A Complex Interplay between Personality Domains,Marital Status and a Variant in CHRNA5 on the Risks of Cocaine,Nicotine Dependences and Cocaine-Induced Paranoia

Background

Personality correlates highly with both cocaine and nicotine dependencies (CD, ND), and their co-morbid psychopathologies. However, little is known about the nature of these relationships. This study examined if environment (marriage) or genetics (a single SNP, CHRNA5*rs16969968) would moderate the correlation of personality with CD, ND and cocaine-induced paranoia (CIP) in African and European Americans (AAs, EAs).

Methods

1432 EAs and 1513 AAs were examined using logistic regression. Personality was assessed by NEO-PI-R, while CD, ND and CIP were diagnosed according to DSM-IV. ND and CD were examined as binary traits and for the analysis of CIP, subjects were divided into 3 groups: (A) Controls with no CIP; (B) CD cases without CIP; and (C) CD cases with CIP. Multiple testing was Bonferroni-corrected.

Results

For CD and ND in the EA population, marital status proved to be a significant moderator in their relationship with openness only (OR = 1.90, 95%CI = 1.36–2.64, p = 1.54e-04 and OR = 2.12, 95%CI = 1.52–2.90, p = 4.65e-06 respectively). For CIP, marriage was observed to moderate its correlation with openness and neuroticism (OR = 1.39, 95%CI = 1.18–1.63, p = 7.64e-04 and OR = 1.26, 95%CI = 1.12–1.42, p = 1.27e-03 respectively). The correlations moderated by rs16969968 were those of conscientiousness and CD (OR = 1.62, 95%CI: 1.23–2.12, p = 8.94e-04) as well as CIP (OR = 1.21, 95%CI: 1.11–1.32, p = 4.93e-04 when comparing group A versus group C). No significant interactions were observed in AA population. The Bonferroni-corrected significance threshold was set to be 1.67e-03.

Conclusion

The role of personality in CD and CIP may be interceded by both environment and genetics, while in ND by environment only.     

The intrinsic neuronal organisation of the nucleus of the basal optic root in the domestic chicken; a light and electron microscopic study using anterograde tracers and postembedding GABA-immunostaining

The intrinsic neuronal organisation in the nucleus of the basal optic root of chickens was investigated. The divergent connections with various areas and the functional complexity of the nucleus require a complex intrinsic structural arrangement. Therefore, an analysis of Golgi impregnated material, ultrastructure, GABA-immunocytochemistry and biotinylated dextran-amine anterograde tracer analysis of the nucleus was carried out. In the Golgi analysis, a characteristic dendritic ramification pattern of two types of putative projection neurons was observed. These neurons form dendritic nests with their overlapping dendritic terminal sections, that develop synaptic fields with the optic fibre terminals. These synaptic fields were confirmed by electron microscopy. GABA-immunopositive terminals synapse with distinct loci of the dendritic trees of projection neurons; they may therefore play an important role in the inhibitory-modulatory system of the nucleus of the basal optic root. The GABA-immunopositive terminals derive from small and/or elongated local circuit neurons which receive retinal afferents, and from myelinated fibres afferents to the nucleus of unknown origin.     

An integrated NAD+-dependent enzyme-functionalized field-effect transistor (ENFET) system: development of a lactate biosensor

An integrated NAD+-dependent enzyme field-effect transistor (ENFET) device for the biosensing of lactate is described. The aminosiloxane-functionalized gate interface is modified with pyrroloquinoline quinone (PQQ) that acts as a catalyst for the oxidation of NADH. Synthetic amino-derivative of NAD+ is covalently linked to the PQQ monolayer. An affinity complex formed between the NAD+/PQQ-assembly and the NAD+-cofactor-dependent lactate dehydrogenase (LDH) is crosslinked and yields an integrated biosensor ENFET-device for the analysis of lactate. Biocatalyzed oxidation of lactate generates NADH that is oxidized by PQQ in the presence of Ca2+-ions. The reduced catalyst, PQQH2, is oxidized by O2 in a process that constantly regenerates PQQ at the gate interface. The biocatalyzed formation of NADH and the O2-stimulated regeneration of PQQ yield a steady-state pH gradient between the gate interface and the bulk solution. The changes in the pH of the solution near the gate interface and, consequently, the gate potential are controlled by the substrate (lactate) concentration in the solution. The device reveals the detection limit of 1 x 10(-4) M for lactate and the sensitivity of 24+/-2 mV dec(-1). The response time of the device is as low as 15 s.     

An explanation for the narrow carbohydrate substrate specificity of adenine phosphoribosyltransferase from Thermus thermophilus from the model of the enzyme,substrate, and magnesium cation cofactor complex

Communicated by Ramaswamy H. Sarma     

Quality of DNA Extracted from Mouthwashes

Background

A cost effective, safe and efficient method of obtaining DNA samples is essential in large scale genetic analyses. Buccal cells are an attractive source of DNA, as their collection is non-invasive and can be carried out by mail. However, little attention has been given to the quality of DNA extracted from mouthwashes.

Methodology

Mouthwash-derived DNA was extracted from 500 subjects participating in a genetic study of high myopia. DNA quality was investigated using two standard techniques: agarose gel electrophoresis and quantitative polymerase chain reaction (qPCR).

