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)

Mutant Syrian hamster cells resistant to N-(phosphonacetyl)-L-aspartate (PALA), a transition state analog inhibitor of aspartate transcarbamylase, overproduce CAD, a multifunctional protein which catalyzes the first three reactions of de novo UMP biosynthesis. Increased levels of a single mRNA cause the overproduction of CAD in all PALA-resistant mutants examined thus far. A recombinant plasmid containing a 2,3-kilobase insert complementary to the 3'-proximal region of this 7.9-kilobase mRNA has been prepared and used to show that the CAD gene is amplified in each of the 10 PALA-resistant mutants examined. Rates of association of CAD sequences in DNA isolated from PALA-sensitive and PALA-resistant cells with labeled plasmid DNA indicated that the degree of amplification is approximately equal to the degree of overproduction of protein and mRNA in each mutant. The patterns of digestion of these DNAs with restriction enzymes confirmed this result and showed that the lower limit for the size of the amplified unit is 19 kilobases, much larger than the mRNA. A comparison of restriction endonuclease digests of the cloned cDNA with digests of genomic DNA indicated that part of this difference is attributable to intervening sequences in the CAD gene. A 10.2-kilobase RNA which contains CAD sequences is found in cytoplasmic fractions from some PALA-resistant mutants but not in wild type cells. Restriction patterns were analyzed by a new method in which fragments of DNA are transferred from agarose gels to diazo paper with a high efficiency which is independent of size.
 ...
PMID:Gene amplification causes overproduction of the first three enzymes of UMP synthesis in N-(phosphonacetyl)-L-aspartate-resistant hamster cells. 38 11

Fischer rat embryo fibroblasts subjected to temporary anoxia followed by an aerobic recovery period show genomic instability in the form of highly elevated CAD gene amplification rates. As revealed by flow cytometric analysis this is associated with DNA breakage in vivo, followed by repair during the recovery period. Such genomic instability parallels expression of a M(r) 29,000/31,000 endonuclease; this enzyme requires no added divalent metal ion and has a pH optimum of about 6.5. The same endonuclease was found to be expressed within healing wounds and in four of ten human colorectal cancers but was not seen in eight normal colorectal tissue samples. Our results indicate that DNA breakage resulting from endogenous endonuclease activity can have a substantial effect in modulating genomic instability.
 ...
PMID:An anoxia inducible endonuclease and enhanced DNA breakage as contributors to genomic instability in cancer. 786 98

Induction of apoptosis by death receptors such as Fas or tumour necrosis factor (TNF) R1 leads to distinct changes in cell morphology, activation of the caspase protease cascade, and the degradation of nuclear chromatin by activated nucleases. Here, we describe the purification and cDNA cloning of a novel 40 kDa endonuclease from Jurkat cells that is activated by caspases. This protein, designated caspase-activated nuclease (CPAN), is sufficient to degrade naked DNA and to induce apoptotic morphology and DNA fragmentation in naive nuclei. CPAN is highly homologous to a recently described mouse nuclease, CAD [1], and may represent the human homologue. Our data on the human cDNA as well as additional data on the mouse homologue suggest that a 30 amino-acid portion of the recently published mouse sequence [1] is incorrect. We show that the activity of human CPAN is regulated by DFF45 [2], an inhibitor necessary for CPAN expression and stabilization in an inactive state in living cells. Proteolytic cleavage of DFF45 by caspases in vitro leads to dissociation of DFF45 fragments from CPAN and activation of CPAN as an endonuclease. CPAN is a tightly regulated endonuclease with unique characteristics that might represent a distinctive family of endonucleases.
 ...
PMID:CPAN, a human nuclease regulated by the caspase-sensitive inhibitor DFF45. 956 Mar 46

