Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cytotoxic T lymphocytes (CTL) kill their target cells via a contact-dependent mechanism that results in the perturbation of the target cell's plasma membrane and the fragmentation of the target cell's DNA into nucleosomal particles. The membrane disruption is presumed to be due to the action of perforin, while the DNA fragmentation is thought to be by the activation of an endogenous nuclease(s). DNA topoisomerases I and II are nuclear enzymes with inherent endonuclease activities. We have investigated their role in the CTL-induced DNA fragmentation process. We report that in CTL killing assays, the treatment of target cells with topoisomerase I and II inhibitors blocks the CTL-induced DNA fragmentation process, but not the lysis of the target cell.
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PMID:Inhibition of cytotoxic T lymphocyte-induced target cell DNA fragmentation, but not lysis, by inhibitors of DNA topoisomerases I and II. 130 53

When activated with either Con A, a CD3-specific mAb, or Ag-pulsed B lymphoma (LK35.2) cells, CD4 (Th1) clones quickly induce DNA fragmentation in target cells followed by release of 51Cr-labeled intracellular materials. Both activated CD4 clones and CD8 (CTL) cells fragment target DNA into electrophoretically identical "ladder" pattern made of approximately 200 bp. The effect of various metabolic inhibitors on the ability of CD4 and CD8 cells to induce target DNA fragmentation was studied. Little effect was observed with the DNA synthesis inhibitor, mitomycin C. The RNA synthesis inhibitor, actinomycin D, and the protein synthesis inhibitor, cycloheximide, strongly inhibited the ability of CD4 cells, but not CD8 cells, to induce target DNA fragmentation. In contrast, target DNA fragmentation by CD8 cells, but not by CD4 cells, was inhibited by cholera toxin. Although cyclosporin A inhibited CD4 cells to fragment target DNA during the early phase (90 min) of E:T interaction, this inhibition was not sustained in the later phase (210 min) of the assay. Zinc ions inhibited the ability of both CD4 and CD8 cells to fragment target DNA. Treatment of effectors and targets with these inhibitors, followed by washings, demonstrated that the action of these inhibitors on effector cells alone is sufficient to inhibit target DNA fragmentation. The strong correlation among these parameters of DNA fragmentation and Cr-release assays supports the hypothesis of programed cell death. Although distinct cytolytic pathways are used by CD4 and CD8 cells to kill targets, both pathways deliver a signal that activates endonuclease(s), fragments target DNA, causes Cr-release, and lyses target cells. Taken together with our previous studies, the present findings demonstrate that activated cytolytic CD4 clones do not use perforin, serine proteases, and TNF as mediators for resistant target DNA fragmentation.
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PMID:Distinct pathways of CD4 and CD8 cells induce rapid target DNA fragmentation. 167 Oct 51

Extracellular ATP is shown here to induce programmed cell death (or apoptosis) in thymocytes and certain tumor cell lines. EM studies indicate that the ATP-induced death of thymocytes and susceptible tumor cells follows morphological changes usually associated with glucocorticoid-induced apoptosis of thymocytes. These changes include condensation of chromatin, blebbing of the cell surface, and breakdown of the nucleus. Cytotoxicity assays using double-labeled cells show that ATP-mediated cell lysis is accompanied by fragmentation of the target cell DNA. DNA fragmentation can be set off by ATP but not the nonhydrolysable analogue ATP gamma S nor other nucleoside-5'-triphosphates. ATP-induced DNA fragmentation but not ATP-induced 51Cr release can be blocked in cells pretreated with inhibitors of protein or RNA synthesis or the endonuclease inhibitor, zinc; whereas pretreatment with calmidazolium, a potent calmodulin antagonist, blocks both DNA fragmentation and 51Cr release. The biochemical and morphological changes caused by ATP are preceded by a rapid increase in the cytoplasmic calcium of the susceptible cell. Calcium fluxes by themselves, however, are not sufficient to cause apoptosis, as the pore-forming protein, perforin, causes cell lysis without DNA fragmentation or the morphological changes associated with apoptosis. Taken together, these results indicate that ATP can cause cell death through two independent mechanisms, one of which, requiring an active participation on the part of the cell, takes place through apoptosis.
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PMID:Extracellular ATP as a trigger for apoptosis or programmed cell death. 198 62

We have partially characterized the granules of the human NK cell line, YT-INDY, and assessed granule-mediated lysis and DNA fragmentation of assorted targets. Biochemical studies demonstrated significant quantities of granzyme B (asp-ase) and a heretofore undescribed chymase but no tryptase (i.e., granzyme A or 3) or distinct met-ase. YT-INDY expressed mRNA for granzyme B, perforin and CCPX. The existence of perforin was confirmed by immunoblot. The granules lysed both human and murine NK-sensitive and NK-resistant targets. YT-INDY and NK3.3, two human cytotoxic cells, were also lysed. EGTA reduced lysis by only 50%, suggesting that a perforin-independent lytic pathway is associated with the granules. In addition, 4-(2-aminoethyl) benzenesulfonylfluoride hydrochloride (AEBSF), an inhibitor that selectively blocked the chymase and 3,4-dichloroisocoumarin (DCI), an inhibitor that inactivated both chymase and asp-ase activities, marginally affected lysis. By gel electrophoresis and 125I-labeled deoxyuridine release assay, only murine cells (SP2/0 and YAC-1) underwent DNA fragmentation, and cleavage was completely inhibited by DCI, whereas EGTA, AEBSF and aurintricarboxylic acid (ATA) had no effect. The results, therefore, underscore the central role of granzyme B in granule-mediated DNA fragmentation, emphasize that the protease acts via an ATA-resistant endonuclease pathway and stress that nucleolysis does not invariably accompany granule-mediated cytolysis. Finally, ATA inhibited the asp-ase activity of isolated but not granule-associated granzyme B. ATA, therefore, is not a specific endonuclease inhibitor and results obtained with ATA should be viewed cautiously.
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PMID:Human granzyme B is essential for DNA fragmentation of susceptible target cells. 808 28

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

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