Systematic screening for skin, hair, and nail abnormalities in a large-scale knockout mouse program

The International Knockout Mouse Consortium was formed in 2007 to inactivate (“knockout”) all protein-coding genes in the mouse genome in embryonic stem cells. Production and characterization of these mice, now underway, has generated and phenotyped 3,100 strains with knockout alleles. Skin and adnexa diseases are best defined at the gross clinical level and by histopathology. Representative retired breeders had skin collected from the back, abdomen, eyelids, muzzle, ears, tail, and lower limbs including the nails. To date, 169 novel mutant lines were reviewed and of these, only one was found to have a relatively minor sebaceous gland abnormality associated with follicular dystrophy. The B6N(Cg)-Far2tm2b(KOMP)Wtsi/2J strain, had lesions affecting sebaceous glands with what appeared to be a secondary follicular dystrophy. A second line, B6N(Cg)-Ppp1r9btm1.1(KOMP)Vlcg/J, had follicular dystrophy limited to many but not all mystacial vibrissae in heterozygous but not homozygous mutant mice, suggesting that this was a nonspecific background lesion. We discuss potential reasons for the low frequency of skin and adnexal phenotypes in mice from this project in comparison to those seen in human Mendelian diseases, and suggest alternative approaches to identification of human disease-relevant models.This work was supported by grants from the National Institutes of Health (R21-AR063781 and U42-OD011185). Shared services at The Jackson Laboratory are subsidized by a National Cancer Institute Core Grant (P30-CA034196). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Are giraffes pollinators or flower predators of Acacia nigrescens in Kruger National Park, South Africa?

We examined the relationship between giraffes (Giraffa camelopardalis) and Acacia nigrescens in Kruger National Park, South Africa, to determine whether these tall ungulates may be providing a pollination service for the trees, or are simply flower predators. We quantified florivory and subsequent fruit set in the presence and absence of giraffes. Acacia nigrescens flowers are clearly a substantial dietary component for giraffes. Although A. nigrescens flowers contain almost three times as much condensed tannin as leaves, giraffes consume large quantities of flowers (∼85% of flowers within reach), resulting in distinct browse lines on the trees. This substantial florivory is detrimental to the overall fecundity of A. nigrescens, with significantly reduced fruit set at heights on trees that are accessible to giraffes. Fruit set increased above the reach of giraffes, suggesting successful pollination by insects. Giraffes were effectively flower predators of A. nigrescens in the season we examined

An introduction to the theory of higher-dimensional quasiconformal mappings

This book offers a modern, up-to-date introduction to quasiconformal mappings from an explicitly geometric perspective, emphasizing both the extensive developments in mapping theory during the past few decades and the remarkable applications of geometric function theory to other fields, including dynamical systems, Kleinian groups, geometric topology, differential geometry, and geometric group theory. It is a careful and detailed introduction to the higher-dimensional theory of quasiconformal mappings from the geometric viewpoint, based primarily on the technique of the conformal modulus of a curve family. Notably, the final chapter describes the application of quasiconformal mapping theory to Mostow's celebrated rigidity theorem in its original context with all the necessary background. This book will be suitable as a textbook for graduate students and researchers interested in beginning to work on mapping theory problems or learning the basics of the geometric approach to quasiconformal mappings. Only a basic background in multidimensional real analysis is assumed

Serotonin type 4 receptor dimers

Chapitre 7International audienceNumerous class A G protein-coupled receptors and especially biogenic amine receptors have been reported to form homodimers. Indeed, the dimerization process might occur for all the metabotropic serotonergic receptors. Moreover, dimerization appears to be essential for the function of serotonin type 2C (5-HT2C) and type 4 (5-HT4) receptors and required to obtain full receptor activity. Several techniques have been developed to analyze dimer formation and properties. Due to our involvement in deciphering 5-HT4R transduction mechanisms, we improved and set up new procedures to study 5-HT4R dimers, by classical methods or modern tools. This chapter presents detailed protocols to detect 5-HT4R dimers by western blotting and co-immunoprecipitation, including the optimizations that we routinely carry out. We developed an innovative method to achieve functional visualization of 5-HT4R dimers by immunofluorescence, taking advantage of the 5-HT4-RASSL (Receptor Activated Solely by Synthetic Ligand) mutant that was engineered in the laboratory. Finally, we adapted the powerful Time-resolved FRET technology to assess a relative quantification of dimer formation and affinit