Caspase-3 initiates apoptotic DNA fragmentation by proteolytically inactivating DFF45 (DNA fragmentation factor-45)/ICAD (inhibitor of caspase-activated DNase), which releases active DFF40/CAD (caspase-activated DNase), the inhibitor's associated endonuclease. Here, we examined whether other apoptotic proteinases initiated DNA fragmentation via DFF45/ICAD inactivation. In a cell-free assay, caspases-3, -6, -7, -8, and granzyme B initiated benzoyloxycarbonyl-Asp-Glu-Val-Asp (DEVD) cleaving caspase activity, DFF45/ICAD inactivation, and DNA fragmentation, but calpain and cathepsin D failed to initiate these events. Strikingly, only the DEVD cleaving caspases, caspase-3 and caspase-7, inactivated DFF45/ICAD and promoted DNA fragmentation in an in vitro DFF40/CAD assay, suggesting that granzyme B, caspase-6, and caspase-8 promote DFF45/ICAD inactivation and DNA fragmentation indirectly by activating caspase-3 and/or caspase-7. In vitro, however, caspase-3 inactivated DFF45/ICAD and promoted DNA fragmentation more effectively than caspase-7 and endogenous levels of caspase-7 failed to inactivate DFF45/ICAD in caspase-3 null MCF7 cells and extracts. Together, these data suggest that caspase-3 is the primary inactivator of DFF45/ICAD and therefore the primary activator of apoptotic DNA fragmentation.
 ...
PMID:Caspase-3 is the primary activator of apoptotic DNA fragmentation via DNA fragmentation factor-45/inhibitor of caspase-activated DNase inactivation. 1052 51

Several environmental and genetic factors are associated with high levels of cholesterol. Hypercholesterolemia is the main phenotype of Familial Defective Apolipoprotein B and Familial Hypercholesterolemia that are caused by mutations at the apolipoprotein (apo) B and LDL receptor genes, respectively. Identification of the specific genetic alteration associated with hypercholesterolemia is an important issue in clinical diagnosis of high risk for CAD. Apo B gene mutations and polymorphisms are usually screened by SSCP, DGGE, and heteroduplex, which must be confirmed by DNA sequencing or by direct detection using PCR techniques. In this study, we have optimized a PCR-RFLP procedure for identification of 3500Q and 3531 mutations and MspI polymorphism at the apo B gene. The technique can be performed in a single reaction, using the restriction endonuclease MspI for simultaneous detection of 3500Q mutation and MspI polymorphism, and NsiI for detection of 3531 mutation. The procedure was validated by analysis of control DNA samples from individuals carrying these mutations. Screening of 186 Brazilian hypercholesterolemic individuals showed that the frequency of the M-allele (7.8%) of MspI polymorphism was similar to that found in other individuals with CAD. However, neither 3500Q nor 3531 mutations were detected in this group. In conclusion, this procedure is simple and rapid, being easily introduced in clinical laboratories for direct detection of the more frequent mutations at the apo B gene associated with hypercholesterolemia.
 ...
PMID:Rapid detection of 3500Q and 3531 mutations and MspI polymorphism in exon 26 at the apolipoprotein B gene. 1117 Feb 32

The endonuclease DFF40/CAD mediates regulated DNA fragmentation and chromatin condensation in cells undergoing apoptosis. Here we report the enzyme's co-factor requirements, and demonstrate that the ionic changes that occur in apoptotic cells maximize DFF40/CAD activity. The nuclease requires Mg2+, exhibits a trace of activity in the presence of Mn2+, is not costimulated by Ca2+, is inhibited by Zn2+ or Cu2+, and has high activity over a rather broad pH range (7.0-8.5). The enzyme is thermally unstable, and is rapidly inactivated at 42 degrees C. Enzyme activity is markedly affected by ionic strength. At the optimal [K+] of 50-125 mM, which is in the range of the cytoplasmic [K+] for cells undergoing apoptosis, the activity of DFF40/CAD for naked DNA cleavage is about 100-fold higher than at 0 or 200 mM [K+]. Although these ranges of ionic strength do not affect DFF40 homo-oligomer formation, at higher ionic strengths the enzyme introduces single-stranded nicks into supercoiled DNA.
 ...
PMID:Ionic and cofactor requirements for the activity of the apoptotic endonuclease DFF40/CAD. 1133 Aug 26

