Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.30.2 (endonuclease)
18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

IRE1 proteins mediate cellular responses to accumulation of malfolded proteins in the endoplasmic reticulum in the yeast and mammalian unfolded protein responses. A sensitive in vivo u.v. crosslinking assay showed that IRE1 proteins are intimately associated with RNA in mammalian cells. The IRE1-associated RNA fragments recovered by this assay were different in stressed and unstressed cells. The amount of RNA associated with IRE1 that could be revealed by end-labeling with T4 kinase was greater in IRE1-containing complexes isolated from stressed cells. Furthermore, the RNA fragments recovered from complexes found in stressed cells were shorter than those from unstressed cells, revealing a dynamic change in the IRE1-RNA complex during the UPR. Formation of the complex between IRE1 and RNA was dependent on both the kinase and endonuclease domains of IRE1, and involved pre-existing RNA species. When viewed in the context of the known importance of Ire1p-HAC1 mRNA interactions to the yeast unfolded protein response, these findings suggest that full-length mammalian IRE1s also engage RNA molecules as downstream effectors.
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PMID:Alterations in an IRE1-RNA complex in the mammalian unfolded protein response. 1159 Feb 47

Changes in neuronal calcium activity in the various subcellular compartments have divergent effects on affected cells. In the cytoplasm and mitochondria, where calcium activity is normally low, a prolonged excessive rise in free calcium levels is believed to be toxic, in the endoplasmic reticulum (ER), in contrast, calcium activity is relatively high and severe stress is caused by a depletion of ER calcium stores. Besides its role in cellular calcium signaling, the ER is the site where membrane and secretory proteins are folded and processed. These calcium-dependent processes are fundamental to normal cell functioning. Under conditions of ER dysfunction unfolded proteins accumulate in the ER lumen, a signal responsible for activation of the unfolded protein response (UPR) and the ER-associated degradation (ERAD). UPR is characterized by activation of two ER-resident kinases, PKR-like ER kinase (PERK) and IRE1. PERK induces phosphorylation of the eukaryotic initiation factor (eIF2alpha), resulting in a shut-down of translation at the initiation step. This stress response is needed to block new synthesis of proteins that cannot be correctly folded, and thus to protect cells from the effect of unfolded proteins which tend to form toxic aggregates. IRE1, on the other hand, is turned after activation into an endonuclease that cuts out a sequence of 26 bases from the coding region of xbp1 mRNA. Processed xbp1 mRNA is translated into the respective protein, an active transcription factor specific for ER stress genes such as grp78. In acute disorders and degenerative diseases, the ER calcium pool is a primary target of toxic metabolites or intermediates, such as oxygen free radicals, produced during the pathological process. Affected neurons need to activate the entire UPR to cope with the severe form of stress induced by ER dysfunction. This stress response is however hindered under conditions where protein synthesis is suppressed to such an extent that processed xbp1 mRNA is not translated into the processed XBP1 protein (XBP1(proc)). Furthermore, activation of ERAD is important for the degradation of unfolded proteins through the ubiquitin/proteasomal pathway, which is impaired in acute disorders and degenerative diseases, resulting in further ER stress. ER functioning is thus impaired in two different ways: first by the direct action of toxic intermediates, produced in the course of the pathological process, hindering vital ER reactions, and second by the inability of cells to fully activate UPR and ERAD, leaving them unable to withstand the severe form of stress induced by ER dysfunction.
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PMID:Endoplasmic reticulum: a primary target in various acute disorders and degenerative diseases of the brain. 1290 82

Granzyme A, a serine protease in the cytotoxic granules of natural killer cells and cytotoxic T lymphocytes, induces caspase-independent cell death when introduced into target cells by perforin. Granzyme A induces single-stranded DNA damage as well as rapid loss of cell membrane integrity and mitochondrial transmembrane potential through unknown mechanisms. Granzyme A destroys the nuclear envelope by targeting lamins and opens up DNA for degradation by targeting histones. A special target of the granzyme A cell death pathway is an endoplasmic reticulum-associated complex, called the SET complex, which contains three granzyme A substrates, the nucleosome assembly protein SET, the DNA bending protein HMG-2, and the base excision repair endonuclease Ape1. The SET complex also contains the tumor suppressor protein pp32 and the granzyme A-activated DNase NM23-H1, which is inhibited by SET. Granzyme A cleavage of SET releases the inhibition and unleashes NM23-H1. Cleavage of Ape1 by granzyme A interferes with the ability of the target cell to repair itself. The novel cell death pathway initiated by granzyme A provides a parallel pathway for apoptosis, important in destroying targets that overexpress bcl-2 or are otherwise invulnerable to the caspases.
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PMID:Nuclear war: the granzyme A-bomb. 1449 64

