bluuebunny/crossref_metadata_2025 huggingface dataset:
Sequence-dependent energetics of the B-Z transition in supercoiled DNA containing nonalternating purine-pyrimidine sequences.
Authors: Ellison M J , Kelleher R J , Wang A H , Habener J F , Rich A
Published: May 2006

The likelihood that a given DNA sequence will adopt the Z conformation in negatively supercoiled DNA depends on the energy difference between the B form and the Z form for that sequence relative to other sequences in the same molecule. This energy can be viewed simply as a sum of energies for the nearest-neighbor interactions within the sequence plus the energy required to stabilize the B-Z boundaries. Knowledge of these energetic terms would be of value in predicting when sequences become left-handed in response to negative superhelicity. Here we present an approach that can be used to determine the free-energy changes associated with all the nearest-neighbor interactions that can occur in Z-DNA. Synthetic stretches of d(C-G)n containing one or two transversions were cloned into plasmids, and the extent of the B-Z transition as a function of negative superhelicity was determined for each insert by two-dimensional agarose gel electrophoresis. By subjecting the data to statistical mechanical analysis, it was possible to evaluate the energetic penalty resulting from each base-pair (bp) substitution. Guanine to cytosine transversions cost 2.4 kcal (1 cal = 4.18 J)/(mol X bp), whereas guanine to thymine transversions cost 3.4 kcal/(mol X bp), to stabilize in the Z conformation. We have used these numbers, along with energetic values determined by others for the B-Z transition, to predict that certain strictly nonalternating purine and pyrimidine sequences may adopt the Z form readily.
Preimaginal learning as a basis of colony-brood recognition in the ant
Cataglyphis cursor
Authors: Isingrini Michel , Lenoir Alain , Jaisson Pierre
Published: May 2006

In most circumstances, social insects recognize their nestmates. They can discriminate against alien adults and also against alien larvae. Results presented here indicate that the mechanism of colony-brood recognition is acquired in large part during larval life and persists through the metamorphosis into the adult stage. During the first days after emergence of the adult, a weaker form of learning can also occur. These phenomena are discussed in relation to kinship theory. It appears that kin recognition is determined not so much by genetic relatedness as by spatial proximity of the individuals during the early stages of life.
Heteroantibody duplexes target cells for lysis by cytotoxic T lymphocytes.
Authors: Liu M A , Kranz D M , Kurnick J T , Boyle L A , Levy R , Eisen H N
Published: May 2006

Antibodies to the clonally unique variable-region determinants (idiotype) of the antigen-specific alpha beta heterodimeric receptor of a clone of cytotoxic T cells (CTLs) were shown previously to render diverse cells, regardless of their own surface antigens, susceptible to lysis by that clone of CTLs. To extend these findings, we have sought to develop a general means for targeting cells for destruction by any CTL, without regard to its alpha beta idiotype and specificity for antigen. We explored the use of heteroantibody duplexes formed by joining covalently an antibody to the T3 complex (anti-T3), which is associated with the alpha beta receptors on all human mature T cells, and a second antibody, specific for an antigen on the intended target cell. The second antibody selected in this study was specific for the idiotype (Id) of the surface immunoglobulin of a human B-lymphoma (anti-Ig Id). In the presence of the anti-T3/anti-Ig Id heteroantibody duplex the B-lymphoma cells were lysed by a clone of human T8+ CTLs (of unrelated specificity) but not by a noncytotoxic clone of human T4+ helper T cells, and lysis by the CTLs was specifically blocked by the uncoupled anti-T3 or the uncoupled anti-Ig Id antibodies. The extent of the heteroantibody-dependent cytolysis depended both on the heteroantibody concentration and on whether the intended target cells or the CTL effectors were initially preincubated with the heteroantibody. Under optimal conditions, heteroantibody-dependent lysis of the surrogate target (B-lymphoma) cells by the CTLs compared favorably with lysis of their natural target cells by the same CTLs. Overall, our findings suggest that heteroantibody duplexes containing anti-T3 antibody may be capable of targeting selected cells, such as tumor cells, for destruction in vivo by the body's CTLs.
Scintillation proximity radioimmunoassay utilizing 125I-labeled ligands.
Authors: Udenfriend S , Gerber L D , Brink L , Spector S
Published: May 2006

A unique type of radioimmunoassay is described that does not require centrifugation or separation. Microbeads containing a fluorophor are covalently linked to antibody. When an 125I-labeled antigen is added it binds to the beads and, by its proximity, the emitted short-range electrons of the 125I excite the fluorophor in the beads. The light emitted can be measured in a standard scintillation counter. Addition of unlabeled antigen from tissue extracts displaces the labeled ligand and diminishes the fluorescent signal. Application of scintillation proximity immunoassay to tissue enkephalins, serum thyroxin, and urinary morphine is described. Applications of the principle to study the kinetics of interaction between receptors and ligands are discussed.
Atrial natriuretic factor inhibits dehydration- and angiotensin II-induced water intake in the conscious, unrestrained rat.
Authors: Antunes-Rodrigues J , McCann S M , Rogers L C , Samson W K
Published: May 2006

