Elucidation of the mechanism producing menaquinone-4 in osteoblastic cells

Vitamin K is an essential nutrient and a cofactor for the carboxylation of specific glutamyl residues of proteins to γ-glutamyl residues, which activates osteocalcin related to bone formation. Among vitamin K homologues, menaquinone-4 (MK-4) is the most active biologically, up-regulating the gene expression of bone markers, and thus has been clinically used in the treatment of osteoporosis in Japan. Recently, we confirmed that MK-4 was converted from dietary phylloquinone (PK), and then accumulated in various tissues at high concentrations. This system should play an important role in biological functions including bone formation, however, the pathway by which MK-4 is converted remains unclear. In this study, we studied the mechanism of MK-4’s conversion with chemical techniques using deuterated analogues.     

Herpes Simplex Virus 1 VP22 Inhibits AIM2-Dependent Inflammasome Activation to Enable Efficient Viral Replication

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Evidence for a pro-calcitonin

The biosynthesis of calcitonin was studied using radioimmunochemical methods and suspensions of calcitonin-producing cells derived from trout ultimobranchial glands. [¹⁴C]leucine was incorporated into cell proteins in a linear fashion for up to 36 hrs. Acid-extracted cellular radioactivity could be precipitated by trichloroacetic acid and calcitonin antiserum. Chromatography of the cell extracts revealed two distinct peaks of radio-immunoassayable and immunoprecipitable calcitonin activity. One peak coeluted with radioiodinated calcitonin, the other as a higher molecular weight species. The relative incorporation of [¹⁴C]leucine into the higher and lower molecular weight peaks during “pulse-chase” experiments was consistent with a precursor-product relationship between them.     

Spermine modulates the expression of two probable polyamine transporter genes and determines growth responses to cadaverine in Arabidopsis

Key message

Two genes, LAT1 and OCT1 , are likely to be involved in polyamine transport in Arabidopsis. Endogenous spermine levels modulate their expression and determine the sensitivity to cadaverine.

Abstract

Arabidopsis spermine (Spm) synthase (SPMS) gene-deficient mutant was previously shown to be rather resistant to the diamine cadaverine (Cad). Furthermore, a mutant deficient in polyamine oxidase 4 gene, accumulating about twofold more of Spm than wild type plants, showed increased sensitivity to Cad. It suggests that endogenous Spm content determines growth responses to Cad in Arabidopsis thaliana. Here, we showed that Arabidopsis seedlings pretreated with Spm absorbs more Cad and has shorter root growth, and that the transgenic Arabidopsis plants overexpressing the SPMS gene are hypersensitive to Cad, further supporting the above idea. The transgenic Arabidopsis overexpressing L-Amino acid Transporter 1 (LAT1) absorbed more Cad and showed increased Cad sensitivity, suggesting that LAT1 functions as a Cad importer. Recently, other research group reported that Organic Cation Transporter 1 (OCT1) is a causal gene which determines the Cad sensitivity of various Arabidopsis accessions. Furthermore, their results suggested that OCT1 is involved in Cad efflux. Thus we monitored the expression of OCT1 and LAT1 during the above experiments. Based on the results, we proposed a model in which the level of Spm content modulates the expression of OCT1 and LAT1, and determines Cad sensitivity of Arabidopsis.

     

Identification of a neuron-specific human gene, KIAA1110, that is a guanine nucleotide exchange factor for ARF1

To identify neuron-specific genes, we performed gene expression profiling, cDNA microarray and in silico ESTs (expressed sequence tags) analyses. We identified a human neuron-specific gene, KIAA1110 (homologue of rat synArfGEF (Po)), that is a member of the guanine nucleotide exchange factor (GEF) for the ADP-ribosylation factor (ARF). RT-PCR analysis showed that the KIAA1110 gene was expressed specifically in the brain among adult human tissues, whereas no apparent expression was observed in immature neural tissues/cells, such as fetal brain, glioma tissues/cells, and neural stem/precursor cells (NSPCs). The KIAA1110 protein was shown to be expressed in mature neurons but not in undifferentiated NSPCs. Immunohistochemical analysis also showed that KIAA1110 was expressed in neurons of the human adult cerebral cortex. Furthermore, the pull-down assay revealed that KIAA1110 has a GEF activity toward ARF1 that regulates transport along the secretion pathway. These results suggest that KIAA1110 is expressed specifically in mature neurons and may play an important role in the secretion pathway as a GEF for ARF1.     

Cutaneous lymphocyte antigen is expressed on memory/effector B cells in the peripheral blood and monocytoid B cells in the lymphoid tissues.

Cutaneous lymphocyte antigen (CLA) is expressed on a subpopulation of human memory T cells and is involved in the primary step of their skin homing. T cells and some B cells in the peripheral blood express CLA, but the pathophysiologic roles of CLA(+) B cells have not yet been clarified. We examined the relationships among CLA expression in B cells and immunoglobulin heavy chain subtype, the localization of CLA(+) B cells in the peripheral lymphoid tissues, and their functional binding to E-selectin. CLA was expressed on class-switched, memory B cells in the peripheral blood and tonsils as revealed by flow cytometry. Immunohistochemical staining of the lymph nodes with various types of inflammation or reactive hyperplasia showed CLA on the monocytoid B cells, which correspond to memory cells. The functional study revealed that CLA on B cells bound to E-selectin transfectants. E-selectin was detected on some of the high endothelial venules in the monocytoid B-cell-rich lymph nodes. These findings suggest that CLA is also expressed on a subset of memory/effector B cells, in addition to a subset of memory T cells. Such B cells were located in the lymph nodes or tonsils and rarely in chronic dermatitis. Therefore, CLA seems to be related to memory/effector B-cell trafficking to the lymph nodes or tonsils. According to the multistep theory, mechanisms involved in the second or third step might be different between CLA(+) B and T cells.     

Heterotypic cell interactions on a dually patterned surface

It is worth investigating heterotypic cell-cell interactions by mimicking their in vivo structures and environment. In the present study, physiological cellular response and behavior of hepatocytes and endothelial cells were investigated by controlling their contact periphery in a new co-culture system. Rat primary hepatocytes and bovine endothelial cells were co-cultured on a dually patterned surface. Hepatic physiological functions such as albumin secretion and ammonium metabolism were enhanced by increasing heterotypic cell-cell interactions in a patterned co-culture. Furthermore, enhanced hepatic functions through heterotypic interactions are effective within a limited area apart from endothelial cells as evidenced by immunofluorescence staining of hepatic intracellular albumin, indicating that heterotypic interactions act in a paracrine manner. Thus, heterotypic cell communications that play indispensable roles in increasing hepatic physiological functions should be obtained with an increasing periphery of two-cell domains. These findings are important for the reconstruction of complex tissues such as liver and pancreas.