Upon the accumulation of unfolded proteins in the mammalian endoplasmic reticulum (ER), X-box binding protein 1 (XBP1) premessenger RNA (premRNA) is converted to mature mRNA by unconventional splicing that is mediated by the endonuclease inositol-requiring enzyme 1. The transcription factor protein (p) XBP1 spliced (S), which is translated from mature XBP1 mRNA, contains the nuclear localization signal and the transcriptional activation domain and activates the transcription of target genes, including those encoding ER chaperones in the nucleus. We show that pXBP1 unspliced (U) encoded in XBP1 pre-mRNA was constitutively expressed and markedly accumulated at the recovery phase of ER stress. pXBP1(U) contained the nuclear exclusion signal instead of the transcriptional activation domain and shuttled between the nucleus and the cytoplasm. Interestingly, pXBP1(U) formed a complex with pXBP1(S), and the pXBP1(U)-pXBP1(S) complex was sequestered from the nucleus. Moreover, the complex was rapidly degraded by proteasomes because of the degradation motif contained in pXBP1(U). Thus, pXBP1(U) is a negative feedback regulator of pXBP1(S), which shuts off the transcription of target genes during the recovery phase of ER stress.
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PMID:pXBP1(U) encoded in XBP1 pre-mRNA negatively regulates unfolded protein response activator pXBP1(S) in mammalian ER stress response. 1646 60

DNase II is an acid endonuclease that is involved in the degradation of exogenous DNA and is important for DNA fragmentation and degradation during cell death. In an effort to understand its catalytic mechanism, we constructed plasmids encoding nine different histidine (H)-to-leucine (L) mutants for porcine DNase II and examined the enzyme properties of the expressed mutant proteins. Of the mutants, all but H132L were secreted into the medium of expressing cells. Six of the mutated DNase II proteins (H41L, H109L, H206L, H207L, H274L and H322L) showed enzyme activity, whereas the H115L, H132L and H297L mutants exhibited very little activity. The H115L and H297L mutants were found to undergo correct protein folding, but were inactive. To further examine these mutants, we expressed H115A and H297A DNase II mutants; these mutants were inactive, but their DNase activities could be rescued with imidazole, indicating that His115 and His297 are likely to function as a general acid and a general base respectively in the catalytic centre of the enzyme. In contrast with the secreted mutants, the H132L mutant protein was found in cell lysates within 16 h after transfection. This protein was inactive, improperly folded and was drastically degraded via the proteosomal pathway after 24 h. The polypeptide of another substitution for His132 with lysine resulted in the misfolded form being retained in endoplasmic reticulum.
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PMID:Identification of three crucial histidine residues (His115, His132 and His297) in porcine deoxyribonuclease II. 1673 90

Granzyme A (GzmA) activates a caspase-independent cell death pathway with morphological features of apoptosis. Single-stranded DNA damage is initiated when the endonuclease NM23-H1 becomes activated to nick DNA after granzyme A cleaves its inhibitor, SET. SET and NM23-H1 reside in an endoplasmic reticulum-associated complex (the SET complex) that translocates to the nucleus in response to superoxide generation by granzyme A. We now find the 3'-to-5' exonuclease TREX1, but not its close homolog TREX2, in the SET complex. TREX1 binds to SET and colocalizes and translocates with the SET complex. NM23-H1 and TREX1 work in concert to degrade DNA. Silencing NM23-H1 or TREX1 inhibits DNA damage and death of cells treated with perforin (PFN) and granzyme A, but not of cells treated with perforin and granzyme B (GzmB). After granzyme A activates NM23-H1 to make single-stranded nicks, TREX1 removes nucleotides from the nicked 3' end to reduce the possibility of repair by rejoining the nicked ends.
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PMID:The exonuclease TREX1 is in the SET complex and acts in concert with NM23-H1 to degrade DNA during granzyme A-mediated cell death. 1681 37

Accumulation of unfolded proteins in the endoplasmic reticulum triggers the unfolded protein response (UPR). How the UPR is downregulated is not well understood. Inositol requirement 1 (Ire1) is an endoplasmic reticulum transmembrane UPR sensor in Saccharomyces cerevisiae. When the UPR is triggered, Ire1 is autophosphorylated, on Ser 840 and Ser 841, inducing the cytosolic endonuclease activity of Ire1, thereby initiating the splicing and translational de-repression of HAC1 mRNA. Homologous to Atf/Creb1 (Hac1) activates UPR transcription. Here, we report that the dose-dependent cell-cycle regulator 2 (Dcr2) phosphatase functionally and physically interacts with Ire1. We identified genetic interactions between DCR2 and genes, including IRE1, which are involved in secretory processes. Overexpression of DCR2, but not of a catalytically inactive DCR2 allele, significantly delays HAC1 splicing and sensitizes cells to the UPR. Furthermore, Dcr2 physically interacts in vivo with Ire1-S840E,S841E, which mimics phosphorylated Ire1, and Dcr2 de-phosphorylates Ire1 in vitro. Our results are consistent with de-phosphorylation of Ire1 being a mechanism for antagonizing UPR signalling.
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PMID:Dcr2 targets Ire1 and downregulates the unfolded protein response in Saccharomyces cerevisiae. 1699 Aug 50

