The receptor activator of NF-B (RANK) and its ligand RANKL are key molecules for differentiation and activation of osteoclasts. RANKL stimulates transcription factors AP-1 through mitogen-activated protein kinase (MAPK) activation, and NF-B through IB kinase (IKK) activation. Tumor necrosis factor receptorassociated factor 6 (TRAF6) is essential for activation of these kinases. In the interleukin-1 signaling pathway, TAK1 MAPK kinase kinase (MAPKKK) mediates MAPK and IKK activation via interaction with TRAF6, and TAB2 acts as an adapter linking TAK1 and TRAF6. Here, we demonstrate that TAK1 and TAB2 participate in the RANK signaling pathway. Dominant negative forms of TAK1 and TAB2 inhibit NF-B activation induced by overexpression of RANK. In 293 cells stably transfected with full-length RANK, RANKL stimulation facilitates the formation of a complex containing RANK, TRAF6, TAB2, and TAK1, leading to the activation of TAK1. Furthermore, in murine monocyte RAW 264.7 cells, dominant negative forms of TAK1 and TAB2 inhibit NF-B activation induced by RANKL and endogenous TAK1 is activated in response to RANKL stimulation. These results suggest that the formation of the TRAF6-TAB2-TAK1 complex is involved in the RANK signaling pathway and may regulate the development and function of osteoclasts.Skeletal remodeling is a dynamic and continual process that involves the coupled events of bone formation by osteoblasts and bone resorption by osteoclasts. Osteoclasts are professional bone-resorbing polykaryons derived from hematopoietic cells of the monocyte-macrophage lineage (27, 34). The receptor activator of NF-B (RANK) is a member of the tumor necrosis factor (TNF) receptor family and is involved in osteoclastogenesis and lymph node development (1, 10). The ligand for RANK, RANKL (also called osteoclast differentiation factor [46], TNF-related activation induced cytokine [44], and osteoprotegerin ligand [21]), is a TNF receptor family ligand that regulates the functions of dendritic cells and osteoclasts. RANKL is expressed on osteoblasts and bone marrow stromal cells, while its receptor RANK is expressed on osteoclast progenitors or mature osteoclasts. RANKL interacts with RANK via direct cell-cell contact, thereby promoting the differentiation, survival, and bone-resorbing capability of osteoclasts (reviewed in references 13 and 35). RANK interacts with members of the family of TNF receptor-associated factors (TRAFs) that mediate activation of NF-B and c-Jun NH 2 -terminal kinase (JNK) (8,11,17,43). Furthermore, the RANK cytoplasmic tail associates with c-Src kinase, which is responsible for the activation of Akt/PKB, a factor that has an antiapoptotic effect on osteoclasts (42). However, the proximal molecular components of RANK signal transduction and their interactions are not well understood.The TRAF family consists of six distinct proteins, each containing a ring and zinc finger motif in their N terminus and C-terminal TRAF domains that are responsible for self-association and protein interaction. The TRAF protei...

To elucidate the mechanism of cellulose synthesis, we isolated a mutant of Arabidopsis (changed sensitivity to cellulose synthesis inhibitors 1, css1) that showed changed sensitivity to cellulose biosynthesis inhibitor. The analysis of phenotypes indicated that the css1 mutation influenced various fundamental metabolic pathways including amino acid metabolism, triacylglycerol degradation and polysaccharide synthesis (cellulose and starch) during the early stage of plant growth. Unexpectedly, the map-based cloning of the gene responsible for the css1 mutation identified a protein (At-nMat1a) that was assumed to be a splicing factor of the mitochondrial group II intron. In accordance with this result, this mutant exhibited improper splicing of the mitochondrial NAD4 transcript. We noticed that the phenotypes of the css1 mutant are similar to the responses to anoxia that hinders mitochondrial aerobic respiration. It seems that the defect in the function of mitochondria influences various aspects of fundamental cellular metabolism including cellulose synthesis. Our results suggested that sucrose synthase (SuSy), an enzyme involved in the biosynthesis of cellulose, plays key roles in the connection between mitochondria and cellulose synthesis. The isolation of the css1 mutant also provides a useful resource in the study of post-transcriptional gene regulation in mitochondria.

Osteoblast maturation is a multistep series of events characterized by an integrated cascade of gene expression that are accompanied by specific phenotypic alterations. To find new osteoblast-related genes we cloned differentially expressed cDNAs characteristic of specific differentiation stages in the mouse osteoblast-like MC3T3-E1 cells by a differential display method. We identified a novel cDNA encoding a putative glycerophosphodiester phosphodiesterase, GDE3, which specifically was expressed at the stage of matrix maturation. Interestingly, the deduced amino acid sequence contains 539 amino acids including seven putative transmembrane domains and a glycerophosphodiester phosphodiesterase region in one of the extracellular loops. Northern blot analysis revealed that GDE3 was also expressed in spleen as well as primary calvarial osteoblasts and femur. We next transfected HEK293T cells with GDE3 with green fluorescent protein fused to the C terminus. The green fluorescent protein-fused protein accumulated at the cell periphery, and the transfected cells overexpressing the protein changed from a spread form to rounded form with disappearance of actin filaments. Immunofluorescence staining with GDE3 antibody and phalloidin in MC3T3-E1 cells indicated that endogenous GDE3 might be co-localized with the actin cