Proteasome activator PA28 gamma regulates p53 by enhancing its MDM2-mediated degradation.
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ABSTRACT:
SUBMITTER: Zhang Z
PROVIDER: S-EPMC2265109 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:PA28 (also known as 11S, REG or PSME) is a family of proteasome regulators whose members are widely present in many of the eukaryotic supergroups. In jawed vertebrates they are represented by three paralogs, PA28?, PA28?, and PA28?, which assemble as heptameric hetero (PA28??) or homo (PA28?) rings on one or both extremities of the 20S proteasome cylindrical structure. While they share high sequence and structural similarities, the three isoforms significantly differ in terms of their biochemical and biological properties. In fact, PA28? and PA28? seem to have appeared more recently and to have evolved very rapidly to perform new functions that are specifically aimed at optimizing the process of MHC class I antigen presentation. In line with this, PA28?? favors release of peptide products by proteasomes and is particularly suited to support adaptive immune responses without, however, affecting hydrolysis rates of protein substrates. On the contrary, PA28? seems to be a slow-evolving gene that is most similar to the common ancestor of the PA28 activators family, and very likely retains its original functions. Notably, PA28? has a prevalent nuclear localization and is involved in the regulation of several essential cellular processes including cell growth and proliferation, apoptosis, chromatin structure and organization, and response to DNA damage. In striking contrast with the activity of PA28??, most of these diverse biological functions of PA28? seem to depend on its ability to markedly enhance degradation rates of regulatory protein by 20S proteasome. The present review will focus on the molecular mechanisms and biochemical properties of PA28?, which are likely to account for its various and complex biological functions and highlight the common features with the PA28?? paralog.
| S-EPMC7915322 | biostudies-literature
Project description:Sirtuin 7 (SIRT7), an NAD+-dependent deacetylase, plays vital roles in energy sensing, but the underlying mechanisms of action remain less clear. Here, we report that SIRT7 is required for p53-dependent cell-cycle arrest during glucose deprivation. We show that SIRT7 directly interacts with p300/CBP-associated factor (PCAF) and the affinity for this interaction increases during glucose deprivation. Upon binding, SIRT7 deacetylates PCAF at lysine 720 (K720), which augments PCAF binding to murine double minute (MDM2), the p53 E3 ubiquitin ligase, leading to accelerated MDM2 degradation. This effect results in upregulated expression of the cell-cycle inhibitor, p21Waf1/Cip1, which further leads to cell-cycle arrest and decreased cell viability. These data highlight the importance of the SIRT7-PCAF interaction in regulating p53 activity and cell-cycle progression during conditions of glucose deprivation. This axis may represent a new avenue to design effective therapeutics based on tumor starvation.
| S-EPMC7286819 | biostudies-literature
Project description:PA28? is a nuclear activator of the 20S proteasome involved in the regulation of several essential cellular processes, such as cell proliferation, apoptosis, nuclear dynamics, and cellular stress response. Unlike the 19S regulator of the proteasome, which specifically recognizes ubiquitylated proteins, PA28? promotes the degradation of several substrates by the proteasome in an ATP- and ubiquitin-independent manner. However, its exact mechanisms of action are unclear and likely involve additional partners that remain to be identified. Here we report the identification of a cofactor of PA28?, PIP30/FAM192A. PIP30 binds directly and specifically via its C-terminal end and in an interaction stabilized by casein kinase 2 phosphorylation to both free and 20S proteasome-associated PA28?. Its recruitment to proteasome-containing complexes depends on PA28? and its expression increases the association of PA28? with the 20S proteasome in cells. Further dissection of its possible roles shows that PIP30 alters PA28?-dependent activation of peptide degradation by the 20S proteasome in vitro and negatively controls in cells the presence of PA28? in Cajal bodies by inhibition of its association with the key Cajal body component coilin. Taken together, our data show that PIP30 deeply affects PA28? interactions with cellular proteins, including the 20S proteasome, demonstrating that it is an important regulator of PA28? in cells and thus a new player in the control of the multiple functions of the proteasome within the nucleus.
| S-EPMC6048556 | biostudies-literature
Project description:Although ubiquitin receptor Rad23 has been implicated in bringing ubiquitylated p53 to the proteasome, how Rad23 recognizes p53 remains unclear. We demonstrate that XPC, a Rad23-binding protein, regulates p53 turnover. p53 protein in XPC-deficient cells remains ubiquitylated, but its association with the proteasome is drastically reduced, indicating that XPC regulates a postubiquitylation event. Furthermore, we found that XPC participates in the MDM2-mediated p53 degradation pathway via direct interaction with MDM2. XPC W690S pathogenic mutant is specifically defective for MDM2 binding and p53 degradation. p53 is known to become stabilized following UV irradiation but can be rendered unstable by XPC overexpression, underscoring a critical role of XPC in p53 regulation. Elucidation of the proteolytic role of XPC in cancer cells will help to unravel the detailed mechanisms underlying the coordination of DNA repair and proteolysis.
