Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P10415 (Bcl-2)
 33,771 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Fibroblast growth factor receptor signaling is an important mechanism regulating osteoblast function. To gain an insight into the regulatory role of FGF receptor-2 (FGFR2) signaling in osteoblasts, we investigated integrin-mediated attachment and cell survival in human calvarial osteoblasts expressing activated FGFR2. FGFR2 activation reduced osteoblast attachment on fibronectin. This was associated with reduced expression of the alpha5 integrin subunit normally expressed in human calvarial osteoblasts in vivo. Treatment with lactacystin, a potent inhibitor of proteasome, restored alpha5 integrin levels in FGFR2 mutant osteoblasts. Immunoprecipitation analysis showed that alpha5 integrin interacts with both the E3 ubiquitin ligase Cbl and ubiquitin. Immunocytochemistry revealed that alpha5 integrin colocalizes with FGFR2 and Cbl at the leading edge in membrane ruffle regions. Transfection with the 70Z-Cbl mutant lacking the RING domain required for Cbl-ubiquitin interaction, or with the G306E Cbl mutant that abolishes the binding ability of Cbl phosphotyrosine-binding domain restored alpha5 integrin levels. This suggests that Cbl-mediated ubiquitination plays an essential role in alpha5 integrin proteasome degradation induced by FGFR2 activation. Reduced alpha5 integrin expression was associated with an increased Bax/Bcl-2 ratio and increased caspase-9 and -3 activities in FGFR2 mutant osteoblasts. Forced expression of alpha5 integrin rescued cell attachment and corrected both the Bax/Bcl-2 ratio and caspase-3 and caspase-9 activities in FGFR2 mutant osteoblasts. We show that Cbl recruitment induced by FGFR2 activation triggers alpha5 integrin degradation by the proteasome, which results in reduced osteoblast attachment on fibronectin and caspase-dependent apoptosis. This identifies a functional role of the alpha5 integrin subunit in the induction of apoptosis triggered by FGFR2 activation in osteoblasts, and reveals that a Cbl-dependent mechanism is involved in the coordinated regulation of cell apoptosis induced by alpha5 integrin degradation.
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PMID:Cbl-mediated ubiquitination of alpha5 integrin subunit mediates fibronectin-dependent osteoblast detachment and apoptosis induced by FGFR2 activation. 1572 56

The elimination of Mcl-1, an anti-apoptotic Bcl-2 family member, is an early and required step for DNA damage-induced apoptosis. The degradation of Mcl-1 can be blocked by proteasome inhibitors, suggesting a role for the ubiquitin proteasome pathway in apoptosis. Here, we demonstrate that Mcl-1 is ubiquinated at five lysines. Biochemical fractionation of cell extracts allowed us to identify a 482 kDa HECT-domain-containing ubiquitin ligase named Mule (Mcl-1 ubiquitin ligase E3) that is both required and sufficient for the polyubiquitination of Mcl-1. Mule also contains a region similar to the Bcl-2 homology region 3 (BH3) domain that allows Mule to specifically interact with Mcl-1. Elimination of Mule expression by RNA interference stabilizes Mcl-1 protein, resulting in an attenuation of the apoptosis induced by DNA-damage agents. Thus, Mule is a unique BH3-containing E3 ubiquitin ligase apical to Bcl-2 family proteins during DNA damage-induced apoptosis.
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PMID:Mule/ARF-BP1, a BH3-only E3 ubiquitin ligase, catalyzes the polyubiquitination of Mcl-1 and regulates apoptosis. 1598 44

