survival, and growth mainly by inhibiting the PI3K-Akt signaling pathway through dephosphorylation of phosphatidylinositol 3,4,5-triphosphate. In addition to it role in tumor suppression, the PTEN-PI3K pathway controls many cellular functions, some of which may be important for cellular resistance to infection. Currently, the intersection between tumorigenic signaling pathways and cellular susceptibility to infection is not well defined. In this study we report that PTEN signaling regulates infection of both noncancerous and cancerous cells by multiple intracellular mycobacterial pathogens and that pharmacological modulation of PTEN signaling can affect mycobacterial infection. We found that PTEN deficiency renders multiple types of cells hyper-susceptible to infection by Mycoplasma and Mycobacterium bovis Bacillus Calmette-Guérin (BCG). The lipid phosphatase activity of PTEN is required for attenuating infection. Furthermore, we found mycobacterial infection activates host cell Akt phosphorylation, and pharmacological inhibition of Akt or PI3K activity reduced levels of intracellular infection. Intriguingly, inhibition of mTOR, one of the downstream components of the Akt signaling and a promising cancer therapeutic target, also lowered intracellular Bacillus Calmette-Guérin levels in mammary epithelial cancer MCF-7 cells. These findings demonstrate a critical role of PTEN-regulated pathways in pathogen infection. The relationship of PTEN-PI3K-Akt mTOR status and susceptibility to mycobacterial infection suggests that the interaction of mycobacterial pathogens with cancer cells may be influenced by genetic alterations in the tumor cells.     

PDGF receptor signaling networks in normal and cancer cells

For about four decades, platelet-derived growth factors (PDGF) and their receptors have been the subject of intense research, revealing their roles in embryo development and human diseases. Drugs such as imatinib, which selectively inhibit the tyrosine kinase activity of these receptors, have been approved for the treatment of cancers such as gastrointestinal stromal tumors and chronic eosinophilic leukemia. Today, the interest in these factors is still increasing in relationship with new potential clinical applications in cancer, stroke, fibrosis and infectious diseases. This review focuses on the mechanisms of PDGF receptor signaling, with an emphasis on pathways that are important for disease development. Of particular interest, recent studies revealed significant differences between normal and cancer cells regarding signal transduction by these growth factors.