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Major advances in understanding the diversity, distribution, and activity of marine N2-fixing microorganisms (diazotrophs) have been made in the past decades, however, large gaps in knowledge remain about the environmental controls on growth and mortality rates. In order to measure diazotroph net growth rates and microzooplankton grazing rates on diazotrophs, nutrient perturbation experiments and dilution grazing experiments were conducted using free-floating in situ incubation arrays in the vicinity of Station ALOHA in March 2016. Net growth rates for targeted diazotroph taxa as well as Prochlorococcus, Synechococcus and photosynthetic picoeukaryotes were determined under high (H) and low (L) nitrate:phosphate (NP) ratio conditions at four depths in the photic zone (25, 45, 75, and 100 m) using quantitative PCR and flow cytometry. Changes in the prokaryote community composition in response to HNP and LNP treatments were characterized using 16S rRNA variable region tag sequencing. Microzooplankton grazing rates on diazotrophs were measured using a modified dilution technique at two depths in the photic zone (15 and 125 m). Net growth rates for most of the targeted diazotrophs after 48 h were not stimulated as expected by LNP conditions, rather enhanced growth rates were often measured in HNP treatments. Interestingly, net growth rates of the uncultivated prymnesiophyte symbiont UCYN-A1 were stimulated in HNP treatments at 75 and 100 m, suggesting that N used for growth was acquired through continuing to fix N2 in the presence of nitrate. Net growth rates for UCYN-A1, UCYN-C, Crocosphaera sp. (UCYN-B) and the diatom symbiont Richelia (associated with Rhizosolenia) were uniformly high at 45 m (up to 1.6 ± 0.5 d-1), implying that all were growing optimally at the onset of the experiment at that depth. Differences in microzooplankton grazing rates on UCYN-A1 and UCYN-C in 15 m waters indicate that the grazer assemblage preyed preferentially on UCYN-A1. Deeper in the water column (125 m), both diazotrophs were grazed at substantial rates, suggesting grazing pressure may increase with depth in the photic zone. Constraining in situ diazotroph growth and mortality rates are important steps for improving parameterization for diazotrophs in global ecosystem models.

The VAHINE mesocosm experiment, conducted in the low-nutrient low-chlorophyll waters of the Noumea lagoon (coastal New Caledonia) was designed to trace the incorporation of nitrogen (N) fixed by diazotrophs into the food web, using large volume (50 m3) mesocosms. This experiment provided a unique opportunity to study the succession of different N2-fixing microorganisms (diazotrophs) and calculate in situ net growth and mortality rates in response to fertilization with dissolved inorganic phosphate (DIP) over a 23-day period, using quantitative polymerase chain reaction (qPCR) assays targeting widely distributed marine diazotroph lineages. Inside the mesocosms, the most abundant diazotroph was the heterocyst-forming Richelia associated with Rhizosolenia (Het-1) in the first half of the experiment, while unicellular cyanobacterial Group C (UCYN-C) became abundant during the second half of the experiment. Decreasing DIP concentrations following the fertilization event and increasing temperatures were significantly correlated with increasing abundances of UCYN-C. Maximum net growth rates for UCYN-C were calculated to range between 1.23 ± 0.07 and 2.16 ± 0.07 d−1 in the mesocosms, which are among the highest growth rates reported for diazotrophs. Outside the mesocosms in the New Caledonia lagoon, UCYN-C abundances remained low, despite increasing temperatures, suggesting that the microbial community response to the DIP fertilization created conditions favorable for UCYN-C growth inside the mesocosms. Diazotroph community composition analysis using PCR targeting a component of the nitrogenase gene (nifH) verified that diazotrophs targeted in qPCR assays were collectively among the major lineages in the lagoon and mesocosm samples, with the exception of Crocosphaera-like phylotypes, where sequence types not typically seen in the oligotrophic ocean grew in the mesocosms. Maximum net growth and mortality rates for nine diazotroph phylotypes throughout the 23-day experiment were variable between mesocosms, and repeated fluctuations between periods of net growth and mortality were commonly observed. The field population of diazotrophs in the New Caledonian lagoon waters appeared to be dominated by Het-1 over the course of the study period. However, results from both qPCR and PCR analysis indicated a diverse field population of diazotrophs was present in the lagoon at the time of sampling. Two ecotypes of the Braarudosphaera bigelowii symbiont unicellular group A (UCYN-A) were present simultaneously in the lagoon, with the recently described B. bigelowii/UCYN-A2 association present at higher abundances than the B. bigelowii/UCYN-A1 association.