Peptides isolated from atrial extracts possess potent natriuretic and diuretic activities. In general, these peptides, called atrial natriuretic factors (ANFs), oppose the actions of the water-conservatory peptides angiotensin II and vasopressin and are released from the heart in response to atrial stretch as a consequence of increased venous return. The recent description of ANF-like immunoreactivity in brain regions associated with the control of water intake suggested a role for these peptides in the neurogenic mechanisms of thirst. Intracerebroventricular (third ventricle) infusion of 1.0 or 2.0 nmol of ANF in conscious, overnight-dehydrated rats significantly inhibited subsequent water intake over a 2-hr test period. Intravenous infusion of 2.0 nmol, but not 1.0 nmol, of ANF resulted in a similar inhibitory action, suggesting that ANF released from the heart might act centrally to inhibit water intake by an action at one or more of the circumventricular organs. Water intake induced by central infusion of angiotensin II (9.6 and 25 pmol) in normally hydrated rats was significantly inhibited by prior infusion of 2.0 nmol of ANF. Water intake induced by higher doses of angiotensin II was not altered significantly by prior infusion of ANF. These results indicate a possible physiologic role for ANF in the hypothalamic control of water intake and reveal that the cardiac peptides can act centrally, as well as peripherally, to assist in the normalization of extracellular fluid volume.
Purified human factor VIII procoagulant protein: comparative hemostatic response after infusions into hemophilic and von Willebrand disease dogs.
Authors: Brinkhous K M , Sandberg H , Garris J B , Mattsson C , Palm M , Griggs T , Read M S
Published: May 2006

The procoagulant protein F.VIII:C is noncovalently bound to von Willebrand factor (vWF) to give the factor VIII macromolecular complex. New highly purified preparations of isolated human F.VIII:C, devoid of vWF and about 500,000-fold purified, were administered to hemophilia A and von Willebrand disease (vWD) dogs to determine their hemostatic effectiveness and survival in the circulation. Two preparations of F.VIII:C were used: peak 1, with active components of Mr 185,000-280,000, and peak 2, with a single component of Mr 170,000. In hemophilic dogs, with no plasma F.VIII:C but normal vWF, both preparations immediately elevated plasma F.VIII:C to expected levels, promptly stopped induced and spontaneous hemorrhages, and gave sustained plasma levels of F.VIII:C. The isolated F.VIII:C immediately complexed with endogenous vWF in hemophilic plasma and was eliminated exponentially, with a half-life (t1/2) of about 9 hr. Survival of peak 2 F.VIII:C was longer than that of peak 1 material. In contrast, F.VIII:C complexed to vWF in a therapeutic concentrate administered to hemophilic dogs was eliminated biexponentially with first-phase t1/2 of 3.2 hr and second-phase t1/2 of 9 hr. In vWD dogs with no vWF and reduced F.VIII:C levels, the isolated F.VIII:C produced supernormal levels of F.VIII:C without effect on induced bleeding. It was rapidly eliminated from plasma with a t1/2 of about 1 hr, as was the complexed F.VIII:C in the concentrate. These data indicate that isolated F.VIII:C promptly complexes with vWF and in this form is highly effective in controlling hemophilic hemorrhages with good survival in plasma. Without endogenous vWF with which to complex, the F.VIII:C is promptly eliminated.
"Frizzy" genes of Myxococcus xanthus are involved in control of frequency of reversal of gliding motility.
Authors: Blackhart B D , Zusman D R
Published: May 2006

Myxococcus xanthus, a Gram-negative bacterium, has a complex life cycle that includes fruiting body formation. Frizzy (frz) mutants are unable to aggregate normally, instead forming frizzy filamentous aggregates. We have found that these mutants are defective in the control of cell reversal during gliding motility. Wild-type cells reverse their direction of gliding about every 6.8 min; net movement occurs since the interval between reversals can vary widely. The frzA-C, -E and -F mutants reverse their direction of movement very rarely, about once every 2 hr. These mutants cannot aggregate normally and give rise to frizzy filamentous colonies on fruiting agar or motility agar. In contrast, frzD mutants reverse their direction of movement very frequently, about once every 2.2 min; individual cells show little net movement and form smooth-edged "nonmotile" type colonies. Genetic analysis of the frzD locus shows that mutations in this locus can be dominant to the wild-type allele and that its gene product(s) must interact with the other frz gene products. Our results suggest that the frz genes are part of a system responsible for directed movement of this organism.
Nonheritable resistance to chloramphenicol and other antibiotics induced by salicylates and other chemotactic repellents in Escherichia coli K-12.
Authors: Rosner J L
Published: May 2006

Phenotypic resistance to chloramphenicol and ampicillin was induced in sensitive Escherichia coli K-12 strains during incubation with the following substances: acetate, acetylsalicylate (aspirin), benzoate, dimethyl sulfoxide, 1-methyl-2-pyrrolidinone, and salicylate. In addition, acetyl-salicylate and salicylate induced resistance to nalidixic acid and tetracycline. The induction of resistance was highly efficient but varied somewhat with the strain and inducer used. In the presence of inducers, from 10% to 100% of the cells formed colonies on antibiotic media, an increase of 10- to 1000-fold over the controls without inducer. After growth in the absence of these inducers, the cells were normally sensitive to the antibiotics. Thus, the resistance was not due to a heritable change. These inducers also increased the level of chloramphenicol resistance of a strain carrying cat (whose gene product inactivates chloramphenicol by acetylation). All of the inducers are chemotactic repellents for E. coli, and they are detected by the tsr gene product (with the possible exceptions of dimethyl sulfoxide and methylpyrrolidinone, whose modes of detection are not known). Nickel sulfate and cobalt sulfate, repellents that are detected by the tar gene product, neither promoted resistance to chloramphenicol nor prevented the induction of resistance by acetylsalicylate. Since several of the inducers are present in common drugs or foods, it may be of medical importance to evaluate their effects on antibiotic therapies.
Colony formation by primitive hemopoietic progenitor cells in serum-free medium.
Authors: Eliason J F , Odartchenko N
Published: May 2006