We previously isolated a 34-kDa nuclease (AN34) from apoptotic human leukemia cells. Here, we identify AN34 as an N-terminally truncated form of human AP endonuclease (Ape1) lacking residues 1-35 (delta35-Ape1). Although Ape1 has hitherto been considered specific for damaged DNA (specific to AP site), recombinant AN34 (delta35-Ape1) possesses significant endonuclease activity on undamaged (normal) DNA and in chromatin. AN34 also displays enhanced 3'-5' exonuclease activity. Caspase-3 activates AN34 in a cell-free system, although caspase-3 cannot cleave Ape1 directly in vitro. We also found that Ape1 itself preferentially cleaves damaged chromatin DNA isolated from cells treated with apoptotic stimuli and that silencing of Ape1 expression decreases apoptotic DNA fragmentation in DFF40/CAD-deficient cells. Thus, we propose that AN34 and Ape1 participate in the process of chromatin fragmentation during apoptosis.
 ...
PMID:Human apurinic/apyrimidinic endonuclease (Ape1) and its N-terminal truncated form (AN34) are involved in DNA fragmentation during apoptosis. 1284 73

Curcumin is a natural pigment that has been shown to induce cell death in many cancer cells; however, the death mode depends on the cell type and curcumin concentration. Here we show that, in Jurkat cells, 50 micromol/L curcumin severely lowers cell survival and induces initial stage of chromatin condensation. It also induces caspase-3, which is sufficient to cleave DNA fragmentation factor 45 [DFF45/inhibitor of caspase-activated DNase (ICAD)], the inhibitor of DFF40/CAD endonuclease. However, the release of DFF40/CAD from its inhibitor does not lead to oligonucleosomal DNA degradation in curcumin-treated cells. Moreover, curcumin treatment protects cells from UVC-induced oligonucleosomal DNA degradation. In biochemical experiments using recombinant DFF activated with caspase-3, we show that curcumin inhibits plasmid DNA and chromatin degradation although it does not prevent activation of DFF40/CAD endonuclease after its release from the inhibitor. Using DNA-binding assay, we show that curcumin does not disrupt the DNA-DFF40/CAD interaction. Instead, molecular modeling indicates that the inhibitory effect of curcumin on DFF40/CAD activity results from curcumin binding to the active center of DFF40/CAD endonuclease.
 ...
PMID:Curcumin induces caspase-3-dependent apoptotic pathway but inhibits DNA fragmentation factor 40/caspase-activated DNase endonuclease in human Jurkat cells. 1664 63

We have previously reported that the pro-apoptotic pyrrolobenzoxazepine, PBOX-6, induces apoptosis in chronic myelogenous leukaemia (CML) cells which is accompanied by oligonucleosomal DNA fragmentation. In this study we show that PBOX-6-induced oligonucleosomal DNA fragmentation occurs in the absence of caspase and CAD activation in CML cells. Dissection of the signalling pathway has revealed that induction of apoptosis requires the upstream activation of a trypsin-like serine protease that promotes the phosphorylation and inactivation of anti-apoptotic Bcl-2. In addition, in this system chymotrypsin-like serine proteases are dispensable for high molecular weight DNA fragmentation, however are required for the activation of a relatively small manganese-dependent acidic endonuclease that is responsible for oligonucleosomal fragmentation of DNA. Furthermore, we demonstrate mitochondrial involvement during PBOX-6-induced apoptosis and suggest the existence of unidentified mitochondrial effectors of apoptosis.
 ...
PMID:Caspase-activated DNase (CAD)-independent oligonucleosomal DNA fragmentation in chronic myeloid leukaemia cells; a requirement for serine protease and Mn2+-dependent acidic endonuclease activity. 1682 Sep 64

Apoptosis is characterized by cell shrinkage, nuclear condensation and internucleosomal DNA cleavage. Besides the central role of caspases and other proteases, cell death triggers DNA degradation so that DNases have an active role in apoptotic cell death. The best-characterized apoptotic DNase is CAD, a neutral Mg-dependent endonuclease. Its activity is regulated by its inhibitor, ICAD, which is cleaved by caspases. Other neutral DNases have been shown to cleave nuclear DNA in apoptotic conditions: endonuclease G, GADD. In cells, the cytosolic pH is maintained to 7.2, mostly due to the activity of the Na(+)/H(+) exchanger. In many apoptotic conditions, a decrease of the intracellular pH has been shown. This decrease may activate different acid DNases, mostly when pH decreases below 6.5. Three acidic DNases II are so far known: DNase II alpha, DNase II beta and L-DNase II, a DNase II, derived from the serpin LEI (Leukocyte Elastase Inhibitor). Their activation during cell death is discussed in this review.
 ...
PMID:Acid DNases and their interest among apoptotic endonucleases. 1698 34


1 2 Next >>