Inference of S-system models of genetic networks using a cooperative coevolutionary algorithm

MOTIVATION: To resolve the high-dimensionality of the genetic network inference problem in the S-system model, a problem decomposition strategy has been proposed. While this strategy certainly shows promise, it cannot provide a model readily applicable to the computational simulation of the genetic network when the given time-series data contain measurement noise. This is a significant limitation of the problem decomposition, given that our analysis and understanding of the genetic network depend on the computational simulation. RESULTS: We propose a new method for inferring S-system models of large-scale genetic networks. The proposed method is based on the problem decomposition strategy and a cooperative coevolutionary algorithm. As the subproblems divided by the problem decomposition strategy are solved simultaneously using the cooperative coevolutionary algorithm, the proposed method can be used to infer any S-system model ready for computational simulation. To verify the effectiveness of the proposed method, we apply it to two artificial genetic network inference problems. Finally, the proposed method is used to analyze the actual DNA microarray data.     

A new analysis of the depolymerized fragments of lignin polymer using ToF-SIMS

Lignin in plant cell walls is a complex, irregular polymer built from phenylpropanoid C6-C3 units that are connected via various C-C and C-O linkages. A recent study using time-of-flight secondary ion mass spectrometry (ToF-SIMS) with Ga primary ion bombardment showed that lignin polymers can be characterized by specific positive ions possessing a substituted aromatic ring (so-called guaiacyl or syringyl rings), which are the basic building units of lignin. To study the relationship between the characteristic ions of lignin and the common interunit linkages, various lignin dimer model compounds were investigated using ToF-SIMS. The resulting dimer spectra showed that the characteristic ions with a guaiacyl ring at m/z 137 and 151 result from rupture of most common interunit linkages, not only 8-O-4' linkages, which are the most abundant in lignin, but also 8-1', 8-5', and 8-8'. There was no evidence of rupture of 5-5' linkages. These results show that ToF-SIMS offers a new tool for the direct analysis of the depolymerized fragments of lignin polymers. The mechanisms for the fragmentation of lignin dimer models in ToF-SIMS were proposed that allow ToF-SIMS fragmentation rules to be deduced. Adduct ions such as [M + 13]+ ([M + CH]+) were also produced in fragmentation of the dimers and are thought to arise from the combination of the molecules with their stable fragments.     

Expression of class A scavenger receptor is enhanced by high glucose in vitro and under diabetic conditions in vivo: one mechanism for an increased rate of atherosclerosis in diabetes

In the early stage of atherosclerosis, macrophages take up chemically modified low density lipoproteins (LDL) through the scavenger receptors, leading to foam cell formation in atherosclerotic lesions. To get insight into a role of the scavenger receptors in diabetes-enhanced atherosclerotic complications, the effects on class A scavenger receptor (SR-A) of high glucose exposure in vitro as well as the diabetic conditions in vivo were determined in the present study. The in vitro experiments demonstrated that high glucose exposure to human monocyte-derived macrophages led to an increased SR-A expression with a concomitant increase in the endocytic uptake of acetylated LDL and oxidized LDL. The endocytic process was significantly suppressed by an anti-SR-A neutralizing antibody. Stability analyses revealed a significant increased stability of SR-A at a mRNA level but not a protein level, indicating that high glucose-induced up-regulation of SR-A is due largely to increased stability of SR-A mRNA. High glucose-enhanced SR-A expression was prevented by protein kinase C and NAD(P)H oxidase inhibitors as well as antioxidants. High glucose-enhanced production of intracellular peroxides was visualized in these cells, which was attenuated by an antioxidant. The in vivo experiments demonstrated that peritoneal macrophages from streptozotocin-induced diabetic mice increased SR-A expression when compared with those from nondiabetic mice. Endocytic degradation of acetylated LDL and oxidized LDL were also increased with these macrophages but not with the corresponding macrophages from diabetic SR-A knock-out mice. These in vitro and in vivo results probably suggest that reactive oxygen species generated from a protein kinase C-dependent NAD(P)H oxidase pathway plays a role in the high glucose-induced up-regulation of SR-A, leading to the increased endocytic degradation of modified LDL for foam cell formation. This could be one mechanism for an increased rate of atherosclerosis in patients with diabetes.     

