Comparative haploid genetic screens reveal divergent pathways in the biogenesis and trafficking of glycophosphatidylinositol (GPI)-anchored proteins
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE68796 | GEO | 2015/05/13
SECONDARY ACCESSION(S): PRJNA283840
REPOSITORIES: GEO
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Project description:Tissue inhibitor of metalloproteinase 1 (TIMP-1) controls matrix metalloproteinase (MMP) activity through 1:1 stochiometric binding. Human TIMP-1 fused to a glycosylphosphatidylinositol (GPI) anchor (TIMP-1-GPI) shifts the activity of TIMP-1 from the extracellular matrix to the cell surface. TIMP-1-GPI treated renal cell carcinoma cells (RCC) show increased apoptosis and reduced proliferation. Transcriptomic profiling and regulatory pathway mapping were used to identify potential mechanisms driving these effects. Significant changes in inhibitor of DNA binding (IDs), TGF-β1/SMAD and BMP pathways resulted from TIMP-1-GPI treatment. These events were linked to reduced TGF-β1 signaling mediated by inhibition of proteolytic processing of latent TGF-β1 by TIMP-1-GPI. Activity of TIMP-1 from the extracellular matrix to the cell surface. TIMP-1-GPI treated renal cell carcinoma cells (RCC) show increased apoptosis and reduced proliferation. Transcriptomic profiling and regulatory pathway mapping were used to identify potential mechanisms driving these effects. Significant changes in inhibitor of DNA binding (IDs), TGF-β1/SMAD and BMP pathways resulted from TIMP-1-GPI treatment. These events were linked to reduced TGF-β1 signaling mediated by inhibition of proteolytic processing of latent TGF-β1 by TIMP-1-GPI. Renal cell carcinoma cells were transfected with empty vector, rhTimp1 and 2 concentrations of Timp1-GPI fusion protein
2012-08-23 | E-GEOD-40301 | biostudies-arrayexpress
Project description:GPI anchors many proteins to the cell surface. GPI precursor has three mannoses, all of which are modified by ethanolamine-phosphate (EthN-P). It has been believed that EthN-P on the third mannose is always used as a bridge to the protein and EthN-P on the second mannose is removed after GPI is attached to the protein. Based on the finding that PIGB-knockout cells lacking the third mannose and PIGO-knockout cells lacking EthN-P linked to the third mannose expressed low levels of CD59 and DAF, GPI-APs and that those residual GPI-APs were lost by further knockout of PIGG. Using mass-spectrometry we determined that CD59 expressed on PIGB-knockout cells is attached to GPI via EthN-P linked to the second mannose. This linkage structure is also found in wild type cells. Our data modifies the current view of GPI anchors and provides mechanistic basis of inherited GPI deficiency caused by PIGG mutations.
2022-05-31 | PXD021996 | Pride
Project description:Tissue inhibitor of metalloproteinase 1 (TIMP-1) controls matrix metalloproteinase (MMP) activity through 1:1 stochiometric binding. Human TIMP-1 fused to a glycosylphosphatidylinositol (GPI) anchor (TIMP-1-GPI) shifts the activity of TIMP-1 from the extracellular matrix to the cell surface. TIMP-1-GPI treated renal cell carcinoma cells (RCC) show increased apoptosis and reduced proliferation. Transcriptomic profiling and regulatory pathway mapping were used to identify potential mechanisms driving these effects. Significant changes in inhibitor of DNA binding (IDs), TGF-β1/SMAD and BMP pathways resulted from TIMP-1-GPI treatment. These events were linked to reduced TGF-β1 signaling mediated by inhibition of proteolytic processing of latent TGF-β1 by TIMP-1-GPI. Activity of TIMP-1 from the extracellular matrix to the cell surface. TIMP-1-GPI treated renal cell carcinoma cells (RCC) show increased apoptosis and reduced proliferation. Transcriptomic profiling and regulatory pathway mapping were used to identify potential mechanisms driving these effects. Significant changes in inhibitor of DNA binding (IDs), TGF-β1/SMAD and BMP pathways resulted from TIMP-1-GPI treatment. These events were linked to reduced TGF-β1 signaling mediated by inhibition of proteolytic processing of latent TGF-β1 by TIMP-1-GPI.
2012-08-23 | GSE40301 | GEO
Project description:GPI-anchored proteins (GPI-APs) are an important class of glycoproteins that are tethered to the surface of mammalian cells via the lipid glycosylphosphatidylinositol (GPI). GPI-APs have been implicated in many important cellular functions including cell adhesion, cell signaling, and immune regulation. Proteomic identification of mammalian GPI-APs en masse has been limited technically by poor sensitivity for these low abundance proteins and the use of methods that destroys cell integrity. Here we present methodology that permits identification of GPI-APs liberated directly from the surface of intact mammalian cells through exploitation of their appended glycans to enrich for these proteins ahead of LC-MS/MS analyses. We validate our approach in HeLa cells, identifying a greater number of GPI-APs from intact cells than has been previously identified from isolated HeLa membranes and a lipid raft preparation. We further apply our approach to define the cohort of endogenous GPI-APs that populate the distinct apical and basolateral membrane surfaces of polarized epithelial cell monolayers. Our approach provides a new method to achieve greater sensitivity in the identification of low abundance GPI-APs from the surface of live cells and the non-destructive nature of the method provides new opportunities for the temporal or spatial analysis of cellular GPI-AP expression and dynamics.
