06-756 Sigma-AldrichAnti-Frizzled 5 Antibody

The lipoglycoproteins of the WNT family act on seven transmembrane-spanning Class Frizzled receptors. Here, we show that WNT-5A evokes a proliferative response in a mouse microglia-like cell line (N13), which is sensitive to pertussis toxin, thus implicating the involvement of heterotrimeric G proteins of the G(i/o) family. We continue to show that WNT-5A stimulation of N13 membranes and permeabilized cells evokes the exchange of GDP for GTP of pertussis toxin-sensitive G proteins employing [?-(35)S]GTP assay and activity state-specific antibodies to GTP-bound G(i) proteins. Our functional analysis of the PTX-sensitivity of WNT-induced G protein activation and PCR analysis of G protein and FZD expression patterns suggest that WNT-5A stimulation leads to the activation of G(i2/3) proteins in N13 cells possibly mediated by FZD(5), the predominant FZD expressed. In summary, we provide for the first time molecular proof that WNT-5A stimulation results in the activation of heterotrimeric G(i2/3) proteins in mammalian cells with physiological protein stochiometry.

The Wnt signaling network is arguably one of the most complex molecular modules by which cells communicate. Though we become accustomed to novel components, the discovery of an unexpected Wnt receptor ligand, Norrin, is surprising even to the connoisseur.

Frizzled genes encode integral membrane proteins that function in multiple signal transduction pathways. They have been identified in diverse animals, from sponges to humans. The family is defined by conserved structural features, including seven hydrophobic domains and a cysteine-rich ligand-binding domain. Frizzled proteins are receptors for secreted Wnt proteins, as well as other ligands, and also play a critical role in the regulation of cell polarity. Frizzled genes are essential for embryonic development, tissue and cell polarity, formation of neural synapses, and the regulation of proliferation, and many other processes in developing and adult organisms; mutations in human frizzled-4 have been linked to familial exudative vitreoretinopathy. It is not yet clear how Frizzleds couple to downstream effectors, and this is a focus of intense study.

Wnt proteins are a large family of secreted glycoproteins. Wnt proteins bind to the Frizzled receptors and LRP5/6 co-receptors, and through stabilizing the critical mediator beta-catenin, initiate a complex signaling cascade that plays an important role in regulating cell proliferation and differentiation. Deregulation of the canonical Wnt/beta-catenin signaling pathway, mostly by inactivating mutations of the APC tumor suppressor, or oncogenic mutations of beta-catenin, has been implicated in colorectal tumorigenesis. Although oncogenic mutations of beta-catenin have only been discovered in a small fraction of non-colon cancers, elevated levels of beta-catenin protein, a hallmark of activated canonical Wnt pathway, have been observed in most common forms of human malignancies, indicating that activation of this pathway may play an important role in tumor development. Over the past 15 years, our understanding of this signaling pathway has significantly improved with the identification of key regulatory proteins and the important downstream targets of beta-catenin/Tcf transactivation complex. Given the fact that Wnt/beta-catenin signaling is tightly regulated at multiple cellular levels, the pathway itself offers ample targeting nodal points for cancer drug development. In this review, we discuss some of the strategies that are being used or can be explored to target key components of the Wnt/beta-catenin signaling pathway in rational cancer drug discovery.

The neural crest is a transient population of multipotent progenitors arising at the lateral edge of the neural plate in vertebrate embryos. After delamination and migration from the neuroepithelium, these cells contribute to a diverse array of tissues including neurons, smooth muscle, craniofacial cartilage, bone cells, endocrine cells and pigment cells. Considerable progress in recent years has furthered our understanding at a molecular level of how this important group of cells is generated and how they are assigned to specific lineages. Here we review a number of recent studies supporting a role for Wnt signaling in neural crest induction, differentiation, and apoptosis. We also summarize the timing of expression of a number of Wnt ligands and receptors with respect to neural crest induction.