In support of these findings, expression of caveolins 1 and 2 in PC12 cells and DRG neurons was also observed by RT-PCR (Fig. treatment with NGF. Robust manifestation of caveolin-1 and -2 is also observed along the entire cell surface of DRG neurons, including high levels on growth cones. These findings demonstrate that neuronal cells communicate caveolins. Caveolae are 50- to 100-nm vesicular organelles that are located at or near the plasma membrane (1C3). It Tetrahydrobiopterin has been proposed that caveolae play a pivotal part in a number of essential cellular functions, including transmission transduction, lipid rate of metabolism, cellular growth control, and apoptotic cell Tetrahydrobiopterin death. The principal protein components of caveolae are the caveolin family of proteins, Rabbit Polyclonal to PAK5/6 (phospho-Ser602/Ser560) termed caveolin-1, -2, and -3 (1, 2). Caveolin-2 shows the same cells distribution as caveolin-1, colocalizes with caveolin-1, and forms a hetero-oligomeric complex with caveolin-1 (4). In contrast, caveolin-3 is definitely a muscle-specific caveolin-related protein that is primarily indicated in striated muscle mass cell types (cardiac and skeletal) (5C7). It has been proposed that caveolin family members function as scaffolding proteins (8) to organize and concentrate specific lipids [cholesterol and glyco-sphingolipids (9C11)] and lipid-modified signaling molecules (9, 12C16) within caveolae membranes. Caveolins interact directly with a number of caveolae-associated signaling molecules, such as H-Ras, hetero-trimeric G-proteins, epidermal growth factor receptor, protein kinase C, Src-family tyrosine kinases, and nitric oxide synthase isoforms (12C14, 17, 18). In many of these instances, it has been recorded that caveolin-binding can efficiently inhibit the enzymatic activity of these signaling molecules formation of caveolae (20). Therefore, down-regulation of caveolin-1 manifestation and caveolae organelles may be crucial to keeping the transformed phenotype. Based on these and additional observations, we as well as others have proposed the caveolae signaling hypothesis, which claims that caveolar localization of particular inactive signaling molecules could provide a compartmental basis for his or her controlled activation and clarify cross-talk between different signaling pathways (1, 21C23). Therefore, we have suggested that caveolin may function as a negative regulator of many different classes of signaling molecules through the acknowledgement of specific caveolin-binding motifs (3, 24). Are these findings relevant to neuronally centered transmission transduction? Caveolin mRNAs and proteins have been shown to be virtually undetectable in mind tissue by Northern and Western blot analyses by several independent investigators (4C7, 25C28), which in the beginning suggested Tetrahydrobiopterin that neurons do not communicate caveolin proteins. In addition, it has been demonstrated that caveolin-1 is not indicated within neuroblastoma cell lines (29). However, this could be secondary to their transformed phenotype, as both caveolin-1 and caveolae are down-regulated in response to cell transformation (19, 20, 30, 31). Here we demonstrate the manifestation of caveolin-1 and -2 in differentiating Personal computer12 cells and dorsal root ganglion (DRG) neurons by using mono-specific antibody probes. In support of our current findings, it has been previously demonstrated that caveolae-like domains can be purified from neuronal cell plasma membranes and that they consist of receptor tyrosine kinases [including insulin and the neurotrophin receptors, Trk-B and p75 nerve growth element (NGF) receptors], as well as other signaling molecules, and the scrapie prion protein (32, 33). However, these investigators were unable to detect the presence of caveolin proteins (32, 33). Also, it has been demonstrated the p75 NGF receptor is definitely associated with caveolae membranes Tetrahydrobiopterin when heterologously indicated Tetrahydrobiopterin in NIH 3T3 cells and is observed to coimmunoprecipitate with caveolin-1 (34). MATERIALS AND METHODS Materials. Antibodies and their sources were as follows: caveolin-1 (pAb N-20; rabbit antipeptide antibody directed.