A serum-free medium has been developed for clonal growth of murine hemopoietic progenitor cells. In this medium, the number of nonerythroid colonies induced by factors produced by cloned T lymphocytes was 90 +/- 10% of the number generated in serum-containing medium. Erythroid colony number in serum-free cultures containing the T-cell factors but no exogenous erythropoietin was significantly higher than that in cultures with serum, and the cloning efficiency was independent of cell concentration. Further addition of erythropoietin increased erythroid colony number approximately equal to 4-fold. Pure erythropoietin alone stimulated erythroid colony formation, but the cloning efficiency was highly dependent on cell concentration. Analysis of individual colonies generated in serum-free cultures containing the T-cell factors indicated that some contained cells of several hemopoietic lineages, demonstrating that multipotential progenitors can give rise to colonies in this system.
Synthetic peptides from four separate regions of the poliovirus type 1 capsid protein VP1 induce neutralizing antibodies.
Authors: Chow M , Yabrov R , Bittle J , Hogle J , Baltimore D
Published: May 2006

Peptides from different regions of the poliovirus type 1 capsid protein VP1 were synthesized. Antibodies raised against these peptides in rabbits and rats recognized the cognate peptides and denatured VP1. Peptides from four regions of VP1 generated antisera with neutralizing titers specifically against poliovirus type 1. Antisera against all other regions of VP1 failed to neutralize virus infectivity, although some of the antisera clearly bound to native virions. Thus, the neutralizing determinants on VP1 reside in specific noncontiguous regions of the protein and can be defined by specific peptides from these regions.
Intrinsic projections within visual cortex: evidence for orientation-specific local connections.
Authors: Matsubara J , Cynader M , Swindale N V , Stryker M P
Published: May 2006

The functional organization of intrinsic connections within area 18 of cat visual cortex was studied using combined electrophysiological and anatomical techniques. Physiological recordings were first used to map the distribution of orientation preference, ocular dominance, and receptive-field location relative to the cortical surface. Next, localized injections of lectin-conjugated horseradish peroxidase were made into physiologically identified regions within area 18. We found that (i) the local cortical interconnections are made preferentially between cell populations with orthogonal preferred orientations and are independent of the ocular dominance of the cortical cells, (ii) the map of visual space in the cortex is anisotropic with the magnification factor for vertical at least twice that for horizontal visual space, and (iii) the pattern of cortical projections compensates for the functional asymmetry so that a population of interconnected cells represents a roughly circular region of visual space.
Evidence for widespread effects of noradrenaline on axon terminals in the rat frontal cortex.
Authors: Mobley P , Greengard P
Published: May 2006

Noradrenaline, apparently working through activation of beta-adrenergic receptors, alters the state of phosphorylation of a substantial proportion of synapsin I in rat frontal cortex. Since synapsin I is enriched in virtually all axon terminals in this brain region, the results indicate that noradrenaline, a sparsely distributed neurotransmitter, affects a large number of axon terminals in this region. The results also suggest that noradrenaline receptors are present on a substantial percentage of all axon terminals in the cerebral cortex, despite the low density of noradrenergic fibers present. The data support the view that noradrenaline may serve a paracrine function.
Hormonal response region in the mouse mammary tumor virus long terminal repeat can be dissociated from the proviral promoter and has enhancer properties.
Authors: Ponta H , Kennedy N , Skroch P , Hynes N E , Groner B
Published: May 2006

The proviral DNA of mouse mammary tumor virus (MMTV) contains a regulatory region closely associated with its promoter, which subjects transcription to the control of glucocorticoid hormones. Delimitation analysis of a chimeric MMTV long terminal repeat-thymidine kinase gene (LTR-tk) has shown that the hormonal regulation sequence is confined to 202 nucleotides preceding the LTR-specific RNA initiation site. A second RNA initiation site (tk-specific mRNA) placed close to the regulatory MMTV sequence by in vitro recombination is also subjected to hormonal stimulation in transfected cells. A series of plasmids with deletions around the LTR cap site progressing from 3' to 5' was made and functionally tested. In vitro deletion of MMTV LTR sequences comprising the RNA initiation sequence and the "TATA" box do not effect hormonal regulation at the tk-specific mRNA start site. Nucleotides up to position -59 from the LTR initiation site could be deleted without influence on the glucocorticoid regulation, whereas deletions to position -65 abolished the hormonal effect on the tk gene transcription. A short MMTV LTR segment containing nucleotides -236 to -52 from the LTR initiation site was recombined with the tk gene or the alpha-globin gene. This fragment confers hormonal inducibility onto the heterologous genes over distances of 0.4 or 1.1 kilobases. The hormonal response region functions when it is placed either 5' or 3' of the regulated gene in both of the possible orientations and is reminiscent of an enhancer sequence.
Two genes encoding steroid 21-hydroxylase are located near the genes encoding the fourth component of complement in man.
Authors: White P C , Grossberger D , Onufer B J , Chaplin D D , New M I , Dupont B , Strominger J L
Published: May 2006

