Depurination-induced infidelity of deoxyribonucleic acid synthesis with purified deoxyribonucleic acid replication proteins in vitro

Removal of purine bases from phi X174 single-stranded DNA leads to increased reversion frequency of amber mutations when this DNA is copied in vitro with purified DNA polymerases. This depurination-induced mutagenesis is observed at three different genetic loci and with several different purified enzymes, including Escherichia coli DNA polymerases I and III, avian myeloblastosis virus DNA polymerase, and eukaryotic DNA polymerases alpha, beta, and gamma. The extent of mutagenesis correlates with the estimated frequency of bypass of the lesion and is greatest with inherently inaccurate DNA polymerases which lack proofreading capacity. With E. coli DNA polymerase I, conditions which diminish proofreading result in a 3-5-fold increase in depurination-induced mutagenesis, suggesting a role for proofreading in determining the frequency of bypass of apurinic sites. The addition of E. coli single-stranded DNA-binding protein to polymerase I catalyzed reactions with depurinated DNA had no effect on the extent of mutagenesis. Analysis of wild-type revertants produced during in vitro DNA synthesis by polymerase I or avian myeloblastosis virus DNA polymerase on depurinated phi X174 amber 3 DNA indicates a preference for insertion of dAMP opposite the putative apurinic site at position 587. These results are discussed in relation both to the mutagenic potential of apurinic sites in higher organisms and to studies on error-prone DNA synthesis.     

Distribution and innervation of short, interdigitated muscle fibers in parallel-fibered muscles of the cat hindlimb

The cat hindlimb contains several long, biarticular strap muscles composed of parallel muscle fascicles that attach to short tendons. Three of these muscles--sartorius, tenuissimus, and semitendinosus--were studied by dissecting individual gold-stained fibers and determining the surface distribution of acetylcholinesterase-stained end-plate zones. In each muscle, fascicles were composed of muscle fibers that ran only part of the fascicle length and tapered to end as fine strands that interdigitated with other tapering fibers within the muscle mass. Most muscle fibers measured 2-3 cm in length. Fascicles of muscle fibers were crossed by short transverse bands of endplates (1 mm wide by 1-5 mm long) that were spaced at fairly regular intervals from the origin to the insertion of the muscle. The endplate pattern suggested that the fiber fascicles were organized into multiple longitudinal strips. In the sartorius, the temporospatial distribution of electromyographic (EMG) activity evoked by stimulating fine, longitudinal branches of the parent nerve confirmed that each strip was selectively innervated by a small subset of the motor axons. These axons appeared to distribute their endings throughout the entire length of the fascicles, providing for synchronous activation of their in-series fibers.     

The chicken receptor for endocytosis of glycoproteins contains a cluster of N-acetylglucosamine-binding sites

The oligomeric state of the chicken hepatic receptor for N-acetylglucosamine-terminated glycoproteins (the chicken hepatic lectin) has been examined in detergent solution, in various membrane preparations, and in hepatocytes. In detergent solution, the cross-linking reagent, 1,5-difluoro-2,4-dinitrobenzene produces covalent complexes containing up to six receptor polypeptides. This result, along with hydrodynamic studies of the receptor-detergent complex, indicates that the purified receptor is a hexamer. Analysis of large proteolytic fragments of the receptor reveals that portions of the receptor polypeptide near the membrane anchor are essential for hexamer stability. This analysis also demonstrates that each receptor polypeptide has an N-acetylglucosamine-binding site, indicating that the native hexameric receptor contains a cluster of six such sites. Immunoblot analysis of membrane fractions and cells cross-linked with 1,5-difluoro-2,4-dinitrobenzene or dimethyl adipimidate reveals that the receptor is also oligomeric in intact cells and in subcellular fractions representing cell surface and internalized receptor. Although the pattern of cross-linking observed in membranes differs from that observed with purified receptor, experiments indicate that the differences may be explained by the presence of membrane components which compete with receptor for reaction with cross-linking reagent. The presence of a cluster of carbohydrate-binding sites in the hepatocyte membrane can account for the preferential endocytosis of multivalent glycoprotein ligands by hepatocytes.     

Vertical distributions and relations of euphausiid populations off Elephant Island,March 1984

Summary Distributional relationships are described for post-larval and larval Euphausia superba and Thysanoessa sp. (probably macrura) and post-larval Euphausia frigida collected in 0–70/80 m and 0–175/200 m depth ranges with a MOCNESS sampler north of Elephant Island (61°S, 55°W) during 17–23 March 1984. Larval E. superba (predominantly calyptopes stage 2 and 3) were rare shallower than 80 m at night. Day catches of post-larval E. suberba were small and night catches were primarily near the top of the thermocline above 50 m depth. Thysanoessa sp. occurred throughout the 0–200 m depth range and was abundant in the upper 80 m both night and day. E. frigida migrated to the upper 80 m at night from deeper day depths. Larval stages of E. superba and bost-larval stages of all three species demonstrated independent and variable vertical distribution patterns both night and day. Changes in E. superba abundance and distributional patterns could to a certain extent be associated with observed environmental changes. An increase in larval and decrease in post-larval E. superba abundances between 0–80 m was associated with an intrusion of cold water at depth. At night, vertically restricted concentrations of post-larval E. superba were associated with shallow mixed layer depths, and a significant vertical separation of developmental stages and size categories was observed only during periods of stratification in the upper 80 m. Fluctuations in the distribution and abundance of Thysanoessa sp. and distribution of E. frigida did not appear to be influenced by physical parameters within the upper 80 m. Within the 0–80 m depth range, the distributions of these two species differed from each other and from E. superba and showed large tow to tow variability that could not be related to physical parameters in the upper water column.     

