Comparative studies of glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1: evidence for a eutherian mammalian origin for the GPIHBP1 gene from an LY6-like gene

Glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1 (GPIHBP1) functions as a platform and transport agent for lipoprotein lipase (LPL) which functions in the hydrolysis of chylomicrons, principally in heart, skeletal muscle and adipose tissue capillary endothelial cells. Previous reports of genetic deficiency for this protein have described severe chylomicronemia. Comparative GPIHBP1 amino acid sequences and structures and GPIHBP1 gene locations were examined using data from several mammalian genome projects. Mammalian GPIHBP1 genes usually contain four coding exons on the positive strand. Mammalian GPIHBP1 sequences shared 41–96% identities as compared with 9–32% sequence identities with other LY6-domain-containing human proteins (LY6-like). The human N-glycosylation site was predominantly conserved among other mammalian GPIHBP1 proteins except cow, dog and pig. Sequence alignments, key amino acid residues and conserved predicted secondary structures were also examined, including the N-terminal signal peptide, the acidic amino acid sequence region which binds LPL, the glycosylphosphatidylinositol linkage group, the Ly6 domain and the C-terminal α-helix. Comparative and phylogenetic studies of mammalian GPIHBP1 suggested that it originated in eutherian mammals from a gene duplication event of an ancestral LY6-like gene and subsequent integration of exon 2, which may have been derived from BCL11A (B-cell CLL/lymphoma 11A gene) encoding an extended acidic amino acid sequence

Analysis of Common and Specific Mechanisms of Liver Function Affected by Nitrotoluene Compounds

BACKGROUND: Nitrotoluenes are widely used chemical manufacturing and munitions applications. This group of chemicals has been shown to cause a range of effects from anemia and hypercholesterolemia to testicular atrophy. We have examined the molecular and functional effects of five different, but structurally related, nitrotoluenes on using an integrative systems biology approach to gain insight into common and disparate mechanisms underlying effects caused by these chemicals. METHODOLOGY/PRINCIPAL FINDINGS: Sprague-Dawley female rats were exposed via gavage to one of five concentrations of one of five nitrotoluenes [2,4,6-trinitrotoluene (TNT), 2-amino-4,6-dinitrotoluene (2ADNT) 4-amino-2,6-dinitrotoulene (4ADNT), 2,4-dinitrotoluene (2,4DNT) and 2,6-dinitrotoluene (2,6DNT)] with necropsy and tissue collection at 24 or 48 h. Gene expression profile results correlated well with clinical data and liver histopathology that lead to the concept that hematotoxicity was followed by hepatotoxicity. Overall, 2,4DNT, 2,6DNT and TNT had stronger effects than 2ADNT and 4ADNT. Common functional terms, gene expression patterns, pathways and networks were regulated across all nitrotoluenes. These pathways included NRF2-mediated oxidative stress response, aryl hydrocarbon receptor signaling, LPS/IL-1 mediated inhibition of RXR function, xenobiotic metabolism signaling and metabolism of xenobiotics by cytochrome P450. One biological process common to all compounds, lipid metabolism, was found to be impacted both at the transcriptional and lipid production level. CONCLUSIONS/SIGNIFICANCE: A systems biology strategy was used to identify biochemical pathways affected by five nitroaromatic compounds and to integrate data that tie biochemical alterations to pathological changes. An integrative graphical network model was constructed by combining genomic, gene pathway, lipidomic, and physiological endpoint results to better understand mechanisms of liver toxicity and physiological endpoints affected by these compounds

Comparative Analysis of Gene Regulation by the Transcription Factor PPARα between Mouse and Human

Studies in mice have shown that PPARalpha is an important regulator of hepatic lipid metabolism and the acute phase response. However, little information is available on the role of PPARalpha in human liver. Here we set out to compare the function of PPARalpha in mouse and human hepatocytes via analysis of target gene regulation

Gene Expression Programs of Mouse Endothelial Cells in Kidney Development and Disease