The effects of ocean acidification on nitrogen (N2) fixation rates and on the community composition of N2-fixing microbes (diazotrophs) were examined in coastal waters of the North-Western Mediterranean Sea. Nine experimental mesocosm enclosures of ∼50 m3 each were deployed for 20 days during June–July 2012 in the Bay of Calvi, Corsica, France. Three control mesocosms were maintained under ambient conditions of carbonate chemistry. The remainder were manipulated with CO2 saturated seawater to attain target amendments of pCO2 of 550, 650, 750, 850, 1000 and 1250 μatm. Rates of N2 fixation were elevated up to 10 times relative to control rates (2.00 ± 1.21 nmol L−1d−1) when pCO2 concentrations were >1000 μatm and pHT (total scale)

Cyanobacterial mats collected in hypersaline salterns were incubated in a greenhouse under low sulfate concentrations ([SO4]) and examined for their primary productivity and emissions of methane and other major carbon species. Atmospheric greenhouse warming by gases such as carbon dioxide and methane must have been greater during the Archean than today in order to account for a record of moderate to warm paleoclemates, despite a less luminous early sun. It has been suggested that decreased levels of oxygen and sulfate in Archean oceans could have significantly stimulated microbial methanogenesis relative to present marine rates, with a resultant increase in the relative importance of methane in maintaining the early greenhouse. We maintained modern microbial mats, models of ancient coastal marine communities, in artificial brine mixtures containing both modern [SO4=] (ca. 70 mM) and "Archean" [SO4] (less than 0.2 mM). At low [SO4], primary production in the mats was essentially unaffected, while rates of sulfate reduction decreased by a factor of three, and methane fluxes increased by up to ten-fold. However, remineralization by methanogenesis still amounted to less than 0.4 % of the total carbon released by the mats. The relatively low efficiency of conversion of photosynthate to methane is suggested to reflect the particular geometry and chemical microenvironment of hypersaline cyanobacterial mats. Therefore, such mats w-ere probably relatively weak net sources of methane throughout their 3.5 Ga history, even during periods of low- environmental levels oxygen and sulfate.

Summary Mechanisms of dimethyl sulphide (DMS) and methanethiol (MT) production and consumption were determined in moderately hypersaline mats, Guerrero Negro, Mexico. Biological pathways regulated the net flux of DMS and MT as revealed by increases in flux resulting from decreased salinity, increased temperature and the removal of oxygen. Dimethylsulphoniopropionate (DMSP) was not present in these microbial mats and DMS and MT are probably formed by the reaction of photosynthetically produced low-molecular weight organic carbon and biogenic hydrogen sulphide derived from sulphate reduction. These observations provide an alternative to the notion that DMSP or S-containing amino acids are the dominant precursors of DMS in intertidal sediment systems. The major sink for DMS in the microbial mats was biological consumption, whereas photochemical oxidation to dimethylsulphoxide was the major sink for DMS in the overlying water column. Diel flux measurements demonstrated that significantly more DMS is released from the system during the night than during the day. The major consumers of DMS in the presence of oxygen were monooxygenase-utilizing bacteria, whereas under anoxic conditions, DMS was predominantly consumed by sulphate-reducing bacteria and methanethiol was consumed by methanogenic bacteria. Aerobic and anaerobic consumption rates of DMS were nearly identical. Mass balance estimates suggest that the consumption in the water column is likely to be smaller than net the flux from the mats. Volatile organic sulphur compounds are thus indicators of high rates of carbon fixation and sulphate reduction in these laminated sediment ecosystems, and atmospheric sulphur can be generated as a biogenic signature of the microbial mat community.

Acclimation of a marine sediment microbial community to the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) was examined by comparing its ability to metabolize 2,4-D before and after exposure to the herbicide. The experimental treatments (control: 2-week exposure, seawater only; test: 2-week exposure, seawater plus 100 ppm 2,4-D) took place under simulated in situ conditions, in an incubation chamber, which maintained the physical and chemical gradient structure of the sediment community. The surface of the sediment was exposed to recirculating seawater on a tidal cycle. 2,4-D (100 ppm) was added to the seawater so that its availability to the sediment microbes mimicked the natural situation. Before and after treatment, bacterial abundance, productivity, and transformation of 2,4-D were determined. After 2 weeks, bacterial populations were similar in the 2,4-D treatment and the initial sediment sample, but higher in the seawater-only treatment. Bacterial productivity was higher in both 2-week treatments, compared to productivity measured before treatment, but rates were the same in 2,4-D and seawater treatments. In contrast, 2,4-D transformation rates increased 28% in the 2-week 100-ppm 2,4-D treatment, compared to the average of the seawater control and the initial sample. This increased transformation rate indicates that the sediment community acclimated to the selective pressure of 2,4-D treatment by increasing its ability to utilize this compound as a substrate.