| S-EPMC3890342 | biostudies-literature
Project description:PA28? is a proteasome activator involved in the regulation of the cellular proliferation, differentiation and growth. In the present study, we identified and characterized a cDNA from Schistosoma mansoni exhibiting significant homology to PA28? of diverse taxa ranging from mammals (including humans) to simple invertebrates. Designated SmPA28?, this transcript has a 753bp predicted ORF encoding a protein of 250 amino acid residues. Alignment of SmPA28? with multiple PA28? orthologues revealed an average similarity of ~40% among the investigated organisms, and 90% similarity with PA28? from Schistosoma japonicum. In addition, phylogenetic analysis demonstrated a close linkage between SmPA28? to its sister group that contains well-characterized PA28? sequences from Drosophila spp., as well as sharing the same branch with PA28? from S. japonicum. Gene expression profiling of SmPA28? using real-time quantitative PCR revealed elevated steady-state transcript levels in the eggs, miracidia and paired adult worms compared to other stages. In parallel with gene expression profiles, an affinity-purified anti-SmPA28? antibody produced against recombinant protein exhibited strongest reactivity in Western blot analyses to endogenous SmPA28? from miracidia, sporocysts and paired adult worms. Given its known regulatory function in other organisms, we hypothesized that the high level of SmPA28? transcript and protein in these stages may be correlated with an important role of the PA28? in the cellular growth and/or development of this parasite. To address this hypothesis, miracidia were transformed in vitro to sporocysts in the presence of SmPA28? double-stranded RNAs (dsRNAs) and cultivated for 4 days, after which time steady-state transcript and protein levels, and phenotypic changes were evaluated. SmPA28? dsRNA treatment resulted in gene and protein knockdown of ~60% and ~80%, respectively, which were correlated with a significant decrease in larval length compared to its controls. These findings are consistent with a putative role of SmPA28? in larval growth/development of the S. mansoni.
| S-EPMC3712533 | biostudies-literature
Project description:Cellular inhibitor of apoptosis proteins (cIAP1 and cIAP2) function to prevent apoptosis and are often overexpressed in various cancers. However, mutations in cIAP1/2 can activate the alternative NF?B pathway through I?B?-kinase-? (IKK?) and are associated with hematopoetic malignancies. In the current study, we found that knockdown of cIAP2 in human mammary epithelial cells resulted in activation of MDM2 through increased SUMOylation and profound reduction of the pool of MDM2 not phosphorylated at Ser166. cIAP2 siRNA markedly decreased p53 levels, which were rescued by addition of the MDM2 inhibitor, Nutlin3a. An IAP antagonist, which induces cIAP degradation, transiently increased MDM2 mRNA. Simultaneous transfection of siRNA for cIAP2 and IKK? reduced MDM2 protein, while expression of a kinase-dead IKK? strongly increased non-Ser166 P-MDM2. Inhibition of either IKK? or -? partially rescued p53 levels, while concomitant IKK?/? inhibition fully rescued p53 after cIAP2 knockdown. Surprisingly, IKK? knockdown alone increased SUMO-MDM2, suggesting that in the absence of activation, IKK? can prevent MDM2 SUMOylation. cIAP2 knockdown disrupted the interaction between the MDM2 SUMO ligase, PIAS1 and IKK?. Partial knockdown of cIAP2 cooperated with (V12) H-ras-transfected mammary epithelial cells to enhance colony formation. In summary, our data identify a novel role for cIAP2 in maintaining wild-type p53 levels by preventing both an NF?B-mediated increase and IKK?/-?-dependent transcriptional and post-translational modifications of MDM2. Thus, mutations or reductions in cIAP2 could contribute to cancer promotion, in part, through downregulation of p53.
| S-EPMC3507496 | biostudies-literature
Project description:Adhesion G protein-coupled receptors (ADGRs) encompass 33 human transmembrane proteins with long N termini involved in cell-cell and cell-matrix interactions. We show the ADGRB1 gene, which encodes Brain-specific angiogenesis inhibitor 1 (BAI1), is epigenetically silenced in medulloblastomas (MBs) through a methyl-CpG binding protein MBD2-dependent mechanism. Knockout of Adgrb1 in mice augments proliferation of cerebellar granule neuron precursors, and leads to accelerated tumor growth in the Ptch1+/- transgenic MB mouse model. BAI1 prevents Mdm2-mediated p53 polyubiquitination, and its loss substantially reduces p53 levels. Reactivation of BAI1/p53 signaling axis by a brain-permeable MBD2 pathway inhibitor suppresses MB growth in vivo. Altogether, our data define BAI1's physiological role in tumorigenesis and directly couple an ADGR to cancer formation.