Two genes encoding steroid 21-hydroxylase [21-OHase; steroid 21-monooxygenase; steroid, hydrogen-donor: oxygen oxidoreductase (21-hydroxylating); EC 1.14.99.10], a cytochrome P-450 enzyme, have been located within the HLA major histocompatibility complex. Congenital adrenal hyperplasia due to 21-OHase deficiency is a common inherited disorder of cortisol biosynthesis which is in genetic linkage disequilibrium with certain extended HLA haplotypes. These haplotypes include characteristic serum complement allotypes. A series of cosmid clones was isolated from a human genomic library by using a probe encoding part of the fourth component of complement, C4. These clones also hybridized with a probe encoding most of human 21-OHase. Restriction mapping and hybridization analysis showed that there are two 21-OHase genes, each located near the 3' end of one of the two C4 genes. Hybridization with probes specific for the 5' and 3' ends of the 21-OHase gene showed that the 21-OHase and C4 genes all have the same orientation. The 21-OHase genes 3' to C4A and C4B carry T aq I fragments of 3.2 and 3.7 kilobases (kb), respectively. Both of these fragments are found in genomic DNA of most individuals. In DNA from an individual with the severe, "salt-wasting" form of 21-OHase deficiency who was homozygous for HLA-A3;Bw47;C4A*1;C4B*Q0(null); DR7, the 3.7-kb Taq I fragment is absent, whereas hormonally normal individuals homozygous for HLA-A1;B8;C4A*Q0;C4B*1;DR3 do not carry the 3.2-kb Taq I fragment. These data suggest that the 21-OHase "B" gene (3.7-kb Taq I fragment) is functional, but the 21-OHase "A" gene (3.2-kb Taq I fragment) is not.
Isolation of cDNA clones coding for the beta subunit of human beta-hexosaminidase.
Authors: O'Dowd B F , Quan F , Willard H F , Lamhonwah A M , Korneluk R G , Lowden J A , Gravel R A , Mahuran D J
Published: May 2006

The major forms of beta-hexosaminidase (2-acetamido-2-deoxy-beta-D-glucoside acetamidodeoxyglucohydrolase, EC 3.2.1.30) occur as multimers of alpha and beta chains--hexosaminidase A (alpha beta a beta b) and hexosaminidase B 2(beta a beta b). To facilitate the investigation of beta-chain biosynthesis and the nature of mutation in Sandhoff disease, a human hexosaminidase beta-chain cDNA clone was isolated. Hexosaminidase B (10 mg) was treated with CNBr, five peptide fragments were isolated by reverse-phase HPLC, and their amino acid sequences were determined. One of these contained a string of six amino acids from which an oligonucleotide probe was defined. The simian virus 40-transformed human fibroblast cDNA library of Okayama and Berg was screened by colony hybridization with the radiolabeled probe. Thirteen probe-binding clones were selected out of 50,000 clones screened. Four of these designated pHex were shown to be identical at their 3' ends by restriction enzyme mapping, differing only in their 5' extensions (1.4-1.7 kilobases). The nucleotide sequence of a 174-base-pair segment contained the deduced amino acid sequence of two of the five CNBr peptides, indicating that the pHex clones encode the beta subunit of hexosaminidase. In addition, pHex cDNA was found homologous to multiple bands in digests of genomic human DNA totaling 43 kilobases (kb), all of which were mapped to chromosome 5 in somatic cell hybrids, as expected of the HEXB gene. The pHex cDNA also hybridized to a 2.2-kilobase RNA that apparently codes for the pre-beta-polypeptide of hexosaminidase. This RNA species was absent in the fibroblasts of one of three patients with Sandhoff disease examined. We anticipate that these clones will be of value to diagnosis and carrier detection of Sandhoff disease in affected families.
NH2-terminal specificity and axonal localization of adrenocorticotropin binding sites in rat median eminence.
Authors: Van Houten M , Khan M N , Walsh R J , Baquiran G B , Renaud L P , Bourque C , Sgro S , Gauthier S , Chretien M , Posner B I
Published: May 2006

Adrenocorticotropin binding sites in the rat median eminence have been localized in vivo. These binding sites occur in the basalar zone, which is rich in axonal endings. Using competitive binding and quantitative light-microscope radioautography, we found that the median-eminence binding site, in contradistinction to the adrenal receptor, binds specifically the residue 4-10 region of the adrenocorticotropin molecule. Using quantitative electron-microscope radioautography and median-eminence deafferentation, we localized the binding sites to axon terminals in this region. In time-delayed uptake studies using light-microscope radioautography, we failed to observe concentration of radiolabel in neurons of the medial basal hypothalamus after the direct injection of radioiodinated adrenocorticotropin(1-24) into the median eminence.
Nonenzymatic sequence-specific cleavage of single-stranded DNA.
Authors: Chu B C , Orgel L E
Published: May 2006

Ethylenediaminetetraacetic acid and diethylenetriaminepentaacetic acid have been attached covalently to the 5' terminus of the deoxynucleotide sequence C-A-C-A-A-T-T-C-C-A-C-A-C-A-A-C (16-mer) via an ethylenediamine linker. In the presence of Fe2+ and dithiothreitol, these reagents bring about the hybridization-dependent cleavage of the sequence T-C-G-T-A-T-G-T-T-G-T-G-T-G-G-A-A-T-T-G-T-G-A-G-C-G-G-A-T-A-A-C-A-A-T-T- T (37-mer), a sequence that contains an internal subsequence complementary to the 16-mer. The principal cleavage sites on the 37-mer are about four residues on each side of the terminal phosphate group of the 16-mer.
gp140, the C3d receptor of human B lymphocytes, is also the Epstein-Barr virus receptor.
Authors: Frade R , Barel M , Ehlin-Henriksson B , Klein G
Published: May 2006

The relationship between gp140, the membrane C3d receptor (CR2) of human B lymphocytes, and the Epstein-Barr virus receptor (EBVR) was analyzed by using the polyclonal anti-gp140, previously prepared by immunizing rabbits with highly purified gp140 (isolated by some of us) from CR2/EBVR-positive Raji cells. Polyclonal anti-gp72, a C3-binding membrane component, not related to the EBVR but also expressed on the Raji cell surface, was used as a control. Binding of rabbit IgG and EBV on cells was assessed by using immunofluorescence techniques with analysis by flow cytofluorometry. A semiquantitative bioassay was also used to measure the EBV binding. Polyclonal monospecific anti-gp140 IgG inhibits directly the binding of EBV to Raji cells at the same concentration that inhibits the binding of EC3d on cells, whereas a 35 times higher concentration of anti-gp72 IgG or preimmune serum IgG does not. Anti-gp140 IgG treatment also inhibits the induction of EBV-determined nuclear antigen in normal tonsil B lymphocytes or in EBV-negative Ramos cells, whereas high concentrations of anti-gp72 IgG or preimmune serum IgG have no effect. These data strongly suggest that gp140, the CR2 of human B lymphocytes, is also the EBVR.
Characterization of a Q subregion gene in the murine major histocompatibility complex.
Authors: Flaherty L , DiBiase K , Lynes M A , Seidman J G , Weinberger O , Rinchik E M
Published: May 2006

We have used restriction enzyme digests, Southern blot analysis, and gene transfer experiments to identify a class I gene in the Q subregion of the murine major histocompatibility complex. By comparisons of class I genes from Q congeneic strains, five restriction fragment length polymorphisms were identified. Further studies of mutant (Qa-2-) and wild-type (Qa-2+) BALB/c sublines indicated that at least part of the structural or regulatory gene controlling a Q subregion antigen resides on a 3.7-kilobase Xba I DNA fragment and is absent in all tested Qa-2- strains. The spontaneously occurring Qa-2- BALB/cBy mutant appears to have an extensive deletion in this region. The identity of this gene was confirmed by gene transfer experiments as well as by the use of a single-copy probe.
Opacification of gamma-crystallin solutions from calf lens in relation to cold cataract formation.
Authors: Siezen R J , Fisch M R , Slingsby C , Benedek G B
Published: May 2006

To determine the molecular mechanisms for cold cataract formation in the nucleus of the young mammalian lens, we have investigated the thermally reversible opacification of gamma-crystallin solutions isolated from calf lens. Coexistence curves (plots of opacification temperature Tc versus protein concentration) were determined for the individual gamma-crystallin fractions II, III, and IV as well as for the unfractionated gamma-crystallin mixtures isolated from the nucleus and cortex. The coexistence curve of gamma IV-crystallin is remarkably elevated above those of gamma II- and gamma III-crystallin and the gamma-crystallin mixtures. The gamma IV-crystallin fraction is the major determinant of the opacification temperature within the whole lens or isolated cytoplasm. Quasielastic light-scattering spectroscopy of gamma IV-crystallin solutions indicates that above Tc there are two populations of protein aggregates of distinctly different mean size. As the temperature is lowered towards Tc, both populations increase in size. Opacification occurs when the population of large scatterers, which is composed of less than 0.1% protein by weight, reaches an average radius of about 20,000 A.
Protonmotive force-driven active transport of D-glucose and L-proline in the protozoan parasite Leishmania donovani.
Authors: Zilberstein D , Dwyer D M
Published: May 2006

Midlogarithmic phase Leishmania donovani promastigotes accumulate 2-deoxy-D-glucose (2-dGlc) and L-proline, maintaining concentration gradient factors across the surface membrane of 78.7 and 60, respectively. Cyanide (1 mM) and iodoacetate (0.5 mM) inhibited the transport of both substrates. L-proline uptake was also inhibited by 2-dGlc (10 mM). Transport of neither substrate was affected by Na+, phlorizin, or ouabain, indicating the sodium-independent transport of both systems. However, N',N'-dicyclohexylcarbodiimide (DCCD; 20 microM) significantly inhibited the transport of both 2-dGlc and L-proline (70% and 90%, respectively). The ionophores valinomycin (1 microM) and nigericin (5 microM) each partially inhibited the uptake of both substrates. In parallel experiments, nigericin and valinomycin were added concomitantly to promastigotes, each at a concentration that individually inhibited the transport of 2-dGlc and L-proline by less than 30%. Under such conditions, the transport of 2-dGlc and L-proline was inhibited by 69% and 78%, respectively. However, these ionophores had no significant effect on the promastigotes cellular ATP level. Carbonylcyanide p-(trifluoromethoxy)phenylhydrazone (FCCP; 1 microM) inhibited 2-dGlc (79%) and L-proline (85%) transport, whereas ATP levels of such cells were diminished by only 20%. Symport of D-glucose/H+ and L-proline/H+ was measured directly in cells pretreated with KCN and DCCD. Upon addition of D-glucose to such cells, a rapid movement of protons into the organisms occurred and was reversed upon addition of FCCP. Conversely, no proton movement was observed when L-glucose was added to such cells. L-proline, as D-glucose, caused a rapid influx of protons into the promastigotes, indicating that both substrates were cotransported with protons. We conclude that transport of D-glucose and L-proline in L. donovani promastigotes is protonmotive force-driven and is coupled to both delta pH and delta psi.
beta-Adrenergic agonist activity of a monoclonal anti-idiotypic antibody.
Authors: Guillet J G , Kaveri S V , Durieu O , Delavier C , Hoebeke J , Strosberg A D
Published: May 2006

Hybridoma cells bearing monoclonal antibody against the beta-adrenergic ligand alprenolol were used as an immunogen to raise monoclonal anti-idiotypic antibodies. Of six anti-idiotypic antibodies, which inhibit ligand binding, three were able to recognize beta-adrenergic receptors. One of them, mAb2B4, an IgM that could be amplified into ascites, binds to the beta-adrenergic catecholamine receptors of intact epidermoid A431 cells and precipitates receptors solubilized from plasma membranes by digitonin. This antibody identifies the beta 2-adrenergic receptor of A431 cells as a single 55-kDa protein and stimulates adenylate cyclase activity. This stimulation is inhibited by the beta-adrenergic antagonist propranolol.
Ca2+ entry in squid axons during voltage-clamp pulses is mainly Na+/Ca2+ exchange.
Authors: Mullins L J , Requena J , Whittembury J
Published: May 2006

Intact squid axons were injected with aequorin and bathed in 3 mM Ca seawater (a concentration close to that of squid blood). Sodium and potassium currents were pharmacologically blocked and repetitive voltage-clamp pulses of a duration of 1.5 ms were applied (to simulate the duration of an action potential) at amplitudes of +30 to +90 mV and at frequencies of 100/s. In a very fresh axon (low internal Na concentration) no detectable change in aequorin glow resulted from this treatment, whether the axons were in Na-containing or in Na-free seawater. In axons subjected to modest Na loading, repetitive voltage-clamp pulsing did not result in an increased aequorin glow when the pulses were delivered in Na seawater, whereas in Na-free seawater there was an easily measurable increase in aequorin light emission during repetitive pulsing. The increase in aequorin photons emitted per voltage-clamp pulse was e-fold for 22 mV of depolarization, and the process showed no signs of saturating at pulse amplitudes of +180 mV (i.e., at a membrane potential close to ECa). The aequorin light emission per voltage-clamp pulse increased linearly with pulse duration (at constant amplitude).
Evidence that the Mg-dependent low-affinity binding site for ATP and Pi demonstrated on the isolated beta subunit of the F0.F1 ATP synthase is a catalytic site.
Authors: Khananshvili D , Gromet-Elhanan Z
Published: May 2006

Binding sites for one Pi and two ATP or ADP molecules have been identified on the isolated, reconstitutively active beta subunit from the Rhodospirillum rubrum F0.F1 ATP synthase. Chemical modification of this beta subunit by the histidine reagent diethyl pyrocarbonate or by the carboxyl group reagent Woodword's reagent K results in complete inhibition of Pi binding to beta. The same reagents inhibit the binding of ATP to a Mg-dependent low-affinity site but not to a Mg-independent high-affinity site on this beta subunit. The binding stoichiometry of ADP to either site is not affected by these modifications. The beta subunit modified by either one of these reagents retains its capacity to rebind to beta-less chromatophores but not its ability to restore their photo-phosphorylation. These results indicate that the low-affinity Pi binding site on beta is located at the binding site of the gamma-phosphate group of ATP in the Mg-dependent low-affinity nucleotide binding site. This site contains histidine and carboxyl group residues, both of which are required for the binding of Pi and of the gamma-phosphate group of ATP. The same residues must also be involved in the capacity of the isolated beta subunit to restore the catalytic activity of the beta-less ATP synthase. It is therefore concluded that the low-affinity Mg-dependent substrate binding site identified on the isolated beta subunit of the R. rubrum F0.F1 ATP synthase is the catalytic site of this enzyme complex.
Parkinsonism-inducing neurotoxin, N-methyl-4-phenyl-1,2,3,6 -tetrahydropyridine: uptake of the metabolite N-methyl-4-phenylpyridine by dopamine neurons explains selective toxicity.
Authors: Javitch J A , D'Amato R J , Strittmatter S M , Snyder S H
Published: May 2006

N-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) produces neuropathological and clinical abnormalities in humans, monkeys, and mice that closely resemble idiopathic parkinsonism. N-Methyl-4-phenylpyridine (MPP+), a metabolite of MPTP formed by monoamine oxidase B, is accumulated into striatal and cerebral cortical synaptosomes by the dopamine and norepinephrine uptake systems, respectively, whereas MPTP itself is not accumulated. The potencies of drugs in inhibiting [3H]MPP+ or [3H]dopamine uptake into striatal synaptosomes are very similar, as are potencies in inhibiting [3H]MPP+ or [3H]norepinephrine uptake into cortical synaptosomes. The Km values for [3H]MPP+ uptake are 170 and 65 nM and the Vmax values are 2 and 0.1 nmol/g of tissue per min in rat striatum and cortex, respectively, similar to values for [3H]dopamine uptake, Autoradiography of accumulated [3H]MPP+ in slices of rat brain shows high densities in the caudate-putamen and nucleus accumbens. Furthermore, blockade of dopamine uptake by mazindol prevents MPTP-induced damage to nigrostriatal dopamine neurons, indicating that MPP+ concentration into dopamine neurons explains their selective destruction by MPTP.
Alternative view of enzyme reactions.
Authors: Dewar M J , Storch D M
Published: May 2006

Since adsorption of the substrate in the active site of an enzyme can occur only if all solvent is squeezed out from between them, any reaction between them takes place in the absence of any intervening solvent--i.e., as it would in the gas phase. Recent work has shown that ionic reactions in the gas phase often differ greatly from analogous processes in solution. Therefore, current interpretations of enzyme reactions in terms of solution chemistry are misguided. The large rates and specificity of enzyme reactions may be due simply to elimination of the solvent. The cleavage of peptides by chymotrypsin and carboxypeptidase A can be interpreted satisfactorily in this way.
Ultraviolet resonance Raman excitation profiles of nucleic acid bases with excitation from 200 to 300 nanometers.
Authors: Kubasek W L , Hudson B , Peticolas W L
Published: May 2006

Raman spectra are presented for dilute aqueous solutions of the four ribonucleotides AMP, GMP, UMP, and CMP obtained with laser excitation at 299, 266, 253, 240, 229, 218, 209, and 200 nm. Distinct evidence of strong, selective resonance enhancement is obtained. Low-resolution excitation profiles have been constructed for the strongest bands by using the phosphate band at 994 cm-1 as an internal reference. The excitation spectra for many of the vibrational bands are dominated by a peak corresponding to the lowest-energy electronic transition near 260 nm. Smaller peaks are seen for higher-energy electronic transitions. For some modes, the resonance enhancement is dominated by the higher-energy transitions. It is clear from these new data that a full description of the resonance Raman profiles of the nucleic acids will have to include several excited electronic states. Two examples are given of cases where ionic species can be distinguished easily by using far-UV excitation, but these species are indistinguishable with 266-nm excitation. This demonstrates the utility of far-UV resonance Raman spectroscopy for obtaining structural information.
Photoreactivation of UV-induced pyrimidine dimers and erythema in the marsupial Monodelphis domestica.
Authors: Ley R D
Published: May 2006

Post-UV treatment of the gray, short-tailed opossum Monodelphis domestica with photoreactivating light (320-400 nm) suppressed the appearance of UV-induced erythema as evidenced by an increase in the dose of UV required to elicit an erythemal response. The average erythema dose for animals held in the dark following UV exposure was 620 +/- 40 J/m2, whereas 2460 +/- 110 J/m2 were required for erythema induction with animals exposed to 90 min of photoreactivating light post-UV. Pre-UV exposure to photoreactivating light had no effect on the UV induction of erythema. The dose-response for the photoreversal of pyrimidine dimers in epidermal DNA of M. domestica was similar to that for the photoreactivation of erythema induction. These data not only support the notion that DNA is the primary chromophore involved in the induction of erythema but also identify pyrimidine dimers as the major DNA change responsible for its induction. These results also identify M. domestica as a useful whole-animal system with which to determine the role of pyrimidine dimers in other photobiological responses of mammalian skin.
Reduction of gap junctional conductance by microinjection of antibodies against the 27-kDa liver gap junction polypeptide.
Authors: Hertzberg E L , Spray D C , Bennett M V
Published: May 2006

Antibody raised against isolated rat liver gap junctions was microinjected into coupled cells in culture to assess its influence on gap junctional conductance. A rapid inhibition of fluorescent dye transfer and electrical coupling was produced in pairs of freshly dissociated adult rat hepatocytes and myocardial cells as well as in pairs of superior cervical ganglion neurons from neonatal rats cultured under conditions in which electrotonic synapses form. The antibodies have been shown by indirect immunofluorescence to bind to punctate regions of the plasma membrane in liver. By immunoreplica analysis of rat liver homogenates, plasma membranes, and isolated gap junctions resolved on NaDodSO4/polyacrylamide gels, binding was shown to be specific for the 27-kDa major polypeptide of gap junctions. This and similar antibodies should provide a tool for further investigation of the role of cell-cell communication mediated by gap junctions and indicate that immunologically similar polypeptides comprise gap junctions in adult mammalian cells derived from all three germ layers.
Persistence and coexistence of a nonsymbiotic coral in open reef environments
Authors: Wellington G.M. , Trench Robert K.
Published: May 2006

In the Republic of Belau (western Caroline Islands), the nonsymbiotic stony coral
Tubastraea micrantha
(Ehrenberg) occurs commonly on open reef habitats that are dominated by symbiotic corals. This observation calls into question the often-held view that nonsymbiotic corals are restricted to cryptic environments because of their inability to compete successfully for space with rapidly growing symbiotic corals. In this study, we investigated aspects of the biology of
Tubastraea
that might account for its coexistence and persistence on the reef. Our results indicate that despite low calcification rates, linear skeletal extension for
Tubastraea
is surprisingly rapid for a nonsymbiotic species and comparable to those of many symbiotic corals. This factor, coupled with
Tubastraea
's resistance to direct competitive dominance (i.e., by aggressive interactions) and avoidance of predation, promotes its coexistence with symbiotic species. The relative importance of these biological interactions in different coral communities leads to predictions corroborated by observed patterns of local distribution and abundance.
Capillary number and percentage closed in human diabetic sural nerve.
Authors: Dyck P J , Hansen S , Karnes J , O'Brien P , Yasuda H , Windebank A , Zimmerman B
Published: May 2006

The number of capillaries per mm2, minimum intercapillary distance, number of endothelial nuclei per capillary section, and percentage of capillaries closed were evaluated in transverse sections of fascicles of 45 control and 36 diabetic sural nerves. All controls and patients were prospectively studied to ascertain their diabetic and neuropathic status. An index of pathology was introduced and it was found to provide a sensitive and reliable measurement of the presence and severity of neuropathy. The number of capillaries and minimum intercapillary distance of diabetic nerves were not significantly different from those of controls (P greater than 0.05). Diabetic nerves exhibited a small but statistically significant increase in the number of endothelial nuclei per capillary that was positively correlated with the severity of neuropathy. The most striking abnormality was the statistically significant increase in the percentage of capillaries closed in patients with neuropathy as compared to those without neuropathy and controls. Among diabetics, this percentage increased with the severity of neuropathy (P = 0.008). The two capillary abnormalities that have been demonstrated may play a role in the development of diabetic polyneuropathy.
Sequence-specific recognition of B-DNA by oligo(N-methylpyrrolecarboxamide)s.
Authors: Youngquist R S , Dervan P B
Published: May 2006

Four homologous oligopeptide-EDTA molecules, tri-, tetra, penta-, and hexa(N-methylpyrrolecarboxamide)-EDTA, in the presence of Fe(II), O2, and dithiothreitol, cleave 32P-end-labeled restriction fragments from plasmid pBR322 DNA at common locations rich in A X T base pairs that differ in the size of the binding site. From analysis of the cleavage patterns visualized by high-resolution denaturing gel electrophoresis, the oligopeptides with three, four, five, and six N-methylpyrrolecarboxamide units, containing four, five, six, and seven amide NHs, bind sites of A X T-rich DNA consisting of five, six, seven, and eight contiguous base pairs, respectively. The general rule of n amides affording binding site sizes of n + 1 base pairs is consistent with the oligopeptides binding in the minor groove of right-handed DNA, with the amide NH groups forming bridges between the adjacent N-3 and O-2 atoms of adenine or thymine on opposite strands of the DNA helix.
Consensus sequence for Escherichia coli heat shock gene promoters.
Authors: Cowing D W , Bardwell J C , Craig E A , Woolford C , Hendrix R W , Gross C A
Published: May 2006

We have identified promoters for the Escherichia coli heat shock operons dnaK and groE and the gene encoding heat shock protein C62.5. Transcription from each promoter is heat-inducible in vivo, and each is recognized in vitro by RNA polymerase containing sigma 32, the sigma factor encoded by rpoH (htpR) but not by RNA polymerase containing sigma 70. We compared the sequences of the heat shock promoters and propose a consensus promoter sequence, having T-N-t-C-N-C-c-C-T-T-G-A-A in the -35 region and C-C-C-C-A-T-t-T-a in the -10 region. These sequences differ from the consensus sequence recognized by holoenzyme containing sigma 70, the major sigma in E. coli. We suggest that the accumulated consensus sequences of promoters recognized by alternate forms of holoenzyme are compatible with a model in which sigma recognizes only the -10 region of the promoter.
Scavenger receptor-mediated recognition of maleyl bovine plasma albumin and the demaleylated protein in human monocyte macrophages.
Authors: Haberland M E , Fogelman A M
Published: May 2006

Maleyl bovine plasma albumin competed on an equimolar basis with malondialdehyde low density lipoprotein (LDL) in suppressing the lysosomal hydrolysis of 125I-labeled malondialdehyde LDL mediated by the scavenger receptor of human monocyte macrophages. Maleyl bovine plasma albumin, in which 94% of the amino groups were modified, exhibited an anodic mobility in agarose electrophoresis 1.7 times that of the native protein. Incubation of maleyl bovine plasma albumin at pH 3.5 regenerated the free amino groups and restored the protein to the same electrophoretic mobility as native albumin. The demaleylated protein suppressed 75% of the hydrolysis of 125I-labeled malondialdehyde LDL and greater than 80% of 125I-labeled maleyl bovine plasma albumin. The ability of the demaleylated protein to compete was abolished after treatment with guanidine hydrochloride. Although ligands recognized by the scavenger receptor typically are anionic, we propose that addition of new negative charge achieved by maleylation, rather than directly forming the receptor binding site(s), induces conformational changes in albumin as a prerequisite to expression of the recognition domain(s). The altered conformation of the modified protein apparently persists after removal of the maleyl groups. We conclude that the primary sequence of albumin, rather than addition of new negative charge, provides the recognition determinant(s) essential for interaction of maleyl bovine plasma albumin with the scavenger receptor.
B-cell stimulatory factor 1 activates resting B cells.
Authors: Rabin E M , Ohara J , Paul W E
Published: May 2006

B-cell stimulatory factor 1 (BSF-1) is a T-cell-derived lymphokine that acts together with low concentrations of anti-IgM antibodies to stimulate resting B cells to enter the G1 phase of the cell cycle and to synthesize DNA. We show here that supernatants from EL-4 cells, rich in BSF-1 activity, and BSF-1 purified by high-pressure liquid chromatography (HPLC-BSF-1) act on resting B cells, in the absence of anti-IgM antibodies, to prepare them to respond to anti-IgM and BSF-1. A 24-hour preculture with BSF-1 speeds the entry into S phase of B cells subsequently cultured with anti-IgM and BSF-1 by approximately equal to 12 hours and causes substantial increase in cell volume of all resting B cells. Both of these effects, stimulated either by EL-4 supernatants or by HPLC-BSF-1, are inhibited by a monoclonal anti-BSF-1 antibody. These results lead us to propose that BSF-1 should be regarded as a B-cell activation factor.
Hybrid molecules containing the B-domain of insulin-like growth factor I are recognized by carrier proteins of the growth factor.
Authors: De Vroede M A , Rechler M M , Nissley S P , Joshi S , Burke G T , Katsoyannis P G
Published: May 2006

The insulin-like growth factors (IGFs) are polypeptides in plasma that are chemically related to insulin and have mitogenic and insulin-like activity. Unlike insulin, the IGFs circulate in plasma bound to specific high molecular weight carrier proteins that regulate their delivery to target tissues. To define the sites on the IGFs that allow them to be recognized by carrier proteins, we constructed hybrid molecules containing different portions of the insulin, IGF-I, and IGF-II molecules. The presence of the B domain of IGF-I, but not the D domain of IGF-II, enables these insulin-IGF hybrid molecules to be recognized by acid-stripped IGF carrier proteins from rat serum and other sources. By contrast, neither the BIGF-I nor DIGF-II domain is sufficient to enable binding to type II IGF receptors, despite the fact that type II receptors, like the carrier protein, specifically bind IGF-I and IGF-II but do not interact with insulin. By differentiating those sites on the IGF molecule required for binding to IGF carrier protein and receptors, the insulin-IGF hybrid molecules should help delineate the role of the carrier protein in presenting biologically active IGF to target tissues.