Metabolic response against sulfur-containing heterocyclic compounds by the lignin-degrading basidiomycete Coriolus versicolor

The fungal conversions of sulfur-containing heterocyclic compounds were investigated using the lignin-degrading basidiomycete Coriolus versicolor. The fungus metabolized a series of sulfur compounds--25 structurally related thiophene derivatives--via several different pathways. Under primary metabolic conditions, C. versicolor utilized thiophenes, such as 2-hydroxymethyl-, 2-formyl-, and 2-carboxyl-thiophenes, as a nutrient sulfur source for growth; thus, the fungus degraded these compounds more effectively in a non-sulfur-containing medium than in conventional medium. The product analysis revealed that several redox reactions, decarboxylation reactions, and C-S cleavage reactions were involved in the fungal conversion of non-aromatic thiophenes. On the other hand, benzothiophene (BT) and dibenzothiophene (DBT) skeletons were converted to water-soluble products. All the products and metabolic intermediates were more hydrophilic than the starting substrates. These metabolic actions seemed to be a chemical stress response against exogenously added xenobiotics. These metabolic reactions were optimized under ligninolytic conditions, also suggesting the occurrence of a fungal xenobiotic response. Furthermore, the fungus converted a series of BTs and DBTs via several different pathways, which seemed to be controlled by the chemical structure of the substrates. DBT, 4-methylDBT, 4, 6-dimethylDBT, 2-methylBT, and 7-methylBT were immediately oxidized to their S-oxides. BTs and DBTs with the hydroxymethyl substituent were converted to their xylosides without S-oxidation. Those with carboxyl and formyl substituents were reduced to form a hydroxymethyl group, then xylosidated. These observations strongly suggested the involvement of a fungal substrate-recognition and metabolic response mechanism in the metabolism of sulfur-containing heterocyclic compounds by C. versicolor.     

Localization of BAI-associated protein1/membrane-associated guanylate kinase-1 at adherens junctions in normal rat kidney cells: polarized targeting mediated by the carboxyl-terminal PDZ domains

Brain-specific angiogenesis inhibitor (BAI)-associated protein (BAP)1 (also called membrane-associated guanylate kinase [MAGI]-1) is composed of six PSD-95/Dlg-A/ZO-1 (PDZ) domains, two WW domains, and one guanylate kinase (GK) domain. We previously reported that BAP1 is localized at tight junctions in Madine Darby canine kidney (MDCK) cells and intestinal epithelial cells. Here, we have determined the localization of BAP1 in normal rat kidney (NRK) cells that do not form tight junctions. BAP1 was colocalized with E-cadherin along the lateral membrane, suggesting its localization at adherens junctions. Green fluorescent protein (GFP)-BAP1 was distributed in the cytosol in separate NRK cells, and accumulated to the cell-cell contacts when NRK cells have contact with each other. The GFP-BAP1 mutant containing either the first PDZ and GK domains or the WW and second PDZ domains was localized in the cytosol and the nucleus. The GFP-BAP1 mutant containing the second to fourth PDZ domains was distributed in the cytosol. The construct containing the fifth and sixth PDZ domains was localized at the cell-cell contacts along the lateral membrane and slightly in the nucleus, whereas the construct lacking the fifth and sixth PDZ domains was localized in the cytosol and in the nucleus. BAP1 was tyrosine-phosphorylated in vivo, but the tyrosine phosphorylation of BAP1 was not correlated with its localization. These results suggest that the signal in the carboxyl-terminal PDZ domains functions dominantly in vivo to target BAP1 to the lateral membrane, although potential nuclear localization signals exist in the N-terminal region of BAP1.     

Fusion of bone marrow-derived cells with renal tubules contributes to renal dysfunction in diabetic nephropathy

Although diabetic nephropathy (DN) is a major cause of end-stage renal disease, the mechanism of dysfunction has not yet been clarified. We previously reported that in diabetes proinsulin-producing bone marrow-derived cells (BMDCs) fuse with hepatocytes and neurons. Fusion cells are polyploidy and produce tumor necrosis factor (TNF)-α, ultimately causing diabetic complications. In this study, we assessed whether the same mechanism is involved in DN. We performed bone marrow transplantation from male GFP-Tg mice to female C57BL/6J mice and produced diabetes by streptozotocin (STZ) or a high-fat diet. In diabetic kidneys, massive infiltration of BMDCs and tubulointerstitial injury were prominent. BMDCs and damaged tubular epithelial cells were positively stained with proinsulin and TNF-α. Cell fusion between BMDCs and renal tubules was confirmed by the presence of Y chromosome. Of tubular epithelial cells, 15.4% contain Y chromosomes in STZ-diabetic mice, 8.6% in HFD-diabetic mice, but only 1.5% in nondiabetic mice. Fusion cells primarily expressed TNF-α and caspase-3 in diabetic kidney. These in vivo findings were confirmed by in vitro coculture experiments between isolated renal tubular cells and BMDCs. It was concluded that cell fusion between BMDCs and renal tubular epithelial cells plays a crucial role in DN.     