Research indicates that bipolar disorder is characterized by both high levels of impairment and high levels of achievement. A critical, and yet largely unexamined question, is: what psychological mechanisms promote high accomplishment (and low impairment) among bipolar spectrum individuals? The aim of this study was to examine this question. The Authors also conceptually explore how the answer to this question can enhance the development of intervention and prevention strategies for adolescents with a bipolar spectrum condition.Academic transcript data were obtained for 120 college students who had either a bipolar spectrum disorder (N=54) or no major psychopathology (N=66).Bipolar spectrum individuals obtained a lower cumulative grade point average (GPA, t=-2.9, P=0.005) and dropped more classes (t=2.1, P=or0.04) than normal controls. The findings also have relevance to the behavioral approach system (BAS) dysregulation theory of bipolar disorder, as well as research on impulsivity among bipolar individuals. Specifically, follow-up analyses revealed that bipolar individuals exhibiting a combination of high BAS drive and low impulsivity earned higher GPAs than the remaining bipolar individuals. Thus, high BAS sensitivity, when paired with low impulsivity, may not be impairing and may contribute to the high achievement sometimes observed among bipolar individuals.Such information is important for the development of prevention and intervention programs designed adolescents that lower risk for bipolar impairment without decreasing achievement.

Aim: To compare the effectiveness of new and 3-month-old worn brush heads in plaque removal using a rotation-oscillation-powered toothbrush. Material and Methods: A single examiner blinded, randomized, cross-over study was conducted in which 34 adults attended the clinic on two occasions following 48-h periods of no oral hygiene. Following plaque scoring, subjects brushed for 2 min. with either a new brush or a brush they had used for 3 months and plaque was then rescored. Bristle wear was assessed by measuring brushing surface areas on digital images. Results: The mean plaque score percentage reductions with new brush heads were not significantly different from those achieved with 3-month-old heads for total surfaces (new = 38.1%; worn = 37.1%; p = 0.83) and approximal surfaces (new = 31.3%; worn = 30.8%; p = 0.9). Brushing surface area increases of the 3-month-old heads ranged from 0% to 135% (mean 26.9%). Compared with new brush heads, no significant differences were found for plaque score reductions for heads with minor, moderate or marked wear. Conclusion: Rotation-oscillation-powered toothbrushes with 3-month-old brush heads exhibiting various degrees of wear were as effective as new brush heads in plaque removal. Bristle age and wear on a powered toothbrush may not impede the effectiveness of plaque removal.

The authors tested the cognitive vulnerability hypotheses of depression with a retrospective behavioral high-risk design. Individuals without current Axis I diagnoses who exhibited either negative or positive cognitive styles were compared on lifetime prevalence of depressive and other disorders and the clinical parameters of depressive episodes. Consistent with predictions, cognitively high-risk participants had higher lifetime prevalence than low-risk participants of major and hopelessness depression and marginally higher prevalence of minor depression. These group differences were specific to depressive disorders. The high-risk group also had more severe depressions than the low-risk group, but not longer duration or earlier onset depressions. The risk group differences in prevalence of depressive disorders were not mediated by current depressive symptoms.

Recently, we have demonstrated that T30695, a G-tetrad-forming oligonucleotide, is a potent inhibitor of human immunodeficiency virus, type I (HIV-1) integrase and the K(+)-induced loop folding of T30695 plays a key role in the inhibition of HIV-1 integrase (Jing, N., and Hogan, M. E. (1998) J. Biol. Chem. 273, 34992-34999). Here we have modified T30695 by introducing a hydrophobic bulky group, propynyl dU, or a positively charged group, 5-amino dU, into the bases of T residues of the loops, and by substitution of the T-G loops by T-T loops. Physical measurements have demonstrated that the substitution of propynyl dU or 5-amino dU for T in the T residues of the loops did not alter the structure of T30695, and these derivatives also formed an intramolecular G-quartet structure, which is an essential requirement for anti-HIV activity. Measured IC(50) and EC(50) values show that these substitutions did not induce an apparent decrease in the ability to inhibit HIV-1 integrase activity and in the inhibition of HIV-1 replication in cell culture. However, the substitution of T-T loops for T-G loops induced a substantial decrease in both thermal stability and anti-HIV activity. The data analysis of T30695 and the 21 derivatives shows a significant, functional correlation between thermal stability of the G-tetrad structure and the capacity to inhibit HIV-1 integrase activity and between thermal stability of the G-tetrad structure and the capacity to inhibit HIV-1 replication, as assessed with the virus strains HIV-1 RF, IIIB, and MN in cell culture. This relationship between thermostability and activity provides a basis for improving the efficacy of these compounds to inhibit HIV-1 integrase activity and HIV-1 replication in cell culture.