2016-07-07 | PXD001130 | Pride
Project description:To determine GPI structure of human CD59 in wild-type,B3GALT4-KO, PIGZ-B3GALT4-DKO HEK293 cells.
2020-02-17 | PXD014226 | Pride
Project description:To determine GPI structure of human CD59 in PGAP4-KO, PGAP3-KO, PGAP5-KO, and PGAP2-KO CHO cells.
2018-01-30 | PXD008230 | Pride
Project description:GPI anchors many proteins to the cell surface. GPI precursor has three mannoses, all of which are modified by ethanolamine-phosphate (EthN-P). It has been believed that EthN-P on the third mannose is always used as a bridge to the protein and EthN-P on the second mannose is removed after GPI is attached to the protein. In fact, several GPI-anchored proteins are not appreciably reduced on cells defective in PIGG, which transfers EthN-P to the second mannose. Nevertheless, mutations in PIGG cause neuronal abnormalities. Here, we show that EthN-P on the second mannose is used as a preferential bridge for several GPI-anchored proteins. Our data modifies the current view of GPI anchors and provides mechanistic basis of PIGG deficiencies.
2022-05-31 | PXD022032 | Pride
Project description:Lipid rafts are membrane microdomains rich in cholesterol, sphingolipids, glycosylphosphatidylinositol-anchored proteins (GPI-APs), and receptors. They are localized at the plasma membrane and are essential for signal transmission and organogenesis. However, few reports on the specific effects of lipid rafts on organogenesis have been reported. Here, we focused on the odontoblast differentiation that secretes the dentin matrix and generates dentin. We found the GPI-AP, lymphocyte antigen-6 (Ly6)/Plaur domain-containing1 (Lypd1), specifically expressed in the dental papilla, especially preodontoblasts, using single-cell RNA sequence (scRNA-seq). Depletion experiments of lipid rafts and Lypd1 using ex vivo culture and cell cultures showed that differentiation of tooth germ and dental mesenchymal cells was inhibited. Furthermore, the C-terminal structural domain containing the omega site, which is necessary to localize the plasma membrane, of Lypd1 is necessary for odontoblast differentiation and the morphological change like odontoblasts. Stimulating downstream Bone morphogenetic protein (Bmp) signaling by BMP2 caused it to activate the phosphorylation of Smad1/5/8 and promote the expression of odontoblast differentiation markers such as Pannexin 3 (Panx3), Alkaline phosphatase (Alp), and Dentin sialophosphoprotein (Dspp). In contrast, suppression of Lypd1 inhibited the phosphorylation of Smad1/5/8. In addition, the expression of Lypd1 predates any other marker of odontoblast differentiation previously described. That implies that Lypd1 could be employed as a novel earlier differentiation marker of odontoblasts. These results suggest that Lypd1 as GPI-AP of lipid rafts is important for tooth organogenesis and plays a pivotal role in odontoblast differentiation by regulating phosphorylation of Smad1/5/8.
2023-08-25 | GSE206135 | GEO
Project description:GPI anchors many proteins to the cell surface. GPI precursor has three mannoses, all of which are modified by ethanolamine-phosphate (EthN-P). It has been believed that EthN-P on the third mannose is always used as a bridge to the protein and EthN-P on the second mannose is removed after GPI is attached to the protein. In fact, several GPI-anchored proteins are not appreciably reduced on cells defective in PIGG, which transfers EthN-P to the second mannose. Nevertheless, mutations in PIGG cause neuronal abnormalities. Here, we show that EthN-P on the second mannose is used as a preferential bridge for several GPI-anchored proteins. Our data modifies the current view of GPI anchors and provides mechanistic basis of PIGG deficiencies.
2022-05-31 | PXD029851 | Pride
Project description:GPI anchors many proteins to the cell surface. GPI precursor has three mannoses, all of which are modified by ethanolamine-phosphate (EthN-P). It has been believed that EthN-P on the third mannose is always used as a bridge to the protein and EthN-P on the second mannose is removed after GPI is attached to the protein. In fact, several GPI-anchored proteins are not appreciably reduced on cells defective in PIGG, which transfers EthN-P to the second mannose. Nevertheless, mutations in PIGG cause neuronal abnormalities. Here, we show that EthN-P on the second mannose is used as a preferential bridge for several GPI-anchored proteins. Our data modifies the current view of GPI anchors and provides mechanistic basis of PIGG deficiencies.
2022-05-31 | PXD029850 | Pride