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PubMed1996

The effect of complete excision on stage III neuroblastoma: a report of the European Neuroblastoma Study Group.

UNLABELLED: Current opinion is divided about the value of excisional surgery in Evans stage III neuroblastoma. AIMS: To evaluate and correlate the survival of patients with stage III neuroblastoma with the effectiveness of the surgical excision, as assessed by (1) the surgeon (resection data) at the time of operation and (2) the pathologist (excision data). METHODS: The ENSG (European Neuroblastoma Study Group) database of 202 patients from 29 centres with proven stage III were analysed. The data include all patients with neuroblastoma diagnosed between 1982 and 1992 and their subsequent follow-up. RESULTS: Patients were grouped according to the extent of resection (100%, 75% to 99%, and < 75%) and the completeness of excision (complete, microscopic residual, macroscopic residual). There were 123 with resection data, a subgroup of 104 with excision data, and 27 with no excision. There was no statistically significant difference (log rank test) in overall survival (p = 0.11) or event-free survival between the resection subgroups, even when the data from patients without resection were included. Complete excision was associated with a highly significant survival advantage, in terms of overall survival (P = .007) and event-free survival (P = .006). This effect is most obvious among patients with the worst prognosis: older children and those with an abdominal tumour. CONCLUSION: Histological confirmation of complete excision confers a significant survival advantage for patients with stage III neuroblastoma and justifies a painstaking attempt at complete resection.

Adolescent↗

PubMed1998

High and low UV-dose responses in SOS-induction of the precise excision of transposons tn1, Tn5 and Tn10 in Escherichia coli.

UV-inducible precise excision of transposons is a specific SOS-mutagenesis process. It deals with the deletion formation which has previously been demonstrated to involve direct or inverted IS-sequences of transposons. The process was used for revisiting the targeted and untargeted SOS-mutability and its relationship to the key genes for SOS-mutagenesis: the recA, lexA and umuDC. The precise excision of transposons Tn5 and Tn10 from the chromosomal insertion sites ade128 and cyc750 is induced in Escherichia coli K-12 and B cells, wild-type for DNA-repair, both by the low doses of UV-light ranging from 0.25 J m-2 to 2.5 J m-2 and the high doses within the range 5.0-40.0 J m-2. Precise excision of these transposons induced by the range of low doses incapable to induce targeted point mutations reveals its mostly untargeted nature. This process for the transposon Tn1 is not induced by UV-light within the range of doses 0.25-2.5 J m-2 while its induction is possible by UV-fluences ranging from 5.0 to 40.0 J m-2. A dose-response of the precise excision of Tn1 is similar to that of the UV-induced reversion of trpUAA point mutation that is targeted by nature and contrasts to the UV-inducible precise excision of Tn5 and Tn10. Both types of UV-inducible precise excision, demonstrated either by Tn1 or Tn5 and Tn10, are eliminated by mutations in the lexA, recA and umuDC genes indispensable for UV-induced SOS-mutability. The palindromic structures different for the transposons Tn1, Tn5 and Tn10 are discussed to be involved and affect the targeted and untargeted precise excision of transposons induced by UV-light.

Chromosomes, Bacterial↗

PubMed1997

Persistent intraepithelial neoplasia after excision for cervical intraepithelial neoplasia grade III.

OBJECTIVE: To determine the factors associated with subsequent intraepithelial neoplasia among patients who had cervical intraepithelial neoplasia grade III (CIN III) diagnosed on a specimen from previous large-loop excision of the transformation zone. METHODS: We studied all large-loop excisions of the transformation zone performed between May 1991 and December 1993, inclusive. All cases of CIN III were identified. We followed up patients with cytology and colposcopy for 2 years after treatment for high-grade CIN. Findings at follow-up were analyzed. RESULTS: A histologic diagnosis of CIN III was made in 225 patients. The lesion appeared incompletely excised in 105 patients (48.2%). In 76 cases (34.9%), CIN III was found at the endocervical margin. Of the 211 patients reviewed at 6 months, 18 (8.5%) had histologically proven CIN. The incidence of CIN was 16.5% after a report of incomplete excision, compared with 1.9% after a report of complete excision (P < .001). Furthermore, there was no subsequent CIN III in the complete-excision group. Of the 183 patients reviewed at 24 months, seven (3.8%) had histologically proven CIN. All cases of subsequent CIN were associated with dyskaryosis on follow-up cervical cytology. CONCLUSIONS: Positive margins increase the risk of treatment failure. Cytology alone may be adequate for follow-up when CIN III is completely excised.