Change in the content and structure of the self-concept was examined in a 2-year prospective study of depression. A self-descriptive card-sorting task measured the self-concepts of participants who were either high or low in cognitive vulnerability to depression at 2 times, once when their mood was low and once when it was not. Analyses examined change in the positive and negative content of the self-concept and in 3 features of self-structure: differential importance, compartmentalization, and self-complexity. Findings suggest that change in the content of the self-concept simply reflects life circumstances, whereas change in self-structure may help to counteract stress and negative mood.

Four species of Helicobacter--H. muridarum, "H. rappini," H. hepaticus, and H. bilis--have been identified in the gastrointestinal tract of rodents. The association of Helicobacter species with chronic gastrointestinal diseases in mice has raised concern about their impact on research results. In this study, different methods for detection of Helicobacter species in the mouse intestinal tract were compared: polymerase chain reaction (PCR) amplification of 16S rRNA gene sequences, bacterial culture, electron microscopy, and histologic examination (Steiner stain). The PCR method was more sensitive in detecting murine Helicobacter species than was culture, electron microscopy, or histologic examination. Of the cecal specimens identified as Helicobacter species-positive by PCR, approximately 60% were identified as positive by each of the other methods. An 87.5% concordance was obtained by PCR screening of DNA from fecal and cecal specimens. Differentiation among murine Helicobacter species by colony morphologic or histologic features was not possible. Scanning electron microscopy and histologic examination indicated greater numbers of helical microorganisms, specifically H. hepaticus, in the cecum than in the colon. These results indicate that the PCR assays used can be performed on feces as a noninvasive means for rapidly screening large numbers of colony mice for murine Helicobacter species.

Mice harboring a targeted disruption of the epidermal growth factor receptor (EGFR) allele exhibit a severely disorganized hair follicle phenotype, fuzzy coat, and systemic disease resulting in death before 3 weeks. This skin phenotype was reproduced in whole skin grafts and in grafts of EGFR null hair follicle buds onto nude mice, providing a model to evaluate the natural evolution of skin lacking the EGFR. Hair follicles in grafts of null skin did not progress from anagen to telogen and scanning electron micrografts revealed wavy, flattened hair fibers with cuticular abnormalities. Many of the EGFR null hair follicles in the grafted skin were consumed by an inflammatory reaction resulting in complete hair loss in 67% of the grafts by 10 weeks. Localization of follicular differentiation markers including keratin 6, transglutaminase, and the hair keratins mHa2 and hacl-1 revealed a pattern of premature differentiation within the null hair follicles. In intact EGFR null mice, proliferation in the interfollicular epidermis, but not hair follicles, was greatly decreased in the absence of EGFR. In contrast, grafting of EGFR null skin resulted in a hyperplastic response in the epidermis that did not resolve even after 10 weeks, although the wound-induced hyperplasia in EGFR wild-type grafts had resolved within 3 to 4 weeks. Thus, epithelial expression of the EGFR has complex functions in the skin. It is important in delaying follicular differentiation, may serve to protect the hair follicle from immunological reactions, and modifies both normal and wound-induced epidermal proliferation but seems dispensable for follicular proliferation.

We have used an artificial DNA bending agent to monitor the local flexibility of the DNA helix as a function of Mg2+ cation concentration, sequence, and temperature. A DNA bending agent was constructed from a pair of triple helix-forming oligonucleotides connected by a flexible polymeric linker, which, when the linker is short enough, causes a bend in a minor groove region separating the two sites of triple helix formation. The unique aspect of this system is that, since the bent region is not in direct contact with the linker or the triple helix-forming oligonucleotides, the free energy reflecting the bendability of the minor helix groove can be estimated from a comparison of binding affinity between the bent and unbent triple helices. A binding competition experiment and association and dissociation kinetic assays executed at 37 degrees C in the presence of 10 mM Mg2+ have revealed an extremely small difference in binding affinity between bent (50 degrees ) and straight triple helices, suggesting that DNA flexibility with respect to minor groove compression is extremely high and virtually independent of the sequence of the distorted duplex. This unexpectedly small difference in binding affinity was detected over the temperature range from 25 to 65 degrees C, and over a Mg2+ concentration range from 0.3 to 10 mM. Thus, these findings provide evidence that DNA bendability for minor groove compression is inherently high and independent of DNA sequence, temperature, or a 30-fold variation of Mg2+ ion concentration.