The Ire1p transmembrane receptor kinase/endonuclease transduces the unfolded protein response (UPR) from the endoplasmic reticulum (ER) to the nucleus in Saccharomyces cerevisiae. In this study, we analyzed the capacity of a highly basic sequence in the linker region of Ire1p to function as a nuclear localization sequence (NLS) both in vivo and in vitro. This 18-residue sequence is capable of targeting green fluorescent protein to the nucleus of yeast cells in a process requiring proteins involved in the Ran GTPase cycle that facilitates nuclear import. Mutagenic analysis and importin binding studies demonstrate that the Ire1p linker region contains overlapping potential NLSs: at least one classical NLS (within sequences 642KKKRKR647 and/or 653KKGR656) that is recognized by yeast importin alpha (Kap60p) and a novel betaNLS (646KRGSRGGKKGRK657) that is recognized by several yeast importin beta homologues. Kinetic binding data suggest that binding to importin beta proteins would predominate in vivo. The UPR, and in particular ER stress-induced HAC1 mRNA splicing, is inhibited by point mutations in the Ire1p NLS that inhibit nuclear localization and also requires functional RanGAP and Ran GEF proteins. The NLS-dependent nuclear localization of Ire1p would thus seem to be central to its role in UPR signaling.
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PMID:The unfolded protein response transducer Ire1p contains a nuclear localization sequence recognized by multiple beta importins. 1703 34

We have reported previously low expression of death receptors for tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) in fresh isolates and tissue sections of melanoma. This seemed to correlate with relative resistance of freshly isolated melanoma cells to TRAIL-induced apoptosis. We show in this study that the endoplasmic reticulum (ER) stress inducer, tunicamycin, selectively up-regulated the cell surface expression of TRAIL-R2, but not other members of the TNF receptor family, and enhanced TRAIL-induced apoptosis in cultured melanoma cells and fresh melanoma isolates. Tunicamycin-mediated sensitization of melanoma cells to TRAIL-induced apoptosis was associated with increased activation of the caspase cascade and reduction in mitochondrial membrane potential and was inhibited by a recombinant TRAIL-R2/Fc chimeric protein. Up-regulation of TRAIL-R2 on the melanoma cell surface was associated with increased transcription of TRAIL-R2 and its total protein levels. Two signaling pathways of the ER stress-induced unfolded protein response mediated by inositol-requiring transmembrane kinase and endonuclease 1alpha (IRE1alpha) and activation of transcription factor 6 (ATF6), respectively, seemed to be involved. In one melanoma line, there was clear evidence of activation of the IRE1alpha pathway, and small interfering RNA (siRNA) knockdown of IRE1alpha substantially reduced the up-regulation of TRAIL-R2. Similarly, there was evidence for the activation of the ATF6 pathway, and siRNA knockdown of ATF6 had a delayed effect on TRAIL-R2 expression in one but not another melanoma cell line. Moreover, the transcription factor CCAAT/enhancer-binding protein homologous protein seemed to be involved in the up-regulation of TRAIL-R2 by tunicamycin, but its role varied between different melanoma lines. Taken together, our results suggest that agents that induce ER stress may enhance TRAIL-R2 expression and increase the therapeutic response to TRAIL in melanoma.
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PMID:Tunicamycin sensitizes human melanoma cells to tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis by up-regulation of TRAIL-R2 via the unfolded protein response. 1757 57

We have previously reported that sensitivity of melanoma cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis is largely correlated with the levels of expression of TRAIL death receptors, in particular, TRAIL-R2 on the cell surface. However, fresh melanoma isolates and melanoma tissue sections express, in general, low levels of death receptors for TRAIL. We show in this study that the endoplasmic reticulum stress inducer, thapsigargin (TG), selectively up-regulated TRAIL-R2 and enhanced TRAIL-induced apoptosis in melanoma cells. However, the TRAIL-R2 pathway did not appear to be involved in induction of apoptosis by TG alone. Up-regulation of TRAIL-R2 appeared to be cooperatively mediated by the inositol-requiring transmembrane kinase and endonuclease 1alpha (IRE1alpha)- and activation of transcription factor (ATF)-6-signaling pathways of the unfolded protein response (UPR) and the transcription factor CCAAT/enhancer-binding protein-homologous protein (CHOP). The latter played a critical role in the initial phase of the increase in TRAIL-R2 as small interfering RNA (siRNA) knockdown of CHOP blocked up-regulation of TRAIL-R2 only at a relatively early stage (16 h) after exposure to TG. In contrast, IRE1alpha and ATF6 appeared to be crucial in maintaining the increased levels of TRAIL-R2 in that siRNA knockdown of IRE1alpha or ATF6 had no effect on the increase in TRAIL-R2 at the initial phase, but blocked TRAIL-R2 up-regulation at a relatively late stage (36 h). Our results indicate that modulation of the UPR may be useful in sensitizing melanoma cells to TRAIL-induced apoptosis by up-regulation of TRAIL-R2.
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PMID:Thapsigargin sensitizes human melanoma cells to TRAIL-induced apoptosis by up-regulation of TRAIL-R2 through the unfolded protein response. 1765 36


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