Heparin regulates vascular endothelial growth factor165-dependent mitogenic activity, tube formation, and its receptor phosphorylation of human endothelial cells. Comparison of the effects of heparin and modified heparins

Vascular endothelial growth factor (VEGF) is a family of glycoproteins with potent angiogenic activity. We reported previously that heparin has an affinity for VEGF165, the major isoform of VEGF, whereas 2-O-desulfated heparin and 6-O-desulfated heparin have weak but significant affinity (Ashikari-Hada, S., Habuchi, H., Kariya, Y., Itoh, N., Reddi, A. H., and Kimata, K. (2004) J. Biol. Chem. 279, 12346-12354). In this study, we first examined the effect of heparin and modified heparins (completely desulfated N-sulfated heparin, 2-O-desulfated heparin, and 6-O-desulfated heparin) on VEGF165-dependent mitogenic activity and tube formation on type I collagen gels of human umbilical vein endothelial cells. Both were enhanced by heparin, but not by modified heparins, suggesting that both the 2-O-sulfate group of hexuronic acid and the 6-O-sulfation group of N-sulfoglucosamine in heparin/heparan sulfate are necessary for VEGF165 activity. We then examined the activation of VEGF receptor (VEGFR) to understand the mechanism. We have made several new findings; 1) heparin yielded a 1.7-fold enhancement of VEGF165-induced phosphorylation of VEGFR-2; 2) depletion of cell surface heparan sulfate by heparinase/heparitinase treatment and preferential reduction of trisulfated disaccharide units of cell surface HS by sodium chlorate treatment resulted in the reduction of such phosphorylation, suggesting the involvement of a heparin-like domain in the phosphorylation of VEGFR-2; and 3) VEGF121, an isoform without the exon 7-encoded region, which has no capacity to bind to heparin, did not show these effects. It is therefore likely that a heparin-like domain of heparan sulfate/heparin forms a complex with VEGF165 and VEGFR-2 via the exon 7-encoded region, thereby enhancing VEGF165-dependent signaling.     

Evidence for Cd101 but not Fcgr1 as candidate for type 1 diabetes locus, Idd10

Among polygenes conferring susceptibility to type 1 diabetes in the NOD mouse, Idd10 on distal chromosome 3 has been shown to be important for disease susceptibility. In this study, we investigated the candidacy of Fcgr1 and Cd101 for Idd10, by congenic mapping and candidate gene sequencing. Among seven NOD-related strains studied, the IIS mouse was found to possess a recombinant Idd10 interval with the same sequence at Fcgr1 as the NOD mouse, but a different sequence at Cd101 from that in the NOD mouse with 10 amino acid substitutions. The frequency of type 1 diabetes in NOD mice congenic for IIS Idd10 (NOD.IISIdd10) was significantly reduced as compared to that in the NOD mouse, despite the presence of the identical Fcgr1 sequence. These data indicate that IIS mice possess a resistant allele at Idd10, and suggest that Cd101, but not Fcgr1, is responsible for the Idd10 effect.     

GAT (GGA and Tom1) domain responsible for ubiquitin binding and ubiquitination

GGAs (Golgi-localizing, gamma-adaptin ear domain homology, ADP-ribosylation factor (ARF)-binding proteins) are a family of monomeric adaptor proteins involved in membrane trafficking from the trans-Golgi network to endosomes. The GAT (GGA and Tom1) domains of GGAs have previously been shown to interact with GTP-bound ARF and to be crucial for membrane recruitment of GGAs. Here we show that the C-terminal subdomain of the GAT domain, which is distinct from the N-terminal GAT subdomain responsible for ARF binding, can bind ubiquitin. The binding is mediated by interactions between residues on one side of the alpha3 helix of the GAT domain and those on the so-called Ile-44 surface patch of ubiquitin. The binding of the GAT domain to ubiquitin can be enhanced by the presence of a GTP-bound form of ARF. Furthermore, GGA itself is ubiquitinated in a manner dependent on the GAT-ubiquitin interaction. These results delineate the molecular basis for the interaction between ubiquitin and GAT and suggest that GGA-mediated trafficking is regulated by the ubiquitin system as endosomal trafficking mediated by other ubiquitin-binding proteins.     