Preparation of G-rich TFOs containing cholesterol at the end presents challenging problems due to low yield in synthesis and loss in purification due to aggregation. These compounds also have low solubility which can result in variability in their uptake properties and cellular efficacy. To overcome these problems we have designed and synthesized cholesteryl -CPG (6) and -TentaGel (7) supports and phosphoramidite (8) containing a triglycyl linker.

The effect on human immunodeficiency virus 1 (HIV-1) viral transcription and subsequent gene expression mediated by mixed purine-pyrimidine oligodeoxyribonucleotides (oligodeoxynucleotides) designed to form collinear DNA triplexes with purine-rich elements in the viral promoter was evaluated in intact mammalian cell lines (MT4 and U937). Oligonucleotides HIV31 (5'-GTTTTTGGGTGTTGTGGGTGTGTGTGGTTTG-3') and HIV38 (5'-TGGGTGGGGTGGGGTGGGGGGGTGTGGGGTGTGGGGTG-3') were designed to interact with the transcription initiation site (-16 to + 13) and nuclear factor Sp1 binding site (-81 to -44) of HIV-1, respectively. Oligonucleotides, synthesized with a 3' amine blocking group (5'-R-O-PO2-OCH (CHOH)-CH2-NH+3-3') to prevent degradation by cellular nucleases, were readily taken up by MT4 cells from the culture medium, achieving measured intranuclear concentrations higher than the medium in less than 2 h of incubation. The 3' amine modified oligonucleotides were recoverable from the cells after 24 h as greater than 90% intact material. Treatment of acutely infected MT4 cells with either HIV31 or HIV38 significantly inhibited viral-associated cytopathology and P24 antigen production (p less than 0.001). Additionally, inhibition of P24 antigen release, culture supernatant viral titer, and expression of the intact 9.2-kb HIV-1 mRNA was observed when the chronically infected promonocyte cell line, U937, was treated with 10 microM HIV38. Control oligonucleotides with similar base composition did not inhibit virus expression in either cell line. Furthermore, inhibition of viral expression was not due to alpha-interferon induction resulting from oligonucleotide treatment. Both HIV31 and HIV38 associate with their respective DNA target duplexes at micromolar concentrations, and a strong negative ellipticity near 210 nm, characteristic of DNA triplexes, was observed in the circular dichroism spectrum of either target-oligonucleotide complex. These observations suggest that oligonucleotides, designed to form nucleic acid triplexes with specific proviral target sequences, can selectively inhibit transcription of viral mRNA in intact cells and suppress accumulation of viral products.

A synthetic 27-base-long oligodeoxyribonucleotide, termed PU1, has been shown to bind to duplex DNA to form a triplex at a single site within the human c-myc P1 promoter. PU1 has been administered to HeLa cells in culture to examine the feasibility of influencing transcription of the c-myc gene in vivo. It is shown that uptake of PU1 into the nucleus of HeLa cells is efficient and that the compound remains intact for at least 4 hr. In nuclei extracted from PU1-treated cells, inhibition of DNase I cleavage is detected within the c-myc P1 promoter at the target site for triplex formation. The inhibition is shown to be both site and oligodeoxyribonucleotide specific. After cellular uptake of PU1, it is shown that steady-state mRNA arising from the c-myc P1 initiation site is selectively reduced relative to total mRNA, relative to mRNA from the alternative c-myc P2 initiation site, and relative to mRNA derived from the beta-actin promoter. Significant mRNA repression is not seen upon treating cells with oligodeoxyribonucleotides that fail to bind to the P1 promoter target. Taken together, these data suggest that triplex formation can occur between an exogenous oligodeoxyribonucleotide and duplex DNA in the nucleus of treated cells.

Excised cucumber ( Cucumis sativus L. cv 447 Wisconsin SMR 18) cotyledons were sensitive to acifluorfen-methyl (methyl 5-[2-chloro-4-(trifluoromethyl)phenoxy]-2-nitrobenzoate) and MC-15608 (methyl 5-[2-chloro-4-(trifluoromethyl)phenoxy]-2-chlorobenzoate). Injury was detected by monitoring efflux of 3- O -methyl-d-[U- 14 C]glucose from herbicide-treated tissue after exposure to light. Efflux kinetics of 3- O -methyl-[ 14 C]glucose from cotyledons treated with either acifluorfen-methyl (AFM) or MC-15608 were similar. Neither herbicide was active in darkness. Cyclic voltammetry was used to study redox behavior of AFM and MC-15608 in protolytic and aprotic solvents. In protic media reduction of AFM was chemically irreversible. Reduction of MC-15608 in protolytic solvent was not observed. In aprotic solvent, the electrode reaction of AFM was quasireversible. The voltammogram of MC-15608 in aprotic solvent was indicative of a multielectron, completely irreversible electrode reaction. Although the physiological data indicate that AFM and MC-15608 have similar effects in vivo , the redox behavior of the two compounds is quite different. These results suggest that mechanisms involving direct reduction and reoxidation of the diphenyl ether molecule are probably not the basis for the action of these herbicides.

