Disruption of the langerin/CD207 gene abolishes Birbeck granules without a marked loss of Langerhans cell function

Langerin is a C-type lectin expressed by a subset of dendritic leukocytes, the Langerhans cells (LC). Langerin is a cell surface receptor that induces the formation of an LC-specific organelle, the Birbeck granule (BG). We generated a langerin(-/-) mouse on a C57BL/6 background which did not display any macroscopic aberrant development. In the absence of langerin, LC were detected in normal numbers in the epidermis but the cells lacked BG. LC of langerin(-/-) mice did not present other phenotypic alterations compared to wild-type littermates. Functionally, the langerin(-/-) LC were able to capture antigen, to migrate towards skin draining lymph nodes, and to undergo phenotypic maturation. In addition, langerin(-/-) mice were not impaired in their capacity to process native OVA protein for I-A(b)-restricted presentation to CD4(+) T lymphocytes or for H-2K(b)-restricted cross-presentation to CD8(+) T lymphocytes. langerin(-/-) mice inoculated with mannosylated or skin-tropic microorganisms did not display an altered pathogen susceptibility. Finally, chemical mutagenesis resulted in a similar rate of skin tumor development in langerin(-/-) and wild-type mice. Overall, our data indicate that langerin and BG are dispensable for a number of LC functions. The langerin(-/-) C57BL/6 mouse should be a valuable model for further functional exploration of langerin and the role of BG.     

Evolution of Plant-Like Crystalline Storage Polysaccharide in the Protozoan Parasite Toxoplasma gondii Argues for a Red Alga Ancestry

Single-celled apicomplexan parasites are known to cause major diseases in humans and animals including malaria, toxoplasmosis, and coccidiosis. The presence of apicoplasts with the remnant of a plastid-like DNA argues that these parasites evolved from photosynthetic ancestors possibly related to the dinoflagellates. Toxoplasma gondii displays amylopectin-like polymers within the cytoplasm of the dormant brain cysts. Here we report a detailed structural and comparative analysis of the Toxoplasma gondii, green alga Chlamydomonas reinhardtii, and dinoflagellate Crypthecodinium cohnii storage polysaccharides. We show Toxoplasma gondii amylopectin to be similar to the semicrystalline floridean starch accumulated by red algae. Unlike green plants or algae, the nuclear DNA sequences as well as biochemical and phylogenetic analysis argue that the Toxoplasma gondii amylopectin pathway has evolved from a totally different UDP-glucose-based metabolism similar to that of the floridean starch accumulating red alga Cyanidioschyzon merolae and, to a lesser extent, to those of glycogen storing animals or fungi. In both red algae and apicomplexan parasites, isoamylase and glucan–water dikinase sequences are proposed to explain the appearance of semicrystalline starch-like polymers. Our results have built a case for the separate evolution of semicrystalline storage polysaccharides upon acquisition of photosynthesis in eukaryotes.This article contains online-only supplementary material.Reviewing Editor:Dr. Patrick Keeling     

Flash photolysis using a light emitting diode: an efficient, compact, and affordable solution

Flash photolysis has become an essential technique for dynamic investigations of living cells and tissues. This approach offers several advantages for instantly changing the concentration of bioactive compounds outside and inside living cells with high spatial resolution. Light sources for photolysis need to deliver pulses of high intensity light in the near UV range (300-380 nm), to photoactivate a sufficient amount of molecules in a short time. UV lasers are often required as the light source, making flash photolysis a costly approach. Here we describe the use of a high power 365 nm light emitting diode (UV LED) coupled to an optical fiber to precisely deliver the light to the sample. The ability of the UV LED light source to photoactivate several caged compounds (CMNB-fluorescein, MNI-glutamate, NP-EGTA, DMNPE-ATP) as well as to evoke the associated cellular Ca(2+) responses is demonstrated in both neurons and astrocytes. This report shows that UV LEDs are an efficient light source for flash photolysis and represent an alternative to UV lasers for many applications. A compact, powerful, and low-cost system is described in detail.     

Mechanism of force generation of a viral DNA packaging motor

A large family of multimeric ATPases are involved in such diverse tasks as cell division, chromosome segregation, DNA recombination, strand separation, conjugation, and viral genome packaging. One such system is the Bacillus subtilis phage phi 29 DNA packaging motor, which generates large forces to compact its genome into a small protein capsid. Here we use optical tweezers to study, at the single-molecule level, the mechanism of force generation in this motor. We determine the kinetic parameters of the packaging motor and their dependence on external load to show that DNA translocation does not occur during ATP binding but is likely triggered by phosphate release. We also show that the motor subunits act in a coordinated, successive fashion with high processivity. Finally, we propose a minimal mechanochemical cycle of this DNA-translocating ATPase that rationalizes all of our findings.     