Frequent overexpression of CADM1/IGSF4 in lung adenocarcinoma

We reported comprehensive screening for antigens (Ags) overexpressed on various carcinomas via isolation of human monoclonal antibodies (mAbs) that may be therapeutic in a previous paper (Proc. Natl. Acad. Sci. USA 105, 7287-7292, 2008). Twenty-one distinct Ags highly expressed on several carcinomas were identified and 356 mAbs with unique sequences turned out to bind to one of the 21 Ags. Among them CADM1/IGSF4 which had been originally referred to as tumor suppressor lung cancer 1 (TSLC1) was included. Therefore we examined the expression of CADM1 in lung cancers in this study. Eight different anti CADM1 mAbs were used for immunohistochemical analysis of 29 fresh lung cancer specimens. Staining patterns were categorized to six groups based on the extent of positive staining and the localization of stained portions. While overexpression of CADM1 was observed on the cell surface of adenocarcinomas at a high frequency, around 60%, positive stainings were rarely observed on that of other lung carcinomas including squamous cell carcinomas. Moreover, some clones among the eight mAbs gave different staining patterns from those by the other clones against the same fresh specimen, suggesting presence of variant forms of CADM1 differentiated by mAbs.     

Toll-like Receptors as a Target of Food-derived Anti-inflammatory Compounds

Toll-like receptors (TLRs) play a key role in linking pathogen recognition with the induction of innate immunity. They have been implicated in the pathogenesis of chronic inflammatory diseases, representing potential targets for prevention/treatment. Vegetable-rich diets are associated with the reduced risk of several inflammatory disorders. In the present study, based on an extensive screening of vegetable extracts for TLR-inhibiting activity in HEK293 cells co-expressing TLR with the NF-κB reporter gene, we found cabbage and onion extracts to be the richest sources of a TLR signaling inhibitor. To identify the active substances, we performed activity-guiding separation of the principal inhibitors and identified 3-methylsulfinylpropyl isothiocyanate (iberin) from the cabbage and quercetin and quercetin 4′-O-β-glucoside from the onion, among which iberin showed the most potent inhibitory effect. It was revealed that iberin specifically acted on the dimerization step of TLRs in the TLR signaling pathway. To gain insight into the inhibitory mechanism of TLR dimerization, we developed a novel probe combining an isothiocyanate-reactive group and an alkyne functionality for click chemistry and detected the probe bound to the TLRs in living cells, suggesting that iberin disrupts dimerization of the TLRs via covalent binding. Furthermore, we designed a variety of iberin analogues and found that the inhibition potency was influenced by the oxidation state of the sulfur. Modeling studies of the iberin analogues showed that the oxidation state of sulfur might influence the global shape of the isothiocyanates. These findings establish the TLR dimerization step as a target of food-derived anti-inflammatory compounds.     

(4-Piperidinyl)-piperazine: A new platform for acetyl-CoA carboxylase inhibitors

Acetyl-CoA carboxylases (ACCs), the rate limiting enzymes in de novo lipid synthesis, play important roles in modulating energy metabolism. The inhibition of ACC has demonstrated promising therapeutic potential for treating obesity and type 2 diabetes mellitus in transgenic mice and preclinical animal models. We describe herein the synthesis and structure–activity relationships of a series of disubstituted (4-piperidinyl)-piperazine derivatives as a new platform for ACC1/2 non-selective inhibitors.     

Rearrangements of the DNA in carbon ion-induced mutants of Arabidopsis thaliana

To elucidate the nature of structural alterations in plants, three carbon ion-induced mutations in Arabidopsis thaliana, gl1-3, tt4(C1), and ttg1-21, were analyzed. The gl1-3 mutation was found to be generated by an inversion of a fragment that contained GL1 and Atpk7 loci on chromosome 3. The size of the inverted fragment was a few hundred kilobase pairs. The inversion was found to accompany an insertion of a 107-bp fragment derived from chromosome 2. The tt4(C1) mutation was also found to be due to an inversion. The size of the intervening region between the breakpoints was also estimated to be a few hundred kilobase pairs. In the case of ttg1-21, it was found that a break occurred at the TTG1 locus on chromosome 5, and reciprocal translocation took place between it and chromosome 3. From the sequences flanking the breakpoints, the DNA strand breaks induced by carbon ions were found to be rejoined using, if present, only short homologous sequences. Small deletions were also observed around the breakpoints. These results suggest that the nonhomologous end-joining (NHEJ) pathway operates after plant cells are exposed to ion particles.