Les collagenes du derme: au-dela de leurs proprietes structurales. [Dermis collagens: beyond their structural properties]

The extracellular matrix is a complex network composed of macromolecules such as collagens, proteoglycans and elastin that strongly interact with each other and with cells to maintain the structural integrity of many tissues. These interactions also sustain important cell programs such as migration, proliferation, differentiation and apoptosis. The skin, and more specifically the dermis, contains an extreme diversity of macromolecules that reflects the importance of the composition and organization of the matrix components in providing physical properties and function of the tissues. The most abundant matrix components are the collagens that form a super-family of 27 different members which are divided into different subgroups. The fibrillar collagens, types I, III and V, the FACIT collagens, types XII, XIV and XVI, and collagen VI are all expressed in the collagen-rich dermis. Although the structural features of these collagens are now well characterized, their functions remain elusive. Mutations in human collagen genes give rise to numerous connective tissue diseases including dermis disorders. For example, clinical manifestations in the classical Elhers-Danlos syndrome caused by collagen V gene mutations occur predominantly in the dermis. However, the genotype-phenotype relationship is not clearly established as well as the relation between the distribution and the function of the collagens in dermis. There is no doubt that the ongoing and future work using in vivo approaches will provide new cues regarding the function of collagens in dermis

Definition of a consensus DNA-binding site for the Escherichia coli pleiotropic regulatory protein, FruR.

International audienceThe FruR regulator of Escherichia coli controls the initiation of transcription of several operons encoding a variety of proteins involved in carbon and energy metabolism. The sequence determinants of the FruR-binding site were analysed by using 6x His-tagged FruR and a series of double-stranded randomized oligonucleotides. FruR consensus binding sites were selected and characterized by several consecutive rounds of the polymerase chain reaction-assisted binding-site selection method (BSS) using nitrocellulose-immobilized DNA-binding protein. FruR was demonstrated to require, for binding, an 8 bp left half-site motif and a 3 bp conserved right half-site with the following sequence: 5'-GNNGAATC/GNT-3'. In this sequence, the left half-site AATC/ consensus tetranucleotide is a typical motif of the DNA-binding site of the regulators of the GalR-Lacl family. On the other hand, the high degree of degeneracy found in the right half-site of this palindrome-like structure indicated that FruR, which is a tetramer in solution, interacts asymmetrically with the two half-sites of its operator. However, potentially FruR-target sites showing a high degree of symmetry were detected in 13 genes/operons. Among these, we have focused our interest on the pfkA gene, encoding phosphofructo-kinase-1, which is negatively regulated by FruR

Evolution of the Skin Microstructural Organization During a Mechanical Assay

International audienceSkin is a complex multi-layered tissue, consisting of three main parts: the epidermis, the dermis and the hypodermis. The dermis is responsible for most of the complex mechanical properties of skin, such as viscoelasticity, non-linearity and anisotropy. At the microscopic level the dermis consists for the greater part of extracellular matrix, compounded mainly of collagen fibers forming an orderless network. The mechanical properties of skin have been studied in the past, but their exact link with the microscopic organization is still an open question. The goal of our study is to measure the evolution of the microstructure during a mechanical assay and to improve existing mechanical models of skin with relevant parameters identified at the microscopic level.We perform uniaxial tensile test on ex vivo mouse skin. The mechanical tests are performed in situ under a second harmonic generation microscope. This allows us to determine quantitatively and simultaneously the mechanical response and the microstructural reorganization of the tissue. This technique can be used to better understand the link between pathological alterations of collagen synthesis, fibers organization, and alteration of the biomechanical properties of skin, as in the Ehlers-Danlos syndrome (EDS)

Three-dimensional structure of the DNA-binding domain of the fructose repressor from Escherichia coli by 1H and 15N NMR.

International audienceFruR is an Escherichia coli transcriptional regulator that belongs to the LacI DNA-binding protein family. By using 1H and 15N NMR spectroscopy, we have determined the three-dimensional solution structure of the FruR N-terminal DNA-binding domain consisting of 57 amino acid residues. A total of 809 NMR-derived distances and 54 dihedral angle constraints have been used for molecular modelling with the X-PLOR program. The resulting set of calculated structures presents an average root-mean-square deviation of 0.37 A at the main-chain level for the first 47 residues. This highly defined N-terminal part of the structure reveals a similar topology for the three alpha-helices when compared to the 3D structures of LacI and PurR counterparts. The most striking difference lies in the connection between helix II and helix III, in which three additional residues are present in FruR. This connecting segment is well structured and contains a type III turn. Apart from hydrophobic interactions of non-polar residues with the core of the domain, this connecting segment is stabilised by several hydrogen bonds and by the aromatic ring stacking between Tyr19 of helix II and Tyr28 of the turn. The region containing the putative "hinge helix" (helix IV), that has been described in PurR-DNA complex to make specific base contacts in the minor groove of DNA, is unfolded. Examination of hydrogen bonds highlights the importance of homologous residues that seem to be conserved for their ability to fulfill helix N and C-capping roles in the LacI repressor family

Multiscale characterization of skin mechanics through in situ imaging

International audienceThe complex mechanical properties of skin have been studied intensively over the past decades. They are intrinsically linked to the structure of the skin at several length scales, from the macroscopic layers (epidermis, dermis and hypodermis) down to the microstructural organization at the molecular level. Understanding the link between this microscopic organization and the mechanical properties is of significant interest in the cosmetic and medical fields. Nevertheless, it only recently became possible to directly visualize the skin’s microstructure during mechanical assays, carried out on the whole tissue or on isolated layers. These recent observations have provided novel information on the role of structural components of the skin in its mechanical properties, mainly the collagen fibers in the dermis, while the contribution of others, such as elastin fibers, remains elusive. In this chapter we present current methods used to observe skin’s microstructure during a mechanical assay, along with their strengths and limitations, and we review the unique information they provide on the link between structure and function of the skin