Adult↗

PubMed1995

Early, complete burn wound excision partially restores cytotoxic T lymphocyte function.

BACKGROUND: Cytotoxic lymphocytes (CTLs) are an important component of immune function, involved in antigen recognition and resistance to viral infection. Burn injury suppresses cell-mediated immunity, induces allograft tolerance, and increases the risk of viral infection, but the mechanisms are not well understood. This study analyzes the effect of burn size and burn wound excision on CTL activity. METHODS: Anesthetized CBA mice (n = 12) received a 0%, 20%, or 40% body surface area contact burn. Additional mice (n = 16) received a 40% burn that was totally, partially, or not excised 72 hours after burn. Excised areas were covered with normal, syngeneic skin. Two weeks later harvested splenocytes were cocultured with allogeneic stimulators. CTL activity was determined by a 51Cr release assay, in which CTL effectors were tested on allogeneic, radiolabeled targets. Dilution curves of CTL activity were compared by ANOVA: RESULTS: Both 20% and 40% burns significantly inhibited CTL activity (p < 0.05). Total but not partial excision of a 40% burn restored CTL activity (p < 0.01). Both total and partial wound excision also improved survival (p < 0.05). CONCLUSIONS: Burn injury inhibits CTL activity in a size-dependent manner, and total wound excision significantly improves both CTL function and survival after injury. This study suggests a mechanism for the immunosuppressive effects of burn injury and provides an immunologic rationale for early, complete burn wound excision.

Animals↗

PubMed1987

Early burn excision and grafting.

The current state of knowledge and experience with early burn wound closure leads to some conclusions that are proved and others that are suggested. The issues that are proved include the following: Small (less than 20 per cent TBSA) full-thickness burns and indeterminate (deep partial-thickness versus full-thickness) burns, if treated by an experienced surgeon, can be excised safely and grafted with a decrease in hospital stay, cost to the patient, and time away from work or school. Early excision and grafting dramatically decreases the number of painful ward debridements required by all patients. Patients with burns between 20 and 40 per cent TBSA appear to have fewer infectious wound complications and a shortened hospital stay if treated with early excision and grafting. In addition to the above conclusions, about which there is little disagreement, there are other suggestive data and clinical impressions that do not yet have "hard data" to support them. These issues include the following: Scarring is less in wounds closed early, leading to better appearance and the need for fewer reconstructive procedures. There presently is not a good measure of "cosmetic appearance," and comparative studies await an acceptable scale to measure results. Mortality from wound infection is less in patients with major burns. Because wounds exceeding the donor sites cannot be closed permanently and completely until donor sites can be reharvested, proof will come only when a durable permanent cover can be applied in a timely fashion. Data now suggest that mortality has decreased, but none of the studies has been conclusive. Mortality from other complications of major burns may decrease with early excision and grafting. Decreasing stress, hypermetabolism, and decreasing the overall bacterial load of such patients enables them to resist other complications better. The only present data supporting this conclusion, however, come from animal studies. The current state of the art suggests that small, deep burns can be excised and grafted by general or plastic surgeons in community hospitals as long as they are interested in the procedure and the hospital has the proper support facilities. Burns of cosmetically or functionally important areas (face, hands, feet, and joints) should be excised by someone with a special interest in burns, and burns in excess of 10 per cent TBSA should be excised only in facilities with excellent support facilities, including an experienced anesthesiologist, trained nursing personnel, good critical-care facilities, and a safe blood bank.

Burns↗

PubMed2003

Mapped serial excision for periocular lentigo maligna and lentigo maligna melanoma.