| S-EPMC6002773 | biostudies-literature
Project description:Optic fissure fusion is a critical event during retinal development. Failure of fusion leads to coloboma, a potentially blinding congenital disorder. Pax2a is an essential regulator of optic fissure fusion and the target of numerous morphogenetic pathways. In our current study, we examined the negative regulator of pax2a expression, Nz2, and the mechanism modulating Nlz2 activity during optic fissure fusion. Upregulation of Nlz2 in zebrafish embryos resulted in downregulation of pax2a expression and fissure fusion failure. Conversely, upregulation of pax2a expression also led to fissure fusion failure suggesting Pax2 levels require modulation to ensure proper fusion. Interestingly, we discovered Nlz2 is a target of the E3 ubiquitin ligase Siah. We show that zebrafish siah1 expression is regulated by Hedgehog signaling and that Siah1 can directly target Nlz2 for proteasomal degradation, in turn regulating the levels of pax2a mRNA. Finally, we show that both activation and inhibition of Siah activity leads to failure of optic fissure fusion dependent on ubiquitin-mediated proteasomal degradation of Nlz2. In conclusion, we outline a novel, proteasome-mediated degradation regulatory pathway involved in optic fissure fusion.
| S-EPMC6602337 | biostudies-literature
Project description:Under normal, non-stressed conditions, intracellular p53 is continually ubiquitinated by MDM2 and targeted for degradation. However, in response to severe genotoxic stress, p53 protein levels are markedly increased and apoptotic cell death is triggered. Inhibiting the ubiquitination of p53 under conditions where DNA damage has occurred is therefore crucial for preventing the development of cancer, because if cells with severely damaged genomes are not removed from the population, uncontrolled growth can result. However, questions remain about the cellular mechanisms underlying the regulation of p53 stability. In this study, we show that p53-inducible gene 3 (PIG3), which is a transcriptional target of p53, regulates p53 stability. Overexpression of PIG3 stabilized both endogenous and transfected wild-type p53, whereas a knockdown of PIG3 lead to a reduction in both endogenous and UV-induced p53 levels in p53-proficient human cancer cells. Using both in vivo and in vitro ubiquitination assays, we found that PIG3 suppressed both ubiquitination- and MDM2-dependent proteasomal degradation of p53. Notably, we demonstrate that PIG3 interacts directly with MDM2 and promoted MDM2 ubiquitination. Moreover, elimination of endogenous PIG3 in p53-proficient HCT116 cells decreased p53 phosphorylation in response to UV irradiation. These results suggest an important role for PIG3 in regulating intracellular p53 levels through the inhibition of p53 ubiquitination.
| S-EPMC5499618 | biostudies-literature
Project description:The proteasome is a multisubunit proteolytic enzyme comprising activator complexes bound to the 20 S catalytic core. The functions of the proteasomal activator (PA) 700 in ubiquitin/ATP-dependent protein degradation and of the PA28 alpha/beta activators in antigen presentation are well defined. However, the function of a third PA, PA28 gamma, remains elusive. We now show that mitogen-activated protein kinase (MAPK)/extracellular-signal-regulated kinase (ERK) kinase kinase 3 (MEKK3), a MAPK kinase kinase (MAPKKK) involved in MAPK kinase 7 (MKK7)-c-Jun N-terminal kinase ('JNK') and MKK6-p38 signalling, can bind PA28 gamma but not PA28 alpha. In contrast, B-Raf, a MAPKKK specific for the MAPK/ERK kinase ('MEK')-ERK module, binds PA28 gamma and alpha. The PA28 gamma-binding domain of MEKK3 is located within its N-terminal regulatory domain (amino acids 1-178). Expression of MEKK3 in Cos-7 cells led to an increase in endogenous and co-expressed PA28 gamma protein levels, whereas kinase-deficient MEKK3 had no effect on PA28 gamma expression. Furthermore, in vitro assays indicated that PA28 gamma was a MEKK3 substrate. MEKK3 represents the first protein kinase capable of binding and phosphorylating a PA, and provides a potential mechanism to link stress-activated protein kinase signalling with the PA28 gamma-dependent proteasome.
| S-EPMC1223459 | biostudies-other