Frizzled Proteins are bona fide G Protein-Coupled Receptors

Receptors of the Frizzled family initiate Wnt ligand-dependent signaling controlling
multiple steps in organism development and highly conserved in evolution.
Misactivation of the Wnt/Frizzled signaling is cancerogenic. Frizzled receptors
launch several signaling cascades: the canonical pathway regulating beta-catenin-dependent transcription; the planar cell polarity pathway polarizing the
cytoskeleton within the epithelial plane; and the calcium pathway. Frizzled
receptors possess seven transmembrane domains and their signaling depends on
trimeric G proteins in various organisms. However, Frizzleds constitute a
distinct group within the G protein-coupled receptors (GPCR) superfamily, and
Frizzled signaling can be G protein-independent in some experimental setups, which led to concerns about the GPCR nature of Frizzled. Here we demonstrate
that human Frizzled receptors can directly bind the trimeric Go protein in a
pertussis toxin-sensitive manner. Furthermore, addition of Wnt ligands elicits
Frizzled-dependent guanine nucleotide exchange on Go. An excess of secreted
Frizzled-related protein (a Wnt antagonist) prevents Go activation, as does
pretreatment of Go with pertussis toxin. These experiments provide a biochemical
proof of the GPCR activities of Frizzled receptors and establish an in vitro assay to
monitor Frizzled activation by Wnt ligands, applicable for the high-throughput
agonist/antagonist screening

The sol-gel synthesis of cotton/TiO2 composites and their antibacterial properties

Presentwork is devoted to investigation of structure and functional properties of hybrid nanomaterials based on the TiO2-modified cellulose fibers of cotton. The titania hydrosol was successfully prepared using the titanium tetraisopropoxide as precursor and the nitric acid as peptizing agent via the low-temperature sol–gel synthesis in aqueous medium and applied to cotton fabric. For cross-linking of titania nanoparticles to cotton the 1,2,3,4-butanetetracarboxylic acid (BTCA) was used as a spacer. The morphology and composition of the surface pure and TiO2 modified cotton fibers were investigated by the scanning electron microscopy (SEM). The cotton/TiO2 composite was characterized by the dielectric permittivity. For the estimation of total titania concentration, all samples were calcined at 650 °C. The antimicrobial activity of the treated TiO2 cotton fibers was investigated against Escherichia coli as a model Gram-negative bacteria after exposure to UV-irradiation for 10 mi

Competing Activities of Heterotrimeric G Proteins in Drosophila Wing Maturation

Drosophila genome encodes six alpha-subunits of heterotrimeric G proteins. The Gαs alpha-subunit is involved in the post-eclosion wing maturation, which consists of the epithelial-mesenchymal transition and cell death, accompanied by unfolding of the pupal wing into the firm adult flight organ. Here we show that another alpha-subunit Gαo can specifically antagonize the Gαs activities by competing for the Gβ13F/Gγ1 subunits of the heterotrimeric Gs protein complex. Loss of Gβ13F, Gγ1, or Gαs, but not any other G protein subunit, results in prevention of post-eclosion cell death and failure of the wing expansion. However, cell death prevention alone is not sufficient to induce the expansion defect, suggesting that the failure of epithelial-mesenchymal transition is key to the folded wing phenotypes. Overactivation of Gαs with cholera toxin mimics expression of constitutively activated Gαs and promotes wing blistering due to precocious cell death. In contrast, co-overexpression of Gβ13F and Gγ1 does not produce wing blistering, revealing the passive role of the Gβγ in the Gαs-mediated activation of apoptosis, but hinting at the possible function of Gβγ in the epithelial-mesenchymal transition. Our results provide a comprehensive functional analysis of the heterotrimeric G protein proteome in the late stages of Drosophila wing development

Wnt secretion and gradient formation.

Concentration gradients formed by the lipid-modified morphogens of the Wnt family are known for their pivotal roles during embryogenesis and adult tissue homeostasis. Wnt morphogens are also implicated in a variety of human diseases, especially cancer. Therefore, the signaling cascades triggered by Wnts have received considerable attention during recent decades. However, how Wnts are secreted and how concentration gradients are formed remains poorly understood. The use of model organisms such as Drosophila melanogaster has provided important advances in this area. For instance, we have previously shown that the lipid raft-associated reggie/flotillin proteins influence Wnt secretion and spreading in Drosophila. Our work supports the notion that producing cells secrete Wnt molecules in at least two pools: a poorly diffusible one and a reggie/flotillin-dependent highly diffusible pool which allows morphogen spreading over long distances away from its source of production. Here we revise the current views of Wnt secretion and spreading, and propose two models for the role of the reggie/flotillin proteins in these processes: (i) reggies/flotillins regulate the basolateral endocytosis of the poorly diffusible, membrane-bound Wnt pool, which is then sorted and secreted to apical compartments for long-range diffusion, and (ii) lipid rafts organized by reggies/flotillins serve as "dating points" where extracellular Wnt transiently interacts with lipoprotein receptors to allow its capture and further spreading via lipoprotein particles. We further discuss these processes in the context of human breast cancer. A better understanding of these phenomena may be relevant for identification of novel drug targets and therapeutic strategies

Reggie-1/flotillin-2 promotes secretion of the long-range signalling forms of Wingless and Hedgehog in Drosophila

The lipid-modified morphogens Wnt and Hedgehog diffuse poorly in isolation yet can spread over long distances in vivo, predicting existence of two distinct forms of these mophogens. The first is poorly mobile and activates short-range target genes. The second is specifically packed for efficient spreading to induce long-range targets. Subcellular mechanisms involved in the discriminative secretion of these two forms remain elusive. Wnt and Hedgehog can associate with membrane microdomains, but the function of this association was unknown. Here we show that a major protein component of membrane microdomains, reggie-1/flotillin-2, plays important roles in secretion and spreading of Wnt and Hedgehog in Drosophila. Reggie-1 loss-of-function results in reduced spreading of the morphogens, while its overexpression stimulates secretion of Wnt and Hedgehog and expands their diffusion. The resulting changes in the morphogen gradients differently affect the short- and long-range targets. In its action reggie-1 appears specific for Wnt and Hedgehog. These data suggest that reggie-1 is an important component of the Wnt and Hedgehog secretion pathway dedicated to formation of the mobile pool of these morphogens