Light-dependent reduction of p -nitro blue tetrazolium chloride (NBT) and 2,6-dichlorophenol-indophenol (DCPIP) was stimulated by nitrofen (2,4-dichlorophenyl p -nitrophenyl ether) in the presence or absence of the photosensitizer riboflavin. Enhancement of NBT reduction occurred at a concentration as low as 17 μ M nitrofen. Nitrofen had no effect on dark reduction of NBT by dithionite, ascorbate, or reduced phenazine methosulfate. NBT reduction proceeded in solution saturated with either air or nitrogen. Stimulation by nitrofen in the presence or absence of riboflavin also occurred independently of oxygen. Superoxide dismutase (superoxide:superoxide oxidoreductase; EC 1.15.1.1) inhibited light-dependent, aerobic reduction of NBT and DCPIP with and without riboflavin. The nitrogen-stimulated component was also eliminated. Thus, it appears nitrofen can effect reduction of NBT (and DCPIP) anaerobically by transferring electrons directly to NBT (and DCPIP) through an oxygen-independent mechanism or aerobically via superoxide radicals.

Benzo[alpha]pyrene diol epoxide (BPDE) is a well-studied environmental carcinogen that binds covalently to DNA. Here we describe a photochemical technique that allows us to map BPDE-binding sites within cloned gene sequences. The technique is based upon our observation that, when irradiated with laser light at 355 nm, one single-strand DNA cut is produced at each BPDE binding site. In initial experiments we have studied the distribution of such cuts in cloned DNA from the chicken adult beta-globin gene. We find that BPDE binding in this gene sequence is distinctly nonrandom. While several prominent BPDE-binding sites are evident, a 300-base-pair sequence immediately 5' to the RNA cap site is most strongly attacked by the carcinogen. This region is believed to contain important transcriptional control sequences. We discuss the possibility that sequence-specific binding to such regulatory elements may be an important feature of the mechanism of the carcinogen.

We have prepared covalent complexes between defined length DNA fragments and a diol epoxide derivative of the carcinogenic polycyclic aromatic hydrocarbon, benzo(a)pyrene. We have studied the structure of these complexes, using transient electric dichroism, circular dichroism, fluorescence quenching, thermal denaturation, polyacrylamide gel electrophoresis, and nuclease digestion. Our observations suggest that th covalently bound carcinogen is intercalated within the helix, forming a wedge-shaped complex. Binding of the carcinogen distorts the structure of the DNA over a region extending beyond the immediate binding site. The most striking aspect of this distortion is that it produces a bend in the helix.

Redox behavior of nitrofen (2,6-dichlorophenyl-p-nitrophenyl ether) at a glassy carbon working electrode in aprotic and protolytic solvents was examined using cyclic voltammetry. The electrode reaction of 2 mM nitrofen in dimethylformamide (0.1 M tetra-n-butylammonium hexafluorophosphate as supporting electrolyte) was quasireversible. Measured half-wave potential was −1.11 V vs saturated calomel reference electrode (SCE) and peak potential separation was 0.110 V. The reduction product was chemically stable on the time scale of the experiment; cathodic peak current equalled anodic peak current. Linear sweep voltammetry employing a rotating, glassy carbon disk electrode indicated that electron stoichiometry for reduction of nitrofen in dimethylformamide and acetonitrile was one. Reduction of nitrofen under aprotic conditions was most likely a one-electron reduction to the nitro radical anion. Reduction of 1 mM nitrofen in 50% ( v v ) ethanol/water buffered at pH 5.0, 7.0, and 10.2 with 0.25 M sodium acetate, 0.1 M potassium phosphate, and 0.1 M sodium carbonate was chemically irreversible and characteristic of an EC … -type reaction mechanism; heterogeneous electron transfer followed by homogeneous chemical reactions. Cathodic peak potentials at pH 5.0 and 7.0 were −0.760 and −0.820 V vs SCE. At pH 10.2 two cathodic peaks were observed at −0.840 and −1.17 V vs SCE. In protolytic solvent at pH 5.0 cathodic peak current varied linearly as a function of the square root of the scan rate. Thus, the electrode reaction was limited by diffusion. Electron stoichiometry for reduction of nitrofen in protolytic solvent was four. The fate of new redox species generated in protolytic solvent at pH 7.0 was examined upon subsequent multiple scans at 200 mV/s. Initial reduction at −0.820 V vs SCE was most likely centered at the nitro group, yielding the four-electron hydroxylamine product. Oxidation at 0.020 V vs SCE was thought to correspond to the two-electron oxidation of hydroxylamine to the nitroso derivative.

