Localization Of Signaling Proteins Research Articles

Ciliary signaling proteins are mislocalized in the brains of Bardet-Biedl syndrome 1-null mice.

In the brain, primary cilia are found on most, if not all, central neurons. The importance of neuronal cilia is underscored by the fact that human diseases caused by primary cilia dysfunction, which are known as ciliopathies, are associated with neuropathologies, including neuropsychiatric disorders and learning and memory deficits. Neuronal cilia are enriched for certain G protein-coupled receptors and their downstream effectors, suggesting they sense and respond to neuromodulators in the extracellular milieu. GPCR ciliary localization is disrupted in neurons from mouse models of the ciliopathy Bardet-Biedl syndrome, with GPCRs failing to localize to cilia, indicating the Bardet-Biedl syndrome proteins are required for trafficking of G protein-coupled receptors into neuronal cilia. Yet, dopamine receptor 1 accumulates in cilia in the absence of Bardet-Biedl syndrome proteins, suggesting Bardet-Biedl syndrome proteins are required for normal ciliary import and export. To further explore the roles of the Bardet-Biedl syndrome proteins in neuronal cilia, we examined localization of ciliary signaling proteins in a new constitutive Bbs1 knockout mouse model. Interestingly, we find that two additional ciliary G protein-coupled receptors (Gpr161 and Gpr19) abnormally accumulate in cilia on Bardet-Biedl syndrome neurons. In addition, we find that the GPCR signaling protein β-arrestin accumulates in a subset of cilia in the brain, suggesting the presence of additional unidentified ciliary G protein-coupled receptors. These results confirm the importance of the Bardet-Biedl syndrome proteins in establishing ciliary GPCR pathways and indicate that loss of Bbs1 leads to complex changes in the localization of signaling proteins in the brain.

Transport and localization of signaling proteins in ciliated cells

Most cells in the human body elaborate cilia which serve a wide variety of functions, including cell and tissue differentiation during development, sensing physical and chemical properties of the extracellular milieu and mechanical force generation. Common among cilia is the transduction of external stimuli into signals that regulate the activities of the cilia and the cells that possess them. These functions require the transport and localization of specialized proteins to the cilium, a process that many recent studies have shown to be vital for normal cell function and, ultimately, the health of the organism. Here we discuss several mechanisms proposed for the transport and localization of soluble and peripheral membrane proteins to, or their exclusion from the ciliary compartment with a focus on how the structure of the cytoplasm and the size and shape of proteins influence these processes. Additionally, we examine the impact of cell and protein structure on our ability to accurately measure the relative concentrations of fluorescently tagged proteins amongst various cellular domains, which is integral to our understanding of the molecular mechanisms underlying protein localization and transport.

Intracellular Signaling Pathways and Size, Shape, and Rupture History of Human Intracranial Aneurysms

Size and morphological features are associated with intracranial aneurysm (IA) rupture. The cellular mechanisms of IA development and rupture are poorly known. We studied the expression and phosphorylation of different intracellular signaling molecules in the IA wall compared with IA morphological features to understand better the cellular pathways involved in IA development and wall degeneration. Nine ruptured and 17 unruptured human IA samples were collected intraoperatively. The expression levels and phosphorylation state of 3 mitogen-activated protein kinases (c-Jun N-terminal kinase [JNK], p38, extracellular signal-regulated kinase [ERK]), Bcl-2 antagonist of cell death (Bad), mammalian target of rapamycin (mTOR), cyclic AMP response element binding protein (CREB), and Akt were determined by Western blotting. The localization of signaling proteins was determined by immunofluorescence. From 3-dimensional segmentation of computed tomography angiographic data, size and shape indexes were calculated. We found a 5-fold difference in phospho-Bad levels between ruptured and unruptured IAs. Phospho-mTOR was downregulated 2.5-fold in ruptured IAs. Phospho-p54 JNK, phospho-p38, and phospho-Akt levels correlated positively with IA size. Phospho-CREB levels were significantly associated with nonsphericity and ellipticity indexes. Phospho-Akt and phospho-p38 correlated negatively with undulation index. The signaling pathway profile (apoptosis, cell proliferation, stress signaling) differs between ruptured and unruptured IAs and is associated with IA geometry. Our results increase the knowledge of IA development and wall degeneration.

Out of the Caves

Caveolae, "little caves" consisting of flask-shaped invaginations of the plasma membrane, form distinct membrane domains that have been implicated in nitric oxide (NO), insulin, and calcium signal transduction, presumably through localization of signaling proteins in caveolae and their specific interactions with caveolar proteins or lipids. Caveolins are integral membrane proteins that are found in caveolae and are required for caveolar formation in endothelial cells and myocytes. Elimination of caveolins thus interferes with both the formation of caveolae and any specific interactions between caveolins and signaling proteins. Bauer et al. therefore specifically overexpressed caveolin-1 (Cav-1) in mouse vascular endothelium to discriminate between the effects of caveolin per se and that of caveolar existence on endothelial cell signaling and angiogenesis. Cav-1 transgenic mice (Cav-1 TG mice), which showed enhanced expression of both endothelial Cav-1 and Cav-2, showed no increase in caveolae number compared with that in wild-type mice; however, they displayed impaired endothelial nitric oxide synthase (eNOS) signaling. Higher concentrations of acetylcholine were required to dilate aortic rings from Cav-1 TG, and constriction of aortic rings was less responsive to a NOS inhibitor. Vascular endothelial growth factor (VEGF)-stimulated vascular permeability and both ischemia- and VEGF-mediated angiogenesis were attenuated in Cav-1 TG mice, as well as VEGF-mediated phosphorylation of Akt and of eNOS (an Akt substrate). VEGF-mediated phosphorylation of extracellular signal-regulated kinase 1 and 2 (ERK1/2), however, was enhanced. Thus, the authors conclude that Cav-1 likely exerts direct effects on cell signaling independent of its role in caveolar formation. P. M. Bauer, J. Yu, Y. Chan, R. Hickey, P. N. Bernatchez, R. Looft-Wilson, Y. Huang, F. Giordano, R. V. Stan, W. C. Sessa, Endothelial-specific expression of caveolin-1 impairs microvascular permeability and angiogenesis. Proc. Natl. Acad. Sci. U.S.A. 102 , 204-209 (2005). [Abstract] [Full Text]

Multiple spatiotemporal modes of actin reorganization by NMDA receptors and voltage-gated Ca2+ channels.

Cytoskeleton is believed to contribute to activity-dependent processes underlying neuronal plasticity, such as regulations of cellular morphology and localization of signaling proteins. However, how neuronal activity controls actin cytoskeleton remains obscure. Taking advantage of confocal imaging of enhanced GFP-actin in the primary culture of hippocampal neurons, we show that synaptic activity induces multiple types of actin reorganization, both at the spines and at the somatic periphery. Activation of N-methyl-d-aspartate receptors, accompanied with a local rise in [Ca(2+)]i, was sufficient to trigger a slow and sustained recruitment of actin into dendritic spines. In contrast, opening of voltage-gated Ca(2+) channels rapidly and reversibly enhanced cortical actin at the somatic periphery but not in the spines, in keeping with a high transient rise in somatic [Ca(2+)]i. These data suggest that spatiotemporal dynamics of [Ca(2+)]i, triggered by activation of N-methyl-d-aspartate receptors and voltage-gated Ca(2+) channels, provides the molecular basis for activity-dependent actin remodeling.

2P168大腸菌走化性の適応過程におけるシグナル伝達タンパク質の細胞内局在

2P168大腸菌走化性の適応過程におけるシグナル伝達タンパク質の細胞内局在

Cut, Tagged, and Sent Off

Activation of a set of proteases known as caspases is an important part of the cellular signaling mechanisms that lead to cell death or apoptosis. One caspase target is the protein BID, which, after cleavage, translocates from the cytosol to the mitochondrial membrane. There, it interacts with other proteins to influence mitochondrial function and the release of cytochrome c . Zha et al . now provide a surprising mechanism for how cleavage of BID results in mitochondrial targeting. Cleavage of BID exposes an NH 2 -terminal glycine residue, which then undergoes a lipid modification (myristoylation) that enhances its binding to mitochondrial membranes. Furthermore, a mutant that cannot be myristoylated has reduced apoptotic activity. Previously, myristoylation of mammalian proteins has only been observed as a cotranslational modification. Myristoylation may be acutely regulated and may serve as a switch that alters localization of signaling proteins. Zha, J., Weiler, S., Oh, K.J., Wei, M.C., and Korsmeyer, S.J. (2000) Posttranslational N-myristoylation of BID as a molecular switch for targeting mitochondria and apoptosis. Science 290 : 1761-1765. [Abstract] [Full Text]

Cholesterol Depletion Delocalizes Phosphatidylinositol Bisphosphate and Inhibits Hormone-stimulated Phosphatidylinositol Turnover

Caveolae and detergent-insoluble, glycosphingolipid-enriched domains (DIGs) are cholesterol-enriched membrane domains that have been implicated in signal transduction because a variety of signaling proteins as well as phosphatidylinositol bisphosphate (PtdInsP2) are compartmentalized in these domains. We report here that depletion of cellular cholesterol leads to the inhibition of epidermal growth factor- and bradykinin-stimulated PtdIns turnover in A431 cells. This is associated with the loss of compartmentalization of epidermal growth factor receptors, Gq, and PtdInsP2 in the low density membrane domains. Replacement of cellular cholesterol leads to the reorganization of signaling molecules in the low density domains and the reestablishment of hormone-stimulated PtdIns hydrolysis. Oxysterol derivatives show a variable ability to functionally replace the cholesterol in this system. These data are consistent with the hypothesis that localization of signaling proteins and lipids to cholesterol-enriched domains is required for the proper function of hormone-stimulated PtdIns turnover.