Principal Findings

Whereas the majority of mouthwash-derived DNA samples showed a single band of high molecular weight DNA by gel electrophoresis, 8.9% (95% CI: 7.1–10.7%) of samples contained only a smear of low-to-medium molecular weight, degraded DNA. The odds of DNA degradation in a subject''s second mouthwash sample, given degradation of the first, was significantly greater than one (OR = 3.13; 95% CI: 1.22–7.39; Fisher''s test P = 0.009), suggesting that DNA degradation was at least partially a subject-specific phenomenon. Approximately 12.4% (95% CI: 10.4–14.4%) of mouthwash-derived DNA failed to PCR amplify efficiently (using an ∼200 bp microsatellite marker). However, we found there was no significant difference in amplification success rate between DNA samples judged to be degraded or non-degraded by gel electrophoresis (Fisher''s test P = 0.5).

Conclusions

This study demonstrated that DNA degradation affects a significant minority of saline mouthwashes, and that the phenomenon is partially subject-specific. Whilst the level of degradation did not significantly prevent successful amplification of short PCR fragments, previous studies suggest that such DNA degradation would compromise more demanding applications.     

Regulation of the transient receptor potential channel TRPA1 by its N-terminal ankyrin repeat domain

The transient receptor potential channel A1 (TRPA1) is unique among ion channels of higher vertebrates in that it harbors a large ankyrin repeat domain. The TRPA1 channel is expressed in the inner ear and in nociceptive neurons. It is involved in hearing as well as in the perception of pungent and irritant chemicals. The ankyrin repeat domain has special mechanical properties, which allows it to function as a soft spring that can be extended over a large range while maintaining structural integrity. A calcium-binding site has been experimentally identified within the ankyrin repeats. We built a model of the N-terminal 17 ankyrin repeat structure, including the calcium-binding EF-hand. In our simulations we find the calcium-bound state to be rigid as compared to the calcium-free state. While the end-to-end distance can change by almost 50% in the apo form, these fluctuations are strongly reduced by calcium binding. This increase in stiffness that constraints the end-to-end distance in the holo form is predicted to affect the force acting on the gate of the TRPA1 channel, thereby changing its open probability. Simulations of the transmembrane domain of TRPA1 show that residue N855, which has been associated with familial episodic pain syndrome, forms a strong link between the S4-S5 connecting helix and S1, thereby creating a direct force link between the N-terminus and the gate. The N855S mutation weakens this interaction, thereby reducing the communication between the N-terminus and the transmembrane part of TRPA1.     

Biphasic Kinetic Behavior of E. coli WrbA, an FMN-Dependent NAD(P)H:Quinone Oxidoreductase

The E. coli protein WrbA is an FMN-dependent NAD(P)H:quinone oxidoreductase that has been implicated in oxidative defense. Three subunits of the tetrameric enzyme contribute to each of four identical, cavernous active sites that appear to accommodate NAD(P)H or various quinones, but not simultaneously, suggesting an obligate tetramer with a ping-pong mechanism in which NAD departs before oxidized quinone binds. The present work was undertaken to evaluate these suggestions and to characterize the kinetic behavior of WrbA. Steady-state kinetics results reveal that WrbA conforms to a ping-pong mechanism with respect to the constancy of the apparent Vmax to Km ratio with substrate concentration. However, the competitive/non-competitive patterns of product inhibition, though consistent with the general class of bi-substrate reactions, do not exclude a minor contribution from additional forms of the enzyme. NMR results support the presence of additional enzyme forms. Docking and energy calculations find that electron-transfer-competent binding sites for NADH and benzoquinone present severe steric overlap, consistent with the ping-pong mechanism. Unexpectedly, plots of initial velocity as a function of either NADH or benzoquinone concentration present one or two Michaelis-Menten phases depending on the temperature at which the enzyme is held prior to assay. The effect of temperature is reversible, suggesting an intramolecular conformational process. WrbA shares these and other details of its kinetic behavior with mammalian DT-diaphorase, an FAD-dependent NAD(P)H:quinone oxidoreductase. An extensive literature review reveals several other enzymes with two-plateau kinetic plots, but in no case has a molecular explanation been elucidated. Preliminary sedimentation velocity analysis of WrbA indicates a large shift in size of the multimer with temperature, suggesting that subunit assembly coupled to substrate binding may underlie the two-plateau behavior. An additional aim of this report is to bring under wider attention the apparently widespread phenomenon of two-plateau Michaelis-Menten plots.     

Slipknotted and unknotted monovalent cation-proton antiporters evolved from a common ancestor

While the slipknot topology in proteins has been known for over a decade, its evolutionary origin is still a mystery. We have identified a previously overlooked slipknot motif in a family of two-domain membrane transporters. Moreover, we found that these proteins are homologous to several families of unknotted membrane proteins. This allows us to directly investigate the evolution of the slipknot motif. Based on our comprehensive analysis of 17 distantly related protein families, we have found that slipknotted and unknotted proteins share a common structural motif. Furthermore, this motif is conserved on the sequential level as well. Our results suggest that, regardless of topology, the proteins we studied evolved from a common unknotted ancestor single domain protein. Our phylogenetic analysis suggests the presence of at least seven parallel evolutionary scenarios that led to the current diversity of proteins in question. The tools we have developed in the process can now be used to investigate the evolution of other repeated-domain proteins.