The ability to regulate apoptosis in mammalian cell cultures represents one approach to developing more economical and efficient processes. Genetic modification of cells using anti-apoptotic genes is one method that may be used to improve cellular performance. This study investigates a method to inhibit upstream apoptosis pathways through the overexpression of MDM2, an E3 ubiquitin ligase for p53. Both 293 and CHO cells expressing MDM2 were examined under both batch and spent media conditions. For batch cultures, MDM2 overexpression increased viable cell densities and viabilities over control cells with the largest enhancements observed in CHO cells. When CHO cells were passaged without medium exchange, cells expressing MDM2 reached a viable cell density that was nearly double the control and survived for an extra day in culture. When exposed to spent media initially, both 293-MDM2 and CHO-MDM2 cells continued to grow for 2 days while the control cells stopped growing after the first day. DNA analysis using flow cytometry confirmed that while CHO controls were found to be undergoing DNA fragmentation, CHO-MDM2 cells exhibit DNA degradation at a much slower rate. When compared to Bcl-2-expressing cells, MDM2 expression showed greater protection against apoptosis in passaged culture, spent medium, and following transient p53 overexpression. However, expression of the RING sequence of MDM2 responsible for E3 ligase activity without the other components of the protein was found to be toxic to 293 cells in culture. These results suggest that the overexpression of heterologous MDM2 represents a promising method to delay apoptosis in mammalian cell cultures.
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PMID:Inhibiting the apoptosis pathway using MDM2 in mammalian cell cultures. 1714 74

Netrins and their receptors deleted in colon cancer (DCC), neogenin, UNC5, and integrins are involved in axon guidance, epithelial morphogenesis, vascular pattering, cancer cell survival, invasion, tumor angiogenesis, and metastasis. Here, we considered the possible contribution of the p53-related apoptosis mediators p63 and p73 in the mechanisms underlying the antagonism between netrin-1 and DCC at the cell death control. We have showed that ectopic expression and external addition of netrin-1 in HeLa and HEK-293 cells with inactive p53 lead to impaired cell viability and induction of apoptosis. These responses were associated with up-regulation of the proapoptotic protein TAp73alpha, decreased Bcl-2/Bax ratio, and caspase-3 cleavage, with no change in protein levels of the antiapoptotic NH(2)-terminal-truncated DeltaNp73alpha isoform, p73 adapter Yap-1 and p73 E3 ubiquitin ligase Itch, and p63, as well as the transcripts encoding p63, TAp73alpha, and DeltaNp73alpha. However, the proteasome inhibitor MG132 potentiated, while DCC counteracted, netrin-1-induced TAp73alpha. Consistently, netrin-1 expression correlated with stabilization of the TAp73alpha protein and lower levels of TAp73alpha ubiquitination that was conversely enhanced by DCC, in a netrin-dependent manner. Our data indicate that netrin-1 selectively up-regulates TAp73alpha by preventing its ubiquitination and degradation. Targeted repression of p73alpha by shRNA reversed TAp73alpha and the apoptosis induced by netrin-1, and exacerbated the growth of HeLa tumor xenografts. Apoptosis induced by cisplatin was markedly enhanced in netrin-1 or DCC-expressing cells. Collectively, our data reveal that the transcriptionally active TAp73alpha tumor suppressor is implicated in the apoptosis induced by netrin-1 in a p53-independent and DCC/ubiquitin-proteasome dependent manner.
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PMID:Netrin-1 induces apoptosis in human cervical tumor cells via the TAp73alpha tumor suppressor. 1892 94

Bax, a multi-domain pro-apoptotic Bcl-2 family member, is a key regulator for the release of apoptogenic factors from mitochondria. MOAP-1, which was first isolated from a screen for Bax-associating proteins, interacts with Bax upon apoptotic induction. MOAP-1 is a short-lived protein that is constitutively degraded by the ubiquitin-proteasome system. Apoptotic stimuli upregulate MOAP-1 rapidly through inhibition of its poly-ubiquitination process. However, cellular factors that regulate the stability of MOAP-1 have not yet been identified. In this study, we report the identification of TRIM39 as a MOAP-1-binding protein. TRIM39 belongs to a family of proteins characterized by a Tripartite Motif (TRIM), consisting of RING domain, B-box and coiled-coil domain. Several TRIM family members are known to demonstrate E3 ubiquitin ligase activity. Surprisingly, TRIM39 significantly extends the half-life of MOAP-1 by inhibiting its poly-ubiquitination process. In agreement with its effect on enhancing MOAP-1 stability, TRIM39 sensitizes cells to etoposide-induced apoptosis. Conversely, knockdown of TRIM39 reduces the sensitivity of cells to etoposide-stimulated apoptosis. Furthermore, TRIM39 elevates the level of MOAP-1 in mitochondria and promotes cytochrome c release from isolated mitochondria stimulated by recombinant Bax. Together, these data suggest that TRIM39 can promote apoptosis signalling through stabilization of MOAP-1.
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PMID:TRIM39 is a MOAP-1-binding protein that stabilizes MOAP-1 through inhibition of its poly-ubiquitination process. 1910 Feb 60