PURPOSE: To report the early cure rate for periocular lentigo maligna (LM) and LM melanoma (LMM), using modified Mohs surgery with vertically cut paraffin-embedded sections (mapped serial excision [MSE]). A secondary aim was to identify differences in the clinical features and outcomes between periocular LM and LMM and those found elsewhere on the head and neck. DESIGN: Prospective, noncomparative, interventional case series. PARTICIPANTS: One hundred thirty-five patients undergoing 141 MSE procedures. METHODS: A prospective series of 141 MSE procedures for LM and LMM over a 10-year period (1993-2002) in a single-center Mohs surgical unit. MAIN OUTCOME MEASURES: Recurrence, site, size of LM or LMM, invasiveness, prior recurrence, clear margin of excision, size of final defect, and number of levels required for complete excision. RESULTS: One hundred forty-one MSE procedures, of which 23% (32/141) were for LMM and 19% (27/141) were for periocular lesions. Location or prior recurrence were not predictive of invasive disease; however, the size distribution of the initial lesion (P = 0.0354) and the final defect after MSE (P = 0.0183) were larger in LMM. Thirty-one percent of LM and 14% of LMM less than 1 mm thick required larger than 5-mm and 1-cm margins, respectively, for complete excision. Mean follow-up of 32 months (range, 1-100 months) revealed 4 recurrences (3%), of which two were periocular (P = 0.188). CONCLUSIONS: Our review is the largest prospective series of MSE for LM and LMM and suggests that it is the treatment of choice in these forms of melanoma. Mapped serial excision offers a high early cure rate in conjunction with tissue conservation, which is of particular relevance in the periocular region. There were no significant differences between periocular LM and LMM and those found elsewhere in the head and neck region. It also appears that the current recommendations of 5-mm margins for in situ melanoma (LM) and 1-cm margins for melanoma less than 1 mm thick are insufficient for complete excision of LM or LMM, emphasizing the importance of margin-controlled excision of these lesions.

Adult↗

PubMed1997

Major suppression of pro-alpha1(I) type I collagen gene expression in the dermis after keloid excision and immediate intrawound injection of triamcinolone acetonide.

BACKGROUND: A keloid is a benign tumor that contains excess collagen, primarily type I collagen. A common therapy is intralesional injection of a glucocorticosteroid, such as triamcinolone acetonide (TA). Surgical excision is also common; often a glucocorticosteroid is injected weeks after excision when wound repair has already begun. OBJECTIVE: Our purpose was to determine the efficacy of TA in reducing the pro-alpha1(I) type I collagen mRNA in the dermis, when TA is injected into the wound bed immediately after surgical excision of the keloid. METHODS: Six patients with previously untreated keloids were studied. Three were treated with 10 mg/ml of TA immediately after excision of the keloid (experimental group); the other three patients were not treated with TA until 2 weeks after excision (control). Punch biopsy specimens were obtained from the TA-treated sites 2 weeks after removal of the keloid and from the wounds of the control group of patients before they were treated with TA. Sections were prepared for in situ hybridization analysis of the pro-alpha1(I) collagen mRNA, as well as for histochemical analysis of collagen fibers. RESULTS: All keloids showed greatly elevated levels of pro-alpha1(I) type I collagen mRNA in the dermis. Postsurgical wounds injected with TA after removal of the keloid expressed decreased pro-alpha1(I) collagen transcripts, compared with skin not treated with TA. The collagen bundles were also thinner and less dense in the TA-treated skin. CONCLUSION: Downregulation of the type I collagen gene expression is elicited by immediate TA injection after keloid excision. This suggests that prevention of recurrent keloid growth is possible if surgical excision is accompanied by immediate TA injection into the wound bed and that healing of the wound is not apparently compromised by inhibition of type I collagen gene expression.

Administration, Topical↗

PubMed1998

Excision of the tol2 transposable element of the medaka fish, Oryzias latipes, in zebrafish, Danio rerio.