Endothelial cells are remarkably heterogeneous in both morphology and function, and they play critical roles in the formation of multiple organ systems. In addition endothelial cell dysfunction can contribute to disease processes, including diabetic nephropathy, which is a leading cause of end stage renal disease. In this report we define the comprehensive gene expression programs of multiple types of kidney endothelial cells, and analyze the differences that distinguish them. Endothelial cells were purified from Tie2-GFP mice by cell dissociation and fluorescent activated cell sorting. Microarrays were then used to provide a global, quantitative and sensitive measure of gene expression levels. We examined renal endothelial cells from the embryo and from the adult glomerulus, cortex and medulla compartments, as well as the glomerular endothelial cells of the db/db mutant mouse, which represents a model for human diabetic nephropathy. The results identified the growth factors, receptors and transcription factors expressed by these multiple endothelial cell types. Biological processes and molecular pathways were characterized in exquisite detail. Cell type specific gene expression patterns were defined, finding novel molecular markers and providing a better understanding of compartmental distinctions. Further, analysis of enriched, evolutionarily conserved transcription factor binding sites in the promoters of co-activated genes begins to define the genetic regulatory network of renal endothelial cell formation. Finally, the gene expression differences associated with diabetic nephropathy were defined, providing a global view of both the pathogenic and protective pathways activated. These studies provide a rich resource to facilitate further investigations of endothelial cell functions in kidney development, adult compartments, and disease

Inactivation of Icmt inhibits transformation by oncogenic K-Ras and B-Raf

Isoprenylcysteine carboxyl methyltransferase (Icmt) methylates the carboxyl-terminal isoprenylcysteine of CAAX proteins (e.g., Ras and Rho proteins). In the case of the Ras proteins, carboxyl methylation is important for targeting of the proteins to the plasma membrane. We hypothesized that a knockout of Icmt would reduce the ability of cells to be transformed by K-Ras. Fibroblasts harboring a floxed Icmt allele and expressing activated K-Ras (K-Ras-Icmt(flx/flx)) were treated with Cre-adenovirus, producing K-Ras-Icmt(Δ/Δ) fibroblasts. Inactivation of Icmt inhibited cell growth and K-Ras–induced oncogenic transformation, both in soft agar assays and in a nude mice model. The inactivation of Icmt did not affect growth factor–stimulated phosphorylation of Erk1/2 or Akt1. However, levels of RhoA were greatly reduced as a consequence of accelerated protein turnover. In addition, there was a large Ras/Erk1/2-dependent increase in p21(Cip1), which was probably a consequence of the reduced levels of RhoA. Deletion of p21(Cip1) restored the ability of K-Ras-Icmt(Δ/Δ) fibroblasts to grow in soft agar. The effect of inactivating Icmt was not limited to the inhibition of K-Ras–induced transformation: inactivation of Icmt blocked transformation by an oncogenic form of B-Raf (V599E). These studies identify Icmt as a potential target for reducing the growth of K-Ras– and B-Raf–induced malignancies

The GPIHBP1–LPL Complex Is Responsible for the Margination of Triglyceride-Rich Lipoproteins in Capillaries

Triglyceride-rich lipoproteins (TRLs) undergo lipolysis by lipoprotein lipase (LPL), an enzyme that is transported to the capillary lumen by an endothelial cell protein, GPIHBP1. For LPL-mediated lipolysis to occur, TRLs must bind to the lumen of capillaries. This process is often assumed to involve heparan sulfate proteoglycans (HSPGs), but we suspected that TRL margination might instead require GPIHBP1. Indeed, TRLs marginate along the heart capillaries of wild-type but not Gpihbp1 -/- mice, as judged by fluorescence microscopy, quantitative assays with infrared-dye-labeled lipoproteins, and EM tomography. Both cell-culture and in vivo studies showed that TRL margination depends on LPL bound to GPIHBP1. Notably, the expression of LPL by endothelial cells in Gpihbp1 -/- mice did not restore defective TRL margination, implying that the binding of LPL to HSPGs is ineffective in promoting TRL margination. Our studies show that GPIHBP1-bound LPL is the main determinant of TRL margination

The GPIHBP1-LPL complex is responsible for the margination of triglyceride-rich lipoproteins in capillaries

Triglyceride-rich lipoproteins (TRLs) undergo lipolysis by lipoprotein lipase (LPL), an enzyme that is transported to the capillary lumen by an endothelial cell protein, GPIHBP1. For LPL-mediated lipolysis to occur, TRLs must bind to the lumen of capillaries. This process is often assumed to involve heparan sulfate proteoglycans (HSPGs), but we suspected that TRL margination might instead require GPIHBP1. Indeed, TRLs marginate along the heart capillaries of wild-type but not Gpihbp1-/- mice, as judged by fluorescence microscopy, quantitative assays with infrared-dye-labeled lipoproteins, and EM tomography. Both cell-culture and in vivo studies showed that TRL margination depends on LPL bound to GPIHBP1. Notably, the expression of LPL by endothelial cells in Gpihbp1-/- mice did not restore defective TRL margination, implying that the binding of LPL to HSPGs is ineffective in promoting TRL margination. Our studies show that GPIHBP1-bound LPL is the main determinant of TRL margination. © 2014 Elsevier Inc