Pharmacological profile of engineered 5-HT4 receptors and identification of 5-HT4 receptor-biased ligands

International audienceG protein-coupled receptors (GPCRs) can activate simultaneously multiple signaling pathways upon agonist binding. The combined use of engineered GPCRs, such as the Receptors Activated Solely by Synthetic Ligands (RASSLs), and of biased ligands that activate only one pathway at a time might help deciphering the physiological role of each G protein signaling. In order to find serotonin type 4 receptor (5-HT4R) biased ligands, we analyzed the ability of several compounds to activate the Gs and Gq/11 pathways in COS-7 cells that transiently express wild type 5-HT4R, the 5-HT4R-D 100 A mutant (known also as 5-HT4-RASSL, or Rs1) or the 5-HT4R-T 104 A mutant, which modifies agonist-induced 5-HT4R activation. This analysis allowed completing the pharmacological profile of the two mutant 5-HT4Rs, but we did not find any biased ligand for the mutant receptors. Conversely, we identified the first biased agonists for wild type 5-HT4R. Indeed, RS 67333 and prucalopride acted as partial agonists to induce cAMP accumulation, but as antagonists on inositol phosphate production. Moreover, they showed very different antagonist potencies that could be exploited to study the activation of the Gs pathway, with or without concomitant block of Gq/11 signaling

Extreme Features of the Galdieria sulphuraria Organellar Genomes: A Consequence of Polyextremophily?

Nuclear genome sequencing from extremophilic eukaryotes has revealed clues about the mechanisms of adaptation to extreme environments, but the functional consequences of extremophily on organellar genomes are unknown. To address this issue, we assembled the mitochondrial and plastid genomes from a polyextremophilic red alga, Galdieria sulphuraria strain 074 W, and performed a comparative genomic analysis with other red algae and more broadly across eukaryotes. The mitogenome is highlyreduced in size and genetic content and exhibits the highest guanine–cytosine skew of any known genome and the fastest substitution rate among all red algae. The plastid genome contains a large number of intergenic stem-loop structures but is otherwise rather typical in size, structure, and content in comparison with other red algae. We suggest that these unique genomic modifications result not only from the harsh conditions in which Galdieria lives but also from its unusual capability to grow heterotrophically, endolithically, and in the dark. These conditions place additional mutational pressures on the mitogenome due to the increased reliance on the mitochondrion for energy production, whereas the decreased reliance on photosynthesis and the presence of numerous stem-loop structures may shield the plastome from similar genomic stress

Conformational Toggle Switches Implicated in Basal Constitutive and Agonist-Induced Activated States of 5-Hydroxytryptamine-4 Receptors

International audienceThe extended classic ternary complex model predicts that a G protein-coupled receptor (GPCR) exists in only two interconvertible states: an inactive R, and an active R(*). However, different structural active R(*) complexes may exist in addition to a silent inactive R ground state (Rg). Here we demonstrate, in a cellular context, that several R(*) states of 5-hydroxytryptamine-4 (5-HT(4)) receptors involve different side-chain conformational toggle switches. Using site-directed mutagenesis and molecular modeling approaches, we show that the basal constitutive receptor (R(*)basal) results from stabilization of an obligatory double toggle switch (Thr3.36 from inactive g- to active g+ and Trp6.48 from inactive g+ to active t). Mutation of either threonine or tryptophan to alanine resulted in a lowering of the activity of the R(*)basal similar to the Rg. The T3.36A mutation shows that the Thr3.36 toggle switch plays a minor role in the stabilization of R(*) induced by 5-HT (R(*)-5-HT) and BIMU8 (R(*)-BIMU8) and is fully required in the stabilization of R(*) induced by (S)-zacopride, cisapride, and 1-(4-amino-5-chloro-2-methoxyphenyl)-3-(1-butyl-4-piperidinyl)-1-propanone (RS 67333) (R(*)-benzamides). Thus, benzamides stabilize R(*)-benzamides by forming a specific hydrogen bond with Thr3.36 in the active g+ conformation. Conversely, R(*)-BIMU8 was probably the result of a direct conformational transition of Trp6.48 from inactive g+ to active t by hydrogen bonding of this residue to a carboxyl group of BIMU8. We were surprised that the Trp6.48 toggle switch was not necessary for receptor activation by the natural agonist 5-HT. R(*)-5-HT is probably attained through other routes of activation. Thus, different conformational arrangements occur during stabilization of R(*)basal, R(*)-5-HT, R(*)-benzamides, and R(*)-BIMU8