Foxl2 gene and the development of the ovary: a story about goat, mouse, fish and woman

In this review, we describe recent results concerning the genetics of sex determination in mammals. Particularly, we developed the study of the FOXL2 gene and its implication in genetic anomalies in goats (PIS mutation) and humans (BPES). We present the expression of FOXL2 in the ovaries of different species.     

Notch1 is essential for postnatal hair follicle development and homeostasis

Notch genes encode evolutionarily conserved large, single transmembrane receptors, which regulate many cell fate decisions and differentiation processes during fetal and postnatal life. Multiple Notch receptors and ligands are expressed in both developing and adult epidermis and hair follicles. Proliferation and differentiation of these two ectodermal-derived structures have been proposed to be controlled in part by the Notch pathway. Whether Notch signaling is involved in postnatal hair homeostasis is currently unknown. Here, we investigate and compare the role of the Notch1 receptor during embryonic hair follicle development and postnatal hair homeostasis using Cre-loxP based tissue specific and inducible loss-of-function approaches. During embryonic development, tissue-specific ablation of Notch1 does not perturb formation and patterning of hair follicle placodes. However, Notch1 deficient hair follicles invaginate prematurely into the dermis. Embryonic as well as postnatal inactivation of Notch1 shortly after birth or in adult mice results in almost complete hair loss followed by cyst formation. The first hair cycle of Notch1 deficient mice is characterized by shortened anagen and a premature entry into catagen. These data show that Notch1 is essential for late stages of hair follicle development during embryogenesis as well as for post-natal hair follicle development and hair homeostasis.     

Inhibition of endothelial cell movement and tubulogenesis by human recombinant soluble melanotransferrin: involvement of the u-PAR/LRP plasminolytic system

We have previously demonstrated that human recombinant soluble melanotransferrin (hr-sMTf) interacts with the single-chain zymogen pro urokinase-type plasminogen activator (scu-PA) and plasminogen. In the present work, the impact of exogenous hr-sMTf on endothelial cells (EC) migration and morphogenic differentiation into capillary-like structures (tubulogenesis) was assessed. hr-sMTF at 10 nM inhibited by 50% the migration and tubulogenesis of human microvessel EC (HMEC-1). In addition, in hr-sMTf-treated HMEC-1, the expression of both urokinase-type plasminogen activator receptor (u-PAR) and low-density lipoprotein receptor-related protein (LRP) are down-regulated. However, fluorescence-activated cell sorting analysis revealed a 25% increase in cell surface u-PAR in hr-sMTf-treated HMEC-1, whereas the binding of the urokinase-type plasminogen activator (u-PA)*plasminogen activator inhibitor-1 (PAI-1) complex is decreased. This reduced u-PA-PAI-1 binding is correlated with a strong inhibition of the HMEC-1 plasminolytic activity, indicating that exogenous hr-sMTf treatment alters the internalization and recycling processes of free and active u-PAR at the cellular surface. Overall, these results demonstrate that exogenous hr-sMTf affects plasminogen activation at the cell surface, thus leading to the inhibition of EC movement and tubulogenesis. These results are the first to consider the potential use of hr-sMTf as a possible therapeutic agent in angiogenesis-related pathologies.     

Effect of hyperosmotic shock on phosphoenolpyruvate carboxykinase gene expression and gluconeogenic activity in the crab muscle

Chasmagnathus granulata phosphoenolpyruvate carboxykinase (PEPCK) cDNA from jaw muscle was cloned and sequenced, showing a specific domain to bind phosphoenolpyruvate in addition to the kinase-1 and kinase-2 motifs to bind guanosine triphosphate (GTP) and Mg(2+), respectively, specific for all PEPCKs. In the kinase-1 motifs the GK was changed to RK. The first 19 amino acids of the putative enzyme contain hydrophobic amino acids and hydroxylated residues specific to a mitochondrial type signal. The PEPCK is expressed in hepatopancreas, muscles, nervous system, heart, and gills. Hyperosmotic stress for 24 h increased the PEPCK mRNA level, gluconeogenic and PEPCK activities in muscle.