TDAG8 is a proton-sensing and psychosine-sensitive G-protein-coupled receptor

T cell death-associated gene 8 (TDAG8) has been reported to be a receptor for psychosine. Ovarian cancer G-protein-coupled receptor 1 (OGR1) and GPR4, G-protein-coupled receptors (GPCRs) closely related to TDAG8, however, have recently been identified as proton-sensing or extracellular pH-responsive GPCRs that stimulate inositol phosphate and cAMP production, respectively. In the present study, we examined whether TDAG8 senses extracellular pH change. In the several cell types that were transfected with TDAG8 cDNA, cAMP was markedly accumulated in response to neutral to acidic extracellular pH, with a peak response at approximately pH 7.0-6.5. The pH effect was inhibited by copper ions and was reduced or lost in cells expressing mutated TDAG8 in which histidine residues were changed to phenylalanine. In the membrane fractions prepared from TDAG8-transfected cells, guanosine 5'-O-(3-thiotriphosphate) binding activity and adenylyl cyclase activity were remarkably stimulated in response to neutral and acidic pH. The concentration-dependent effect of extracellular protons on cAMP accumulation was shifted to the right in the presence of psychosine. The inhibitory psychosine effect was also observed for pH-dependent actions in OGR1- and GPR4-expressing cells but not for prostaglandin E(2)- and sphingosine 1-phosphate-induced actions in any pH in native and sphingosine 1-phosphate receptor-expressing cells. Glucosylsphingosine and sphingosylphosphorylcholine similarly inhibited the pH-dependent action, although to a lesser extent. Psychosine-sensitive and pH-dependent cAMP accumulation was also observed in mouse thymocytes. We concluded that TDAG8 is one of the proton-sensing GPCRs coupling to adenylyl cyclase and psychosine, and its related lysosphingolipids behave as if they were antagonists against protein-sensing receptors, including TDAG8, GPR4, and OGR1.     

Degradation of fodrin by m-calpain in fibroblasts adhering to fibrillar collagen I gel

When skin fibroblasts were cultured on fibrillar collagen I gel, we observed rapid degradation of talin, fodrin and ezrin, which are well-known calpain substrates. The protease m-calpain was activated only in cells adhering to fibrillar collagen, whereas micro-calpain was activated in cells adhering to monomeric or fibrillar collagen at the same level. The calpain inhibitor Z-Leu-Leu-aldehyde inhibited degradation of fodrin, but not talin. Degradation of fodrin, alpha-actinin and ezrin was prevented by over-expression of dominant negative m-calpain. However, over-expression of calpastatin, an endogenous calpain inhibitor, had no effect the degradation of these three proteins. These results suggest that m-calpain is responsible for degradation of their membrane proteins via adhesion to fibrillar collagen I gel.     

Structural and functional relationship among diamines in terms of inhibition of cell growth

Following the report that agmatine has an anti-proliferative effect on cell growth through induction of antizyme [Satriano et al. (1998) J. Biol. Chem. 273, 15313-15316], we examined the effects of 16 different diamines on cell growth. Many diamines had little or no effect on cell growth, but agmatine and 1,6-hexanediamine had anti-proliferative effects, with agmatine having the strongest effect. Inhibition of cell growth occurred after 2 days, and inhibitory effects paralleled the degree of antizyme induction. Decreased spermine levels indicated that induction of spermidine/spermine N(1)-acetyltransferase was also involved in the inhibition of cell growth by agmatine and 1,6-hexanediamine. The frameshift efficiency (ratio of antizyme synthesis with or without frameshift) measured in a rabbit reticulocyte cell-free system was also increased by 1,3-propanediamine and cis-1,4-cyclohexanediamine in addition to agmatine and 1,6-hexanediamine. However, the intracellular levels of 1,3-propanediamine and cis-1,4-cyclohexanediamine were low when these compounds were added to the cell-culture medium. Other diamines had no effect on cell growth or frameshift efficiency. The results suggest that the presence of two amino-groups separated by an appropriate distance is important for the enhancement of frameshifting by diamines.