Gold labeling of antigenic sites has become an increasingly useful tool in the study of cultured cell monolayers. If these mono-layers are grown on flat substrates, major difficulties in both scanning (SEM) and transmission electron microscopy (TEM) specimen preparation and imaging may result. An alternate surface, that of dextran microcarrier beads, eliminates a majority of these difficulties and facilitates correlative TEM and SEM. The SEM procedure for using backscattered electron imaging requires the use of carbon planchets as the cell growth matrix to eliminate background signals. These planchets are expensive and are not an optimal cell-attachment matrix in that they result in loose and abnormally shaped cells. In contrast, the dextran beads were produced specifically for cell culture and, therefore, provide an excellent surface for growth. The beads have an average diameter of 100 μm, allowing attachment directly to aluminum stubs without signal generation from the aluminum to interfere with the gold signal. With TEM preparation, the monolayer poses the major disadvantage. Specimen preparation for thin sectioning is often preceded by extensive manipulation. In the microcarrier bead system, the beads are directly sectionable, and it is possible to cut five to eight full beads per thin section. This increase in cell surface makes quantification of gold labeling easier and also provides a more representative sampling of the monolayer. The ease of preparation, the decrease in reagents used (via cell pooling), and the ability to use one cell preparation for TEM and SEM make this procedure an ideal technique for gold labeling.

A 27-base-long DNA oligonucleotide was designed that binds to duplex DNA at a single site within the 5' end of the human c-myc gene, 115 base pairs upstream from the transcription origin P1. On the basis of the physical properties of its bound complex, it was concluded that the oligonucleotide forms a colinear triplex with the duplex binding site. By means of an in vitro assay system, it was possible to show a correlation between triplex formation at -115 base pairs and repression of c-myc transcription. The possibility is discussed that triplex formation (site-specific RNA binding to a DNA duplex) could serve as the basis for an alternative program of gene control in vivo.

New SrH band systems photographed at high dispersion include: (1) an abbreviated $C^{2}\ensuremath{\Sigma}\ensuremath{\rightarrow}N^{2}\ensuremath{\Sigma}$ system with (0,0) band head at 3808A, (2) a many-lined $D^{2}\ensuremath{\Sigma}\ensuremath{\rightarrow}N^{2}\ensuremath{\Sigma}$ system between 5000A and 6700A, (3) an $E^{2}\ensuremath{\Pi}\ensuremath{\rightarrow}N^{2}\ensuremath{\Sigma}$ system consisting of a $\ensuremath{\Delta}v=0$ sequence only with the (0,0) band at 5323A. The (0,0) and (0,1) $C$ bands have three violent perturbations and a sharp cut-off due to predissociation at ${K}^{\ensuremath{'}}=19$. The form and magnitude of the perturbations are computed. Quantum analyses give ${B}_{0}=3.930$ and ${D}_{0}=\ensuremath{-}9.31\ifmmode\times\else\texttimes\fi{}{10}^{\ensuremath{-}4}$ for the $C$ state, ${B}_{1}=3.56$ for the normal state. Quantum assignments of the lines of the six principal branches of the (0,0) band of the $E$ system are presented. For the $E$ state $A=117$, ${{B}^{*}}_{0, \ensuremath{-}\frac{1}{2}}=3.6388$, ${{B}^{*}}_{0, +\frac{1}{2}}=3.8687$ and perturbations at small $J$ values distort the $\ensuremath{\Lambda}$-doubling curve for the $^{2}\ensuremath{\Pi}_{\frac{1}{2}}$ state.