The Tol2 element is a transposable element in Oryzias latipes (the medaka fish) found in the tyrosinase gene locus of the tyrosinase-deficient mutant medaka fish and has been shown to be excised from the genome during medaka embryogenesis (Koga, A., Suzuki, M., Inagaki, H., Bessho, Y., Hori, H., 1996. Transposon element in fish. Nature 383, 30). It is, however, not known whether the Tol2 element is an autonomous element. To determine whether the cloned Tol2 element is an autonomous element and whether excision can occur also in the other fish species, the plasmid DNA harboring the Tol2 element was injected to fertilized eggs of zebrafish, Danio rerio, and the total DNA extracted from the embryos 9-10h after the injection was analyzed by PCR. When a plasmid with the full-length Tol2 element was used for the microinjection, in 39 out of 43 injected embryos, we found generation of short PCR products indicative of the loss of the Tol2 element from the injected plasmid. Ten of these cases were analyzed at the DNA sequence level, and nine of them showed either precise excision of the Tol2 element (three cases) or nearly precise excision of the element with the addition of a few nucleotides of the target duplication (six cases). When a deletion version of the Tol2 element that retained the terminal inverted repeats but lacked about one-fourth of the open reading frame-coding region was used for the microinjection, such short PCR products could not be amplified from any of the injected embryos (0 out of 30). Thus, the Tol2 element is capable of excision in zebrafish embryos, presumably dependent on a putative transposase encoded by the Tol2 element itself. This transient embryonic excision assay using zebrafish should be useful to analyze the structure and the function of the transposase and cis-elements necessary for excision. Also, this study implies the potential use of the Tol2 element in transgenesis and insertional mutagenesis in both zebrafish and the medaka fish.

Animals↗

PubMed1999

Identification of the Tol2 transposase of the medaka fish Oryzias latipes that catalyzes excision of a nonautonomous Tol2 element in zebrafish Danio rerio.

The Tol2 element is found in the genome of the medaka fish, Oryzias latipes, and contains DNA sequences similar to those of transposons of the hAT family. Previously, we have developed a transient embryonic excision assay in zebrafish, in which zebrafish embryos were injected with a plasmid DNA harboring the Tol2 element, and have shown that the Tol2 element is excisable from the injected plasmid DNA (Kawakami, K., Koga, A., Hori, H., Shima, A., 1998. Excision of the Tol2 transposable element of the medaka fish, Oryzias latipes, in zebrafish, Danio rerio. Gene 225, 17-22). Although the Tol2 element is thought to be autonomous, an active transposase encoded by the Tol2 element has not been identified. Here we report the identification and analysis of mRNA transcribed from the Tol2 element in zebrafish embryos. The Tol2 transcript has the capacity to encode a protein of 649 amino acids, whose amino acid sequence is similar to those of transposases of the hAT family. To determine whether the transcript encodes an active enzyme, we developed a novel transient embryonic excision assay in which zebrafish fertilized eggs were co-injected with RNA transcribed in vitro using the Tol2 cDNA as a template and a plasmid DNA harboring a nonautonomous Tol2 element, which has a deletion in the transposase coding region. The nonautonomous Tol2 element could be efficiently excised in the zebrafish only when co-injected with the Tol2 RNA. This result indicates that the Tol2 transcript encodes an active enzyme, a probable transposase, that can catalyze the excision reaction in trans. Further, by the co-injection analysis, we found that the Tol2 sequence lacking the first intron sequence of the transposase gene could not be excised, suggesting that it may contain essential cis-elements.

Amino Acid Sequence↗

PubMed1992

Substrate specificity of the Escherichia coli Fpg protein (formamidopyrimidine-DNA glycosylase): excision of purine lesions in DNA produced by ionizing radiation or photosensitization.

We have investigated the excision of a variety of modified bases from DNA by the Escherichia coli Fpg protein (formamidopyrimidine-DNA glycosylase) [Boiteux, S., O'Connor, T. R., Lederer, F., Gouyette, A., & Laval, J. (1990) J. Biol. Chem. 265, 3916-3922]. DNA used as a substrate was modified either by exposure to ionizing radiation or by photosensitization using visible light in the presence of methylene blue (MB). The technique of gas chromatography/mass spectrometry, which can unambiguously identify and quantitate pyrimidine- and purine-derived lesions in DNA, was used for analysis of hydrolyzed and derivatized DNA samples. Thirteen products resulting from pyrimidines and purines were detected in gamma-irradiated DNA, whereas only the formation of 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyGua) and 8-hydroxyguanine (8-OH-Gua) was observed in visible light/MB-treated DNA. Analysis of gamma-irradiated DNA after incubation with the Fpg protein followed by precipitation revealed that the Fpg protein significantly excised 4,6-diamino-5-formamidopyrimidine (FapyAde), FapyGua, and 8-OH-Gua. The excision of a small but detectable amount of 8-hydroxyadenine was also observed. The detection of these products in the supernatant fractions of the same samples confirmed their excision by the enzyme. Nine pyrimidine-derived lesions were not excised. The Fpg protein also excised FapyGua and 8-OH-Gua from visible light/MB-treated DNA. The presence of these products in the supernatant fractions confirmed their excision.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed2001