Atrogin-1/MAFbx is a major atrophy-related E3 ubiquitin ligase that is expressed specifically in striated muscle. Although the contribution of atrogin-1 to cardiac and muscle hypertrophy/atrophy has been examined extensively, it remains unclear whether atrogin-1 plays an essential role in the simulated ischemia/reperfusion-induced apoptosis of primary cardiomyocytes. Here we showed that atrogin-1 markedly enhanced ischemia/reperfusion-induced apoptosis in cardiomyocytes via activation of JNK signaling. Overexpression of atrogin-1 increased phosphorylation of JNK and c-Jun and decreased phosphorylation of Foxo3a. In addition, atrogin-1 decreased Bcl-2, increased Bax, and enhanced the activation of caspases. Furthermore, JNK inhibitor SP600125 markedly blocked the effect of atrogin-1 on cell apoptosis and the expression of apoptotic-related proteins and caspases. Importantly, atrogin-1 induced sustained activation of JNK through a mechanism that involved degradation of MAPK phosphatase-1 (MKP-1) protein. Atrogin-1 interacted with and triggered MKP-1 for ubiquitin-mediated degradation. In contrast, proteasome inhibitors markedly blocked the degradation of MKP-1. Taken together, these results demonstrate that atrogin-1 promotes degradation of MKP-1 through the ubiquitin-proteasome pathway, thereby leading to persistent activation of JNK signaling and further cardiomyocyte apoptosis following ischemia/reperfusion injury.
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PMID:Atrogin-1/MAFbx enhances simulated ischemia/reperfusion-induced apoptosis in cardiomyocytes through degradation of MAPK phosphatase-1 and sustained JNK activation. 1911 50

The Sindbis viral expression system enables the rapid production of high levels of recombinant protein in mammalian cells; however, this expression is typically limited to transient production due to the cytotoxicity of the virus. Limiting the lethality inherent in the Sindbis virus vector in order to enable long term, sustained expression of recombinant proteins may be possible. In this study, modifications to virus and host have been combined in order to reduce the cytopathic effects. Non-cytopathic replication competent viruses of two Sindbis viral strains, TE and 633, were developed using a non-structural protein (nsP) P726S point mutation in order to obtain persistent heterologous gene expression in infected Baby Hamster Kidney (BHK) cells and Chinese Hamster Ovary (CHO) cells. Cells infected with the P726S variant viruses were able to recover after infection, while cells infected with normal virus died within 3 days. The P726S mutation did not reduce the susceptibility of 5- and 14-day-old mice to 633 and TE viruses in vivo. In addition, animal survival with the P726S variant viruses was increased and GFP expression was sustained for at least 14 days while the 633 and TE infection resulted in short-term GFP expression or an earlier mortality. Modifications to the host BHK and CHO cells themselves were subsequently undertaken by including the anti-apoptotic gene Bcl-2 and a deletion mutant of Bcl-2 (Bcl-2Delta) as another method for limiting the cytopathic effects of the Sindbis virus. The inclusion of anti-apoptotic genes permitted higher production of heterologous GFP protein following Sindbis virus infection, and the combination of the TE-P726S virus and the CHO-Bcl-2Delta cell line showed the greatest improvement in cell survival. Sindbis virus infection also induced ER stress in mammalian cells as detected by increased PERK phosphorylation and ATF4 translation. Overexpression of Parkin, an E3 ubiquitin ligase that can protect cells against agents that induce ER stress, suppressed Sindbis virus-induced cell death in both BHK cells and in vivo studies in mice. Such findings show that viral and host modifications can improve cell survival and production of heterologous proteins, change viral behavior in vitro and in vivo, and assist in the development of new expression or gene delivery vehicles.
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PMID:An improved in vitro and in vivo Sindbis virus expression system through host and virus engineering. 1920 Aug 10

