Two molecular forms of the human interferon-gamma receptor. Ligand binding, internalization, and down-regulation

The receptors for human interferon-gamma (IFN-gamma) on peripheral blood monocytes and various cells of nonhematopoietic origin were thoroughly characterized and compared. The receptors of all cell types exhibited a similar affinity for IFN-gamma (Kd approximately 1 x 10(-10) M), and in all cases receptor-mediated endocytosis and ligand degradation were demonstrated. However, the receptors differed in their molecular weights (95,000 in HeLa cells and 140,000 in monocytes, assuming a 1:1 ligand to receptor ratio) as concluded from experiments of cross-linking to 125I-IFN-gamma. Lower molecular weight species were obtained as well, particularly in monocytes. Such species could represent either degradation products or subunit structures. The monocyte and HeLa receptor responded differently to an excess of ligand. A significant receptor down-regulation was observed when monocytes were incubated with an excess of 125I-IFN-gamma, whereas no such down-regulation was observed in HeLa cells or in normal fibroblasts. This differential response was observed both in the presence or in the absence of a protein synthesis inhibitor. The receptor on monocytes was found to be acid-labile whereas that on HeLa cells was resistant to acid treatment. These and additional experiments indicate that the monocyte receptor is inactivated following internalization, whereas the HeLa receptor retains its structure and recycles back to the cell surface. The difference in the properties and fate of these two receptor subtypes is probably related to the differential functions of IFN-gamma in various cell types.     

Fat facets does a Highwire act at the synapse.

Neuromuscular synapses are highly dynamic structures that respond to both intercellular and intracellular cues to manipulate synaptic form. A variety of post-translational modifications of synaptic proteins are used to regulate synaptic plasticity. A recent report by DiAntonio et al. shows that two ubiquitin pathway proteins, Highwire and Fat facets, may be mutually antagonistic regulators of presynaptic growth at the Drosophila neuromuscular junction. This work adds support to the emerging idea that ubiquitin, a polypeptide that targets proteins for proteasomal degradation, regulates synaptic development.     

Affect and Cognition in Attitude Formation toward Familiar and Unfamiliar Attitude Objects

At large attitudes are built on earlier experience with the attitude object. If earlier experiences are not available, as is the case for unfamiliar attitude objects such as new technologies, no stored evaluations exist. Yet, people are still somehow able to construct attitudes on the spot. Depending on the familiarity of the attitude object, attitudes may find their basis more in affect or cognition. The current paper investigates differences in reliance on affect or cognition in attitude formation toward familiar and unfamiliar realistic attitude objects. In addition, individual differences in reliance on affect (high faith in intuition) or cognition (high need for cognition) are taken into account. In an experimental survey among Dutch consumers (N = 1870), we show that, for unfamiliar realistic attitude objects, people rely more on affect than cognition. For familiar attitude objects where both affective and cognitive evaluations are available, high need for cognition leads to more reliance on cognition, and high faith in intuition leads to more reliance on affect, reflecting the influence of individually preferred thinking style. For people with high need for cognition, cognition has a higher influence on overall attitude for both familiar and unfamiliar realistic attitude objects. On the other hand, affect is important for people with high faith in intuition for both familiar and unfamiliar attitude objects and for people with low faith in intuition for unfamiliar attitude objects; this shows that preferred thinking style is less influential for unfamiliar objects. By comparing attitude formation for familiar and unfamiliar realistic attitude objects, this research contributes to understanding situations in which affect or cognition is the better predictor of overall attitudes.     

Mutation of Threonine 34 in Mouse Podoplanin-Fc Reduces CLEC-2 Binding and Toxicity in Vivo While Retaining Anti-lymphangiogenic Activity

The lymphatic system plays an important role in cancer metastasis and inhibition of lymphangiogenesis could be valuable in fighting cancer dissemination. Podoplanin (Pdpn) is a small, transmembrane glycoprotein expressed on the surface of lymphatic endothelial cells (LEC). During mouse development, binding of Pdpn to the C-type lectin-like receptor 2 (CLEC-2) on platelets is critical for the separation of the lymphatic and blood vascular systems. Competitive inhibition of Pdpn functions with a soluble form of the protein, Pdpn-Fc, leads to reduced lymphangiogenesis in vitro and in vivo. However, the transgenic overexpression of human Pdpn-Fc in mouse skin causes disseminated intravascular coagulation due to platelet activation via CLEC-2. In the present study, we produced and characterized a mutant form of mouse Pdpn-Fc, in which threonine 34, which is considered essential for CLEC-2 binding, was mutated to alanine (PdpnT34A-Fc). Indeed, PdpnT34A-Fc displayed a 30-fold reduced binding affinity for CLEC-2 compared with Pdpn-Fc. This also translated into fewer side effects due to platelet activation in vivo. Mice showed less prolonged bleeding time and fewer embolized vessels in the liver, when PdpnT34A-Fc was injected intravenously. However, PdpnT34A-Fc was still as active as wild-type Pdpn-Fc in inhibiting lymphangiogenesis in vitro and also inhibited lymphangiogenesis in vivo. These data suggest that the function of Pdpn in lymphangiogenesis does not depend on threonine 34 in the CLEC-2 binding domain and that PdpnT34A-Fc might be an improved inhibitor of lymphangiogenesis with fewer toxic side effects.