The analyses of the $B$ and $C$ band systems of SrD and of a portion of the $D$ system of SrH are presented. $B_{e}^{*}=1.9427$ for SrD and 3.8788 for SrH for the normal $^{2}\ensuremath{\Sigma}$ state. The ratio of these, 0.50635, shows about the expected departure from equality with the ratio ${\ensuremath{\rho}}^{2}=0.50603$ of the reduced masses. The spin doubling of this SrD state is regular, ${\ensuremath{\gamma}}_{0}=+0.0613$ being almost exactly in the ratio ${\ensuremath{\rho}}^{2}$ with the value for SrH. The spin doubling of the $B^{2}\ensuremath{\Sigma}$ state of SrD departs markedly from the usual linear variation with $K+\frac{1}{2}$. Multiple perturbations occur in the (0,0) $C$ band of SrD, and there is a sharp cut-off of the branches at ${K}^{\ensuremath{'}}=29$. For this $C^{2}\ensuremath{\Sigma}$ state ${B}_{0}\ensuremath{\cong}1.95$, but exact analysis is hindered by the lack of any unperturbed lines. Two bands (${v}^{\ensuremath{'}},2$) and (${v}^{\ensuremath{'}},3$), with a common upper state, of the $D^{2}\ensuremath{\Sigma}\ensuremath{\rightarrow}N^{2}\ensuremath{\Sigma}$ "many-lined" SrH system have been analyzed. In this $D$ state, for which ${B}_{v}=1.913$, large irregular perturbations occur. Electronic configurations are discussed.

This study investigated the effectiveness of the Hutt Scoring System (Hutt, 1977) in differentiating organic from functional psychiatric patients, the relationship of IQ to Bender-Gestalt errors as measured by this scoring system, and examined the discriminative power of this scoring system for various IQ ranges. One hundred eighty patients were utilized, 90 of whom had been classified as organics and matched with the remaining 90 subjects on the basis of age, education, and IQ range. In addition, the organic subjects' secondary diagnosis was matched with the primary diagnosis of the nonorganic patients. Results between WAIS IQ and Hutt error scores for organic subjects yielded an r = -.34 (p less than .01); between WAIS IQ and Hutt error scores for the nonorganic subjects, r = -.61 (p less than .001). No significant results were obtained for organics and nonorganics simultaneously within the same IQ cells.

Fructans synthesized from newly formed assimilates accumulate in wheat stems as nonstructural carbohydrates. Experiments performed tested the hypothesis that the fructose moiety from translocated sucrose is used preferentially in biosynthesis of these fructans. Results indicated: (a) a large percentage of labeled sucrose was translocated and unloaded in an unaltered state; and (b) sucrose contributed its fructose moiety to fructan synthesis in stems.

We have prepared covalent complexes between defined length DNA fragments and a diol epoxide derivative of the carcinogenic polycyclic aromatic hydrocarbon, benzo(a)pyrene. We have studied the structure of these complexes, using transient electric dichroism, circular dichroism, fluorescence quenching, thermal denaturation, polyacrylamide gel electrophoresis, and nuclease digestion. Our observations suggest that th covalently bound carcinogen is intercalated within the helix, forming a wedge-shaped complex. Binding of the carcinogen distorts the structure of the DNA over a region extending beyond the immediate binding site. The most striking aspect of this distortion is that it produces a bend in the helix.

We have used a photochemical method to map covalent binding sites of the carcinogen benzo[a]pyrenediol epoxide (BPDE) within DNA from the transcriptional control region of the chicken adult beta-globin gene. Our preliminary low-resolution mapping has demonstrated that this region contains highly preferred BPDE binding sites [Boles, T.C., & Hogan, M.E. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 5623-5627]. Here, we find that BPDE binding at individual G residues in this region is influenced by nearest-neighbor interactions and also by longer range interactions that may be attributable to sequence-specific variation of DNA secondary structure. Our findings suggest that long poly(dG) sequences should be preferred sites for BPDE action in other genes.

As the use of isolated cell cultures increases as an experimental model, there has been a proportional increase in the number of matrices used for cell growth support. These matrices vary in shape, texture and porosity, and as a result, affect the cells grown on them.The ultrastructural characteristics of several of these growth matrices were examined using two cell types chosen for their distinct growth habits. Chinese Hamster Ovary cells and Balb 3T3 Mouse Fibroblasts were grown on flat substrates (glass, tissue culture plastic, Millipore filters) as well as spherical (glass, tissue culture plastic, cross-linked dextran) matrices. Cells were plated maintaining egual densities and growth surface area. Once the majority of the cells reached confluency (approx. one week), the cell morphology on each matrix was examined using scanning electron microscopy and digital analysis of cell attachment area.Variation in cell shape was dramatic between matrices, being most noticeable between a textured surface (filter, dextran bead), and that of a smooth (glass) surface (Figs. 1 and 2). Even within smooth surfaces, some variation was observed.