The ability of thymocytes to assess T cell receptor (TCR) signaling strength and initiate the appropriate downstream response is crucial for determining their fate. We have previously shown that a c-Cbl RING finger mutant knock-in mouse, in which the E3 ubiquitin ligase activity of c-Cbl is inactivated, is highly sensitive to TCR-induced death signals that cause thymic deletion. This high intensity signal involves the enhanced tyrosine phosphorylation of the mutant c-Cbl protein promoting a marked increase in the activation of Akt. Here we show that this high intensity signal in c-Cbl RING finger mutant thymocytes also promotes the enhanced induction of two mediators of TCR-directed thymocyte apoptosis, Nur77 and the pro-apoptotic Bcl-2 family member, Bim. In contrast, a knock-in mouse harboring a mutation at Tyr-737, the site in c-Cbl that activates phosphatidylinositol 3-kinase, shows reduced TCR-mediated responses including suppression of Akt activation, a reduced induction of Nur77 and Bim, and greater resistance to thymocyte death. These findings identify tyrosine-phosphorylated c-Cbl as a critical sensor of TCR signal strength that regulates the engagement of death-promoting signals.
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PMID:c-Cbl promotes T cell receptor-induced thymocyte apoptosis by activating the phosphatidylinositol 3-kinase/Akt pathway. 2013 44

Bax, a pro-apoptotic protein from the Bcl-2 family, is central to apoptosis regulation. To suppress spontaneous apoptosis, Bax must be under stringent control that may include regulation of Bax conformation and expression levels. We report that IBRDC2, an IBR-type RING-finger E3 ubiquitin ligase, regulates the levels of Bax and protects cells from unprompted Bax activation and cell death. Downregulation of IBRDC2 induces increased cellular levels and accumulation of the active form of Bax. The ubiquitination-dependent regulation of Bax stability is suppressed by IBRDC2 downregulation and stimulated by IBRDC2 overexpression in both healthy and apoptotic cells. Although mostly cytosolic in healthy cells, upon induction of apoptosis, IBRDC2 accumulates in mitochondrial domains enriched with Bax. Mitochondrial accumulation of IBRDC2 occurs in parallel with Bax activation and also depends on the expression levels of Bcl-xL. Furthermore, IBRDC2 physically interacts with activated Bax. By applying Bax mutants in HCT116 Bax(-/-) cells, combined with the use of active Bax-specific antibodies, we have established that both mitochondrial localization and apoptotic activation of Bax are required for IBRDC2 translocation to the mitochondria.
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PMID:IBRDC2, an IBR-type E3 ubiquitin ligase, is a regulatory factor for Bax and apoptosis activation. 2030 62

Parkin is an E3 ubiquitin ligase that mediates the ubiquitination of protein substrates. The mutations in the parkin gene can lead to a loss of function of parkin and cause autosomal recessive juvenile onset parkinsonism. Recently, parkin was reported to be involved in the regulation of mitophagy. Here, we identify the Bcl-2, an anti-apoptotic and autophagy inhibitory protein, as a substrate for parkin. Parkin directly binds to Bcl-2 via its C terminus and mediates the mono-ubiquitination of Bcl-2, which increases the steady-state levels of Bcl-2. Overexpression of parkin, but not its ligase-deficient forms, decreases autophagy marker LC3 conversion, whereas knockdown of parkin increases LC3 II levels. In HeLa cells, a parkin-deficient cell line, knockdown of parkin does not change LC3 conversion. Moreover, overexpression of parkin enhances the interactions between Bcl-2 and Beclin 1. Our results provide evidence that parkin mono-ubiquitinates Bcl-2 and regulates autophagy via Bcl-2.
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PMID:Parkin mono-ubiquitinates Bcl-2 and regulates autophagy. 2088 74


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