Activation of topoisomerase I by poly [ADP-ribose] polymerase.

Poly(ADP-ribose) polymerase (PARP I) and Topoisomerase I (Topo I) were reisolated from calf thymus to eliminate cross contamination as tested by immunotransblots. The specific activity of Topo I was greatly increased by added PARP I, following saturation kinetics. Recombinant PARP I and isolated PARP I at final purity were indistinguishable in terms of their activation of Topo I. There was a coincidence of experimentally obtained binding constants and computer generated values based on the kinetic model, indicating that the association of PARP I and Topo I is rate limiting in the catalytic activation of Topo I by PARP I. Polypeptide domains of PARP I that are required for protein-protein binding and protein-DNA binding also activate Topo I. Fluorescence resonance energy transfer between fluorophor-labeled PARP I and Topo I was demonstrated. The binding of Topo I to circular SV40 DNA, assayed either by the formation of a) the sum of non-covalently and covalently attached Topo I to DNA or b) by the covalently bound transient intermediate in the presence of camptothecin, was augmented when PARP I protein was bound to SV40 DNA. These binding experiments provide a molecular basis for the kinetic activation of Topo I by PARP I inasmuch as the increased superhelicity of SV40 DNA induced by PARP I may facilitate the formation of a more Topo I-DNA complex that increases the rate of the DNA breakage-reunion cycle of Topo I catalysis.

Reversion of malignant phenotype by 5-iodo-6-amino-1,2-benzopyrone a non-covalently binding ligand of poly(ADP-ribose) polymerase.

A non-covalently binding inhibitory ligand of poly(ADP-ribose) polymerase, 5-iodo-6-amino-1,2-benzopyrone, when incubated at 5-600 microM external concentration with an E-ras-transformed tumorigenic cell line or with human prostatic carcinoma cells for 40 to 60 days converts both cancer cells to a non-tumorigenic phenotype that is characterized by drastic changes in cell morphology, absence of tumorigenicity in nude mice, and a high rate of aerobic glycolysis.

DNA hybridization evidence of hominoid phylogeny: a reanalysis of the data.

Sibley and Ahlquist (1984, 1987) presented the results of a study of 514 DNA-DNA hybrids among the hominoids and Old World monkeys (Cercopithecidae). They concluded that the branching order of the living hominoid lineages, from oldest to most recent, was gibbons, orangutan, gorilla, chimpanzees, and human. Thus, a chimpanzee-human clade was indicated, rather than the chimpanzee-gorilla clade usually suggested from morphological evidence. The positions of the gibbon and orangutan branches in the phylogeny are supported by substantial evidence, but whether the chimpanzee lineage branched most recently from the human lineage or from the gorilla lineage remains controversial. The conclusions of Sibley and Ahlquist (1984, 1987) have been supported by several independent studies cited by Sibley and Ahlquist (1987), plus the DNA sequence data of Hayasaka et al. (1988), Miyamoto et al. (1988), Goodman et al. (1989, 1990), and the DNA-DNA hybridization data of Caccone and Powell (1989). The laboratory and data analysis methods have been criticized by Marks et al. (1988) and Sarich et al. (1989). In response to these critics, and for our own interests, we present a reanalysis of the Sibley and Ahlquist data, including a description of the corrections applied to the "raw counts." The validity of the laboratory methods is supported by the congruence of tree topology and delta values with those of Caccone and Powell (1989), although their tetraethylammonium chloride technique differs from the hydroxyapatite method in several respects. The utility of the T50H distance measure is indicated by its congruence with percent sequence divergence at least to delta T50H 30, as noted by Goodman et al. (1990). The Sibley and Ahlquist uncorrected data indicate that Pan is genetically closer to Homo than to Gorilla, but that Gorilla may be genetically closer to Pan than to Homo. Melting curves are presented for the pertinent experiments, plus one that includes representatives of most of the groups of living primates.

A prospective study of infections in hemodialysis patients: patient hygiene and other risk factors for infection.

We performed a prospective randomized study on 71 patients on chronic outpatient hemodialysis to determine whether a sterile technique was better than a clean technique for preparation of the skin over the vascular access site prior to cannulation. In addition, we wanted to determine overall and site-specific infection rates, microbial etiologies of infection, and risk factors for infection. The overall infection rate was 4.7 infections per 100 dialysis months; the vascular access-site infection rate was 1.3 infections per 100 dialysis months; and the rate for bacteremia was 0.7 cases per 100 dialysis months. Staphylococcus aureus was the most common pathogen, but infections were equally divided between gram-positive cocci and gram-negative bacilli. Advanced age (P = 0.02), a low Karnofsky activity score (P = 0.05), poor hygiene (P = 0.0004) and number of hospitalizations (P = 0.0002) were risk factors for infections in general while only poor hygiene (P = 0.002) was a risk factor for vascular access-site infection. Sterile preparation of the skin over the vascular access site was no more effective at preventing infection than was clean technique (P = 0.80). Maintenance of good personal hygiene may be one of the most important measures for prevention of infections in hemodialysis patients.

Prospective study of microbial colonization of the nose and skin and infection of the vascular access site in hemodialysis patients.

We conducted a prospective study of nasal and skin floras in 71 patients receiving chronic hemodialysis. We wished to determine whether a sterile skin preparation technique was more effective than a clean technique in removing microorganisms from the skin of the vascular access site. We also examined the effect of administration of antibiotics and status of patient hygiene on microbial flora. The presence of Staphylococcus aureus in the nose had a low predictive value for the simultaneous presence of the microorganism on the skin. The status of skin colonization can be accurately assessed only by culture of the skin. Sterile technique was no more effective at removing microorganisms from skin than was clean technique. Antibiotics significantly affected nasal flora but not skin flora. S. aureus was significantly more likely to remain on the skin after application of an antiseptic in patients with poor hygiene than in patients with good hygiene (P = 0.002). Patients with poor hygiene also had a significantly higher concentration of S. aureus on the skin of the vascular access site after application of antiseptic than patients with good hygiene (P = 0.005). We found no evidence to support a change from clean to sterile technique for skin preparation, but improvement in personal hygiene may be an effective strategy for prevention of vascular access infections.

Streptococcal perianal disease in children.

From October 1985 through June 1986, 31 children in a single pediatric practice were treated for perianal signs and symptoms associated with growth of group A beta-hemolytic streptococci from perianal cultures. Signs and symptoms included perianal dermatitis (90%), perianal itching (78%), rectal pain (52%), and blood-streaked stools (35%). Ages ranged from 7 months to 8 years mean 4.25 +/- 1.8 years). There were 24 boys (77%) and seven girls (23%). The 31 cases occurred in 19 families. Intrafamily spread was only to siblings and occurred in 50% of the possible situations. Direct perianal antigen studies had a sensitivity of 89% for predicting positive cultures. Four different T types of group A streptococci were isolated from these cases, but the T type within each family outbreak was identical except in one case. When group A streptococci were found in the pharynx (64% of patients), the T type of the pharyngeal and perianal isolates were identical. Treatment was usually with oral penicillin. Relapses occurred in 39%. Signs of cellulitis were absent in all 31 cases and, therefore, we suggest that the nomenclature for this entity be changed from streptococcal perianal cellulitis to streptococcal perianal disease.