Evidence in Escherichia coli that N3-methyladenine lesions induced by a minor groove binding methyl sulfonate ester can be processed by both base and nucleotide excision repair.

It has been previously reported that a neutral DNA equilibrium binding agent based on an N-methylpyrrolecarboxamide dipeptide (lex) and modified with an O-methyl sulfonate ester functionality (MeOSO(2)-lex) selectively affords N3-methyladenine lesions. To study the interaction of the neutral lex dipeptide with calf thymus DNA, we have prepared stable, nonmethylating sulfone analogues of MeOSO(2)-lex that are neutral and cationic. Thermodynamic studies show that both the neutral and monocationic sulfone compounds bind to DNA with K(b)'s of 10(5) in primarily entropy-driven reactions. To determine how the cytotoxic N3-methyladenine adduct generated from MeOSO(2)-lex is repaired in E. coli, MeOSO(2)-lex was tested for toxicity in wild-type E. coli and in mutant strains defective in base excision repair (tag and/or alkA glycosylases or apn endonuclease), nucleotide excision repair (uvrA), and both base and nucleotide excision repair (tag/alkA/uvrA). The results clearly demonstrate the cellular toxicity of the N3-methyladenine lesion, and the protective role of base excision glycosylase proteins. A novel finding is that in the absence of functional base excision glycosylases, nucleotide excision repair can also protect cells from this cytotoxic minor groove lesion. Interaction between base and nucleotide excision repair systems is also seen in the protection of cells treated with cis-diamminedichloroplatinum(II) but not with anti-(+/-)-r-7,t-8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗

PubMed1986

Photoreversal-dependent release of thymidine and thymidine monophosphate from pyrimidine dimer-containing DNA excision fragments isolated from ultraviolet-damaged human fibroblasts.

To elucidate the enzymatic excision-repair process operative on cyclobutane-type pyrimidine photodimers in human dermal fibroblasts, we have examined excised dimer-containing material recovered in the trichloroacetic acid soluble fraction from far-ultraviolet-irradiated (254 nm, 40 J m-2) and incubated (24 h) cell cultures. The excised DNA photoproducts were found in oligonucleotide fragments with an estimated mean chain length of approximately 3.7 bases. Exposure of these isolated excision fragments, labeled with [3H]thymidine (dT), to a secondary, dimer-photoreversing fluence of far-UV (5.5 kJ m-2) resulted in the release of free dT and thymidine monophosphate (TMP). Photorelease of these two radioactive species was measured by high-performance liquid chromatography, with TMP being detected as the increase in dT following bacterial alkaline phosphatase treatment. These data imply that the photoliberated dT and TMP moieties were attached to the excision fragments solely by the cyclobutane ring of the dimer. No evidence was obtained for the photoliberation of free thymine, thus corroborating a conclusion reached by others that the excision of dimers in human cells is not initiated by scission of an intradimer N-glycosyl bond. The sum of the tritium label recovered in dT plus TMP corresponded to approximately 40% of that disappearing from thymine-containing dimers on photoreversal, suggesting that in about 80% of the isolated excision fragments the dimer is located at one end of the oligonucleotide and contains a break in its internal phosphodiester bond.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed2003

Arabidopsis thaliana Ogg1 protein excises 8-hydroxyguanine and 2,6-diamino-4-hydroxy-5-formamidopyrimidine from oxidatively damaged DNA containing multiple lesions.

A functional homologue of eukaryotic Ogg1 proteins in the model plant Arabidopsis thalianahas recently been cloned, isolated, and characterized [Garcia-Ortiz, M. V., Ariza, R. R., and Roldan-Arjona, T. (2001) Plant Mol. Biol. 47, 795-804]. This enzyme (AtOgg1) exhibits a high degree of sequence similarity in several highly conserved regions with Saccharomyces cerevisiae, Drosophila melanogaster, and human Ogg1 proteins. We investigated the substrate specificity and kinetics of AtOgg1 for excision of modified bases from oxidatively damaged DNA that contained multiple pyrimidine- and purine-derived lesions. Two different DNA substrates prepared by exposure to ionizing radiation in aqueous solution under N2O or air were used for this purpose. Gas chromatography/isotope-dilution mass spectrometry was applied to identify and quantify modified bases in DNA samples. Of the 17 modified bases identified in DNA samples, only 8-hydroxyguanine and 2,6-diamino-4-hydroxy-5-formamidopyrimidine were significantly excised from both DNA substrates. This is in agreement with the substrate specificities of other eukaryotic Ogg1 proteins that had previously been studied under identical conditions. Excision depended on incubation time, enzyme concentration, and substrate concentration and followed Michaelis-Menten kinetics. A significant dependence of excision on the nature of DNA substrate was observed in accord with previous studies on other DNA glycosylases. A comparison of excision kinetics pointed to significant differences between AtOgg1 and other Ogg1 proteins. We also investigated the effect of base-pairing on the excision using double-stranded oligodeoxynucleotides that contained 8-OH-Gua paired with each of the four DNA bases. The activity of AtOgg1 was most effective on the 8-OH-Gua:C pair with some or very low activity on other pairs in agreement with the activity of other Ogg1 proteins. The results unequivocally show that AtOgg1 possesses common substrates with other eukaryotic Ogg1 proteins albeit significant differences between their excision kinetics.

Arabidopsis Proteins↗

PubMed1996

Repair of cisplatin--DNA adducts by the mammalian excision nuclease.

Nucleotide excision repair is one of the many cellular defense mechanisms against the toxic effects of cisplatin. An in vitro excision repair assay employing mammalian cell-free extracts was used to determine that the 1,2-d(ApG) intrastrand cross-link, a prevalent cisplatin-DNA adduct, is excised by the excinuclease from a site-specifically modified oligonucleotide 156 base pairs in length. Repair of the minor interstrand d(G)/d(G) cross-link was not detected by using this system. Proteins containing the high mobility group (HMG) domain DNA-binding motif, in particular, rat HMG1 and a murine testis-specific HMG-domain protein, specifically inhibit excision repair of the intrastrand 1,2-d(GpG) and -d(ApG) cross-links. This effect was also exhibited by a single HMG domain from HMG1. Similar inhibition of repair of a site-specific 1,2-d(GpG) intrastrand cross-link by an HMG-domain protein also occurred in a reconstituted system containing highly purified repair factors. These results indicate that HMG-domain proteins can block excision repair of the major cisplatin-DNA adducts and suggest that such an activity could contribute to the unique sensitivity of certain tumors to the drug. The reconstituted excinuclease was more efficient at excising the 1,3-d(GpTpG) intrastrand adduct than either the 1,2-d(GpG) or d(ApG) intrastrand adducts, in agreement with previous experiments using whole cell extracts [Huang, J. -C., Zamble, D. B., Reardon, J. T., Lippard, S. J., Sancar, A. (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 10394-10398]. This result suggests that structural differences among the platinated DNA substrates, and not the presence of unidentified cellular factors, determine the relative excision repair rates of cisplatin-DNA intrastrand cross-links in the whole cell extracts.

Animals↗

PubMed1998

Catalytic center of DNA polymerase beta for excision of deoxyribose phosphate groups.

The amino-terminal 8-kDa domain of vertebrate DNA polymerase beta (pol beta) has an activity to excise deoxyribose phosphate (dRP) groups from 5'-incised apurinic/apyrimidinic (AP) sites during base excision repair. The excision reaction proceeds via a beta-elimination reaction following formation of a Schiff base between an aldehyde group of the AP site and an amino group of the enzyme. Here we report that the Lys-72 residue of this enzyme is the catalytic center for dRP excision. Substitutions of Lys-72 with Arg or Gln reduced the dRP excision activity to less than 1% of the wild-type 8-kDa domain, while substitutions of Lys-35, Lys-68, or Lys-84 did not abolish its activity. The Lys-72 mutations also significantly decreased Schiff base intermediates trapped by reduction with sodium borohydride. The 8-kDa domain alone was able to bind preferentially to a single-nucleotide gap or 5'-incised synthetic AP site on double-stranded DNA. The Lys-72 mutations did not affect this damage-specific DNA binding activity. When introduced into the intact enzyme, a mutation of Lys-72 to Arg did not affect DNA synthesis activity of pol beta, but eliminated the repair activity. Addition of the wild-type 8-kDa domain to this reaction restored the repair activity. These results indicate a specific role of Lys-72 of pol beta in the dRP excision during base excision repair.

Amino Acid Sequence↗

PubMed2004

Signaling from DNA mispairs to mismatch-repair excision sites despite intervening blockades.

Mismatch-repair (MMR) systems promote genomic stability by correction of DNA replication errors. Thus, MMR proteins--prokaryotic MutS and MutL homodimers or their MutSalpha and MutLalpha heterodimer homologs, plus accessory proteins--specifically couple mismatch recognition to nascent-DNA excision. In vivo excision-initiation signals--specific nicks in some prokaryotes, perhaps growing 3' ends or Okazaki-fragment 5' ends in eukaryotes--are efficiently mimicked in vitro by nicks or gaps in exogenous DNA substrates. In some models for recognition-excision coupling, MutSalpha bound to mismatches is induced by ATP hydrolysis, or simply by binding of ATP, to slide along DNA to excision-initiation sites, perhaps in association with MutLalpha and accessory proteins. In other models, MutSalpha.MutLalpha complexes remain fixed at mismatches and contact distant excision sites by DNA looping. To challenge the hypothesis that recognition complexes remain fixed, we placed biotin-streptavidin blockades between mismatches and pre-existing nicks. In human nuclear extracts, mismatch efficiently provoked the initiation of excision despite the intervening barriers, as predicted. However, excision progress and therefore mismatch correction were prevented.

Adenosine Triphosphatases↗

PubMed1997

Ds excision from extrachromosomal geminivirus vector DNA is coupled to vector DNA replication in maize.

Analysis of transposition products generated after Activator (Ac) excision from the P locus in maize suggest that Ac excises either during or after replication of the P locus. The frequency of excision of the non-autonomous Ac derivative, Dissociation (Ds), from extrachromosomal replicating and nonreplicating vector DNAs in transfected black mexican sweet maize protoplasts was compared to assess directly a role of extrachromosomal vector DNA replication in Ds excision. Replicating (rep+) and nonreplicating (rep-) vector DNAs comprised a Ds element that harbored a geminivirus, wheat dwarf virus (WDV), origin of replication and WDV genes required for viral DNA replication (rep+) or mutant, inactive derivatives of these genes (rep-). Excision of Ds was detected only in those cell nuclei co-transfected with the replicating Ds-vector DNA and a transposase expression vector. Quantitative reconstruction experiments showed that Ds excised at least 3 x 10(5)-fold more frequently from replicating vector DNA as compared with nonreplicating vector DNA. Therefore, these results provide direct evidence for a coupling of Ds excision from extrachromosomal vector DNA to vector DNA replication in maize.

Cell Nucleus↗

PubMed1999

The pathogenesis of local recurrence of melanoma at the primary excision site.

Local recurrence of melanoma at the primary excision site may imply that the primary excision was incomplete or 'inadequate', and that the recurrence was due to retained primary melanoma cells or occult microsatellites in the adjacent tissue. Pathologists frequently report these tumours in the scar as recurrent or residual melanoma, without further qualification, apparently without considering the possibility that they may be metastases and manifestations of systemic disease. In this study, 17 of 19 cases of locally recurrent melanoma at the primary excision site showed the histological features of metastasis rather than residual incompletely excised primary melanoma. Because the prevention of local recurrence is the main reason given in recommendations for wide excision of melanoma beyond complete excision of the primary tumour itself, it is essential that surgeons and pathologists should classify these neoplasms precisely as either persistent incompletely excised primary melanoma or metastatic melanoma.

Adult↗