Analytical model of internally coupled ears.

Lizards and many birds possess a specialized hearing mechanism: internally coupled ears where the tympanic membranes connect through a large mouth cavity so that the vibrations of the tympanic membranes influence each other. This coupling enhances the phase differences and creates amplitude differences in the tympanic membrane vibrations. Both cues show strong directionality. The work presented herein sets out the derivation of a three dimensional analytical model of internally coupled ears that allows for calculation of a complete vibration profile of the membranes. The analytical model additionally provides the opportunity to incorporate the effect of the asymmetrically attached columella, which leads to the activation of higher membrane vibration modes. Incorporating this effect, the analytical model can explain measurements taken from the tympanic membrane of a living lizard, for example, data demonstrating an asymmetrical spatial pattern of membrane vibration. As the analytical calculations show, the internally coupled ears increase the directional response, appearing in large directional internal amplitude differences (iAD) and in large internal time differences (iTD). Numerical simulations of the eigenfunctions in an exemplary, realistically reconstructed mouth cavity further estimate the effects of its complex geometry.

Optimality in mono- and multisensory map formation.

In the struggle for survival in a complex and dynamic environment, nature has developed a multitude of sophisticated sensory systems. In order to exploit the information provided by these sensory systems, higher vertebrates reconstruct the spatio-temporal environment from each of the sensory systems they have at their disposal. That is, for each modality the animal computes a neuronal representation of the outside world, a monosensory neuronal map. Here we present a universal framework that allows to calculate the specific layout of the involved neuronal network by means of a general mathematical principle, viz., stochastic optimality. In order to illustrate the use of this theoretical framework, we provide a step-by-step tutorial of how to apply our model. In so doing, we present a spatial and a temporal example of optimal stimulus reconstruction which underline the advantages of our approach. That is, given a known physical signal transmission and rudimental knowledge of the detection process, our approach allows to estimate the possible performance and to predict neuronal properties of biological sensory systems. Finally, information from different sensory modalities has to be integrated so as to gain a unified perception of reality for further processing, e.g., for distinct motor commands. We briefly discuss concepts of multimodal interaction and how a multimodal space can evolve by alignment of monosensory maps.

Identification of the 'NORE' (N-Oct-3 responsive element), a novel structural motif and composite element.

N-Oct-3 is a neuronal transcription factor widely expressed in the developing mammalian central nervous system, and necessary to maintain neural cell differentiation. The key role of N-Oct-3 in the transcriptional regulation of a multiplicity of genes is primarily due to the structural plasticity of its so-called 'POU' (acronym of Pit, Oct, Unc) DNA-binding domain. We have recently reported about the unusual dual neuro-specific transcriptional regulation displayed by N-Oct-3 [Blaud,M., Vossen,C., Joseph,G., Alazard,R., Erard,M. and Nieto,L. (2004) J. Mol. Biol., 339, 1049-1058]. To elucidate the underlying molecular mechanisms, we have now made use of molecular modeling, DNA footprinting and electrophoretic mobility shift assay techniques. This combined approach has allowed us to uncover a novel mode of homodimerization adopted by the N-Oct-3 POU domain bound to the neuronal aromatic amino acids de-carboxylase and corticotropin-releasing hormone gene promoters and to demonstrate that this pattern is induced by a structural motif that we have termed 'NORE' (N-Oct-3 responsive element), comprising the 14 bp sequence element TNNRTAAATAATRN. In addition, we have been able to explain how the same structural motif can also induce the formation of a heterodimer in association with hepatocyte nuclear factor 3beta(/Forkhead box a2). Finally, we discuss the possible role of the NORE motif in relation to neuroendocrine lung tumor formation, and in particular the development of small cell lung cancer.

Effect of electric field vectoriality on electrically mediated gene delivery in mammalian cells.

Electropermeabilization is a nonviral method used to transfer genes into living cells. Up to now, the mechanism is still to be elucidated. Since cell permeabilization, a prerequired for gene transfection, is triggerred by electric field, its characteristics should depend on its vectorial properties. The present investigation addresses the effect of pulse polarity and orientation on membrane permeabilization and gene delivery by electric pulses applied to cultured mammalian cells. This has been directly observed at the single-cell level by using digitized fluorescence microscopy. While cell permeabilization is only slightly affected by reversing the polarity of the electric pulses or by changing the orientation of pulses, transfection level increases are observed. These last effects are due to an increase in the cell membrane area where DNA interacts. Fluorescently labelled plasmids only interact with the electropermeabilized side of the cell facing the cathode. The plasmid interaction with the electropermeabilized cell surface is stable and is not affected by pulses of reversed polarities. Under such conditions, DNA interacts with the two sites of the cell facing the two electrodes. When changing both the pulse polarity and their direction, DNA interacts with the whole membrane cell surface. This is associated with a huge increase in gene expression. This present study demonstrates the relationship between the DNA/membrane surface interaction and the gene transfer efficiency, and it allows to define the experimental conditions to optimize the yield of transfection of mammalian cells.

Characteristic patterns of N Oct-3 binding to a set of neuronal promoters.

N Oct-3, a neurospecific POU protein, homodimerizes in a non-cooperative fashion on the neuronal aromatic l-amino acid decarboxylase gene promoter and generates heterodimers with HNF-3beta. Several other neuronal gene promoters, the corticotropin releasing hormone and the aldolase C gene promoters also contain overlapping binding sites for N Oct-3 and HNF-3beta. We have demonstrated that N Oct-3 presents a non-cooperative homodimerization on these two additional targets and can also give rise to heterodimers with HNF-3beta. Surprisingly, despite the high degree of conservation of the respective POU subunits, the ubiquitous POU protein Oct-1 can only form monomers even in the presence of either N Oct-3 or HNF-3beta on these DNA targets. Our data indicate that this difference is correlated with the specific ability of a portion of the N Oct-3 linker to fold as an alpha-helix, a property shared by class III POU proteins. These results suggest that this novel binding pattern permits the heterodimerization of N Oct-3 and HNF-3beta on the neuronal promoters, which could be a key issue in the development of the nervous system and possibly tumors of neural origin.

Down-regulation of nuclear receptor DNA-binding activity by nitric oxide--HNF4 as a model system.

The numerous physiological roles of nitric oxide (NO) are currently the focus of intensive research. A major pathway by which NO mediates its biological effects is via S-nitrosylation of cysteine residues, and a growing body of evidence suggests that transcription factors are critical targets for such S-nitrosylation. Here we review the ability of NO to down-regulate the activity of transcription factors, and in particular nuclear receptors. Among the latter, the hepatocyte nuclear factor HNF4 stands out as a key regulator of cytochrome P450 (CYP) gene expression. We report on a series of experiments which show that inflammation-induced NO production decreases CYP mRNA transcription, and that NO suppresses the DNA-binding ability of HNF4. Together these data suggest that cysteine-nitrosylation of the HNF4 DNA-binding domain is the primary molecular mechanism responsible for the drop in oxydase activities of hepatic cytochrome P450 enzymes, and the consequent impairment in drug metabolism during inflammation. In order to discuss this hypothesis from a structural perspective, we have built a homology-derived model of the HNF4 DNA-binding domain and computer-simulated the S-nitrosylation of its cysteine residues. Finally, bearing in mind the structural conservation of the nuclear receptor DNA-binding domain, we discuss to what extent results from HNF4 can be extended to other nuclear receptors.