Protein polyubiquitination

About: Protein polyubiquitination is a research topic. Over the lifetime, 45 publications have been published within this topic receiving 1220 citations.

Elevated Expression of ISG15 in Tumor Cells Interferes with the Ubiquitin/26S Proteasome Pathway

Abstract: IFN-stimulatory gene factor 15 (ISG15) is a ubiquitin-like protein, which is conjugated to many cellular proteins. However, its role in protein degradation is unclear. Here, we show that ISG15 is highly elevated and extensively conjugated to cellular proteins in many tumors and tumor cell lines. The increased levels of ISG15 in tumor cells were found to be associated with decreased levels of polyubiquitinated proteins. Specific knockdown of ISG15 expression using ISG15-specific small interfering RNA (siRNA) was shown to increase the levels of polyubiquitinated proteins, suggesting an antagonistic role of ISG15 in regulating ubiquitin-mediated protein turnover. Moreover, siRNA-mediated down-regulation of the major E2 for ISG15 (UbcH8), which blocked the formation of ISG15 protein conjugates, also increased the levels of polyubiquitinated proteins. Together, our results suggest that the ISG15 pathway, which is deregulated during tumorigenesis, negatively regulates the ubiquitin/proteasome pathway by interfering with protein polyubiquitination/degradation.

Protein degradation and protein synthesis in long-term memory formation

Abstract: Long-term memory (LTM) formation requires transient changes in the activity of intracellular signaling cascades that are thought to regulate new gene transcription and de novo protein synthesis in the brain. Consistent with this, protein synthesis inhibitors impair LTM for a variety of behavioral tasks when infused into the brain around the time of training or following memory retrieval, suggesting that protein synthesis is a critical step in LTM storage in the brain. However, evidence suggests that protein degradation mediated by the ubiquitin-proteasome system may also be a critical regulator of LTM formation and stability following retrieval. This requirement for increased protein degradation has been shown in the same brain regions in which protein synthesis is required for LTM storage. Additionally, increases in the phosphorylation of proteins involved in translational control parallel increases in protein polyubiquitination and the increased demand for protein degradation is regulated by intracellular signaling molecules thought to regulate protein synthesis during LTM formation. In some cases inhibiting proteasome activity can rescue memory impairments that result from pharmacological blockade of protein synthesis, suggesting that protein degradation may control the requirement for protein synthesis during the memory storage process. Results such as these suggest that protein degradation and synthesis are both critical for LTM formation and may interact to properly “consolidate” and store memories in the brain. Here, we review the evidence implicating protein synthesis and degradation in LTM storage and highlight the areas of overlap between these two opposing processes. We also discuss evidence suggesting these two processes may interact to properly form and store memories. LTM storage likely requires a coordinated regulation between protein degradation and synthesis at multiple sites in the mammalian brain.

Ubiquitin-conjugating enzyme Ubc13 controls breast cancer metastasis through a TAK1-p38 MAP kinase cascade

Abstract: Metastatic spread is the leading cause of cancer mortality. Breast cancer (BCa) metastatic recurrence can happen years after removal of the primary tumor. Here we show that Ubc13, an E2 enzyme that catalyzes K63-linked protein polyubiquitination, is largely dispensable for primary mammary tumor growth but is required for metastatic spread and lung colonization by BCa cells. Loss of Ubc13 inhibited BCa growth and survival only at metastatic sites. Ubc13 was dispensable for transforming growth factor β (TGFβ)-induced SMAD activation but was required for activation of non-SMAD signaling via TGFβ-activating kinase 1 (TAK1) and p38, whose activity controls expression of numerous metastasis promoting genes. p38 activation restored metastatic activity to Ubc13-deficient cells, and its pharmacological inhibition attenuated BCa metastasis in mice, suggesting it is a therapeutic option for metastatic BCa.

CSN5/Jab1 Is Involved in Ligand-Dependent Degradation of Estrogen Receptor α by the Proteasome

Abstract: Estradiol (E2) regulates target cell proliferation and gene transcription through series of molecular events initiated by the hormone-dependent binding of the estrogen receptor α (ERα), a member of the nuclear receptor superfamily, to its cognate DNA target. In mammary cells, the effects of estradiol can be antagonized by compounds such as 4-hydroxy-tamoxifen (OH-Tam), a tamoxifen metabolite that is a selective estrogen receptor modulator, and ICI 182,780 (ICI), a pure antiestrogen. Both OH-Tam and ICI antagonize estrogen action primarily by competing with estradiol for receptor binding. However, OH-Tam has a partial agonist activity, depending on the tissue and response examined (14), and ERα-OH-Tam complexes accumulate in nuclei (12, 37). In contrast, ICI compounds are totally devoid of agonist activity in the models studied to date (32), and ICI treatment provokes a rapid relocalization of ERα-ICI complexes to a salt-resistant nuclear fraction, followed by rapid degradation by the proteasome (12, 36). A complex pathway leading to transcriptional activation of target genes involves ERα structural changes upon hormone binding, allowing dynamic interactions of the hormone-receptor complex with DNA and coactivators. This will allow chromatin remodeling and the subsequent recruitment of coactivators and the formation of preinitiation complexes (for a review, see reference 24). ERα is downregulated in the presence of E2, its cognate ligand, through the ubiquitin/proteasome (Ub/26S) pathway (1). It has been proposed that ERα-mediated transcription and proteasome-mediated degradation are linked (17) and act to continuously turn over ERα on responsive promoters (20). Recently, it was proposed that the p160 coactivator steroid receptor coactivator 3 (SRC3)/AIB1 mediates agonist-induced ERα degradation (29). The 26S proteasome is a multiprotein complex composed of a 20S core complex and a 19S particle (base plus lid). It is involved in the degradation of short-lived proteins. The lid is thought to recognize ubiquitinated substrates that are then unfolded by the base and degraded in the proteolytic core complex. Protein ubiquitination is catalyzed by three critical factors: a ubiquitin-activating enzyme (E1), a ubiquitin-conjugating enzyme (E2), and a factor E3 or ubiquitin ligase (E3). Ubiquitin ligases termed SKP1-Cullin/CDC53-F-box protein (SCF) complexes catalyze the polyubiquitination of a number of proteins to target them for degradation. The neddylation (addition of the Ub-like protein NEDD8) of the cul1 (cullin1) subunit of the SCF stimulates protein polyubiquitination (23). The COP9 signalosome (CSN), a complex of eight proteins (CSN1 to CSN8), removes NEDD8 from cul1 (18, 27, 40). This deneddylase activity has recently been attributed to the MPM plus motif of the CSN5/Jab1 subunit (8) of CSN. It stops the polyubiquitination process and allows the subsequent degradation of polyubiquitinated substrates by the proteasome. It has been proposed that CSN serves as an assembly and maintenance platform for the cullin-based ubiquitin ligases (38). In addition, this complex exhibits significant sequence homologies to the eight subunits of the 26S proteasome lid complex. It is involved in regulating the stability of proteins such as p27Kip1 (34), c-jun (22), and p53 (4), which are substrates of the Ub/26S proteasome pathway. A kinase activity, inhibited by curcumin, is associated to the CSN complex. Recent data demonstrated an association of inositol 1,3,4-trisphosphate 5/6-kinase (31), casein kinase II (CKII), and protein kinase D (35) with the CSN complex. In the cell, two types of complexes containing CSN5/Jab1 have been described. One is the conventional 450-kDa CSN located in the nucleus and the other is a mini-CSN (approximately 100 kDa) containing only a subset of CSN components that is mainly located in the cytoplasm. The mini-CSN originates from the nuclear export of subunits CSN4 to -8 (33). As part of this mini-CSN, CSN5/Jab1 has been implicated in the nuclear export of the cyclin-dependent kinase inhibitor p27Kip1, playing the role of an adaptor between p27Kip1 and the nuclear transport protein CRM-1. The use of leptomycin B (LMB), a chemical inhibitor of CRM-1-dependent nuclear export, prevented p27Kip1 nuclear export and its CSN5/Jab1-mediated degradation (33). In addition, overexpression of CSN5/Jab1 caused the translocation of Smad7 from the nucleus to the cytoplasm, promoting its degradation (15). Several results suggest that CSN could be involved in the mechanism of action of estrogens: CSN5/Jab1 has been described as a coactivator of nuclear receptors (6), and the neddylation pathway is involved in ERα degradation (9, 10). The aim of our work was to investigate the contribution of CSN5/Jab1 to ERα ligand-dependent degradation and to the mechanism of transcription activation in response to estradiol. Here, we show that ERα and CSN5/Jab1 coimmunoprecipitate and that an increase in CSN5/Jab1 levels increases ligand-induced ERα degradation by the proteasome. Curcumin, which inhibits the kinase activity associated with CSN, prevents hormone-dependent ERα degradation, as well as estradiol-dependent phosphorylation of ERα. CSN5/Jab1 is also involved in the nuclear export of proteins. We show that LMB, an inhibitor of nuclear export, prevents estradiol-induced degradation of ERα, demonstrating that this degradation takes place in the cytoplasm. In contrast, for ERα complexed to ICI, a pure hormone antagonist, LMB had no effect, indicating that this degradation takes place in the nucleus. Finally, we investigated the effects of curcumin and LMB on estradiol-induced transcription. Curcumin, but not LMB, abolished estradiol-induced reporter gene activation. Curcumin prevented ERα binding to its cognate DNA target, indicating a probable involvement of CSN in a phosphorylation event, necessary for ERα binding. These results also demonstrate that ERα degradation is not required to sustain estradiol-induced transcription.

Effects of insulin-like growth factor-I, insulin, and leucine on protein turnover and ubiquitin ligase expression in rainbow trout primary myocytes

Abstract: The effects of insulin-like growth factor-I (IGF-I), insulin, and leucine on protein turnover and pathways that regulate proteolytic gene expression and protein polyubiquitination were investigated...