Response of microbial adhesives and biofilm matrix polymers to chemical treatments as determined by interference reflection microscopy and light section microscopy.

The polymers involved in the adhesion of Pseudomonas fluorescens H2S to solid surfaces were investigated to determine whether differences between cell surface adhesives and biofilm matrix polymers could be detected. Two optical techniques, i.e., interference reflection microscopy (IRM) and light section microscopy (LSM), were used to compare the responses of the two types of polymer to treatment with electrolytes, dimethyl sulfoxide (DMSO), and Tween 20. To evaluate initial adhesive polymers, P. fluorescens H2S cells were allowed to attach to glass cover slip surfaces and were immediately examined with IRM, and their response to chemical solutions was tested. With IRM, changes in cell-substratum separation distance between 0 and ca. 100 nm are detectable as changes in relative light intensity of the image; a contraction of the polymer would be detected as a darkening of the image, whereas expansion would appear as image brightening. To evaluate the intercellular polymer matrix in biofilms, 3-day-old biofilms were exposed to similar solutions, and the resultant change in biofilm thickness was measured with LSM, which measures film thicknesses between 10 and 1,000 microns. The initial adhesive and biofilm polymers were similar in that both appeared to contract when treated with electrolytes and to expand when treated with Tween 20. However, with DMSO treatment, the initial adhesive polymer appeared to contract, whereas there was no change in thickness of the biofilm polymer. These results indicate that both polymers bear acidic groups and thus act electrostatically with cations and are able to enter into hydrophobic interactions.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evaluation of the mechanisms by which gamma-amino-butyric acid in association with phosphatidylserine exerts an antiepileptic effect in the rat

The i.p. injection in rats of GABA (740 mg/Kg) after sonication with an equal amount of phosphatidylserine (PS) has an antiepileptic effect. The injection of plain GABA has no such an effect. Blood, brain and synaptosomal accumulation of exogenous labeled GABA under the two circumstances are evaluated. In the case of GABA/PS injection there is a higher passage of the exogenous labeled neurotransmitter into the blood and brain nerve endings (synaptosomes). A higher synaptosomal accumulation of the exogenous labeled neurotransmitter is found even when GABA and PS are injected separately. Since these accumulation increases occur at a time when there is the antiepileptic effect, they seem relevant to it. Our interpretation of the chain of the events resulting in the antiepileptic action is that the phospholipid facilitates from the beginning the first passage of the exogenous neurotransmitter form the peritoneum to the blood. Then a higher passage to the brain tissue and eventually to the GABA-ergic nerve endings ensues. The brisker accumulation of the exogenous neurotransmitter in the nerve endings could be at the basis of a more efficient GABA-ergic inhibitory control in the brain.     

Hemolymph concentrations of host ecdysteroids are strongly suppressed in precocious prepupae of Trichoplusia ni parasitized and pseudoparasitized by Chelonus near curvimaculatus.

Regulation of ecdysteroid production in lepidopteran prepupae was studied using a parasitic wasp (C. near curvimaculatus) which specifically suppresses host prepupal ecdysteroid production after the induction of precocious host metamorphosis. At the developmental stage at which the hemolymph of the unparasitized metamorphosing host has its maximum titer of prepupal ecdysteroids, the hemolymph of 4th instar "truly parasitized" hosts (hosts with a surviving endoparasite) had a strongly reduced ecdysteroid titer. However, during the photophase about 12 h later, just prior to emergence of the parasite larva, an ecdysteroid peak was observed in the host hemolymph. Fourth instar pseudoparasitized prepupal hosts (in which the endoparasite was not present or died early in development) exhibited a sustained suppression in the hemolymph ecdysteroid titer. Small 5th instar pseudoparasitized hosts, which normally would molt to a 6th instar prior to metamorphosis, but which precociously attained the prepupal stage, also had a strongly reduced ecdysteroid titer. The late increase observed in truly parasitized hosts could be completely prevented by surgical removal of the parasite 24 h earlier, resulting in a titer similar to that in pseudoparasitized hosts. HPLC analysis of ecdysteroids in normal, truly parasitized, and 4th or 5th instar pseudoparasitized prepupae showed that both ecdysone and 20-OH ecdysone* were suppressed in truly and pseudoparasitized prepupae, with ecdysteroid levels being lowest in pseudoparasitized hosts. These data, and those of Brown and Reed-Larsen (Biol Contr 1, 136 [1992]), showing endoparasite secretion of ecdysteroids just prior to its emergence from the host, strongly indicate that: (1) the prepupal peak in truly parasitized hosts originates from the endoparasite, and (2) the low level of ecdysteroids in pseudoparasitized hosts results from the host's intrinsic inability to express a normal level of prepupal ecdysteroid titer. While precocious 4th or 5th instar prepupae of similar size had similarly suppressed ecdysteroid titers, smaller 4th instar prepupae had a lower ecdysteroid titer than larger, precocious 5th instar prepupae. Rare 5th instar pseudoparasitized prepupae that were of nearly normal size showed a prepupal ecdysteroid titer distinctly greater than those of the usual smaller, precocious 5th instar prepupae. The data suggest that the competence of the host to express a normal hemolymph titer of prepupal ecdysteroids is more closely correlated with the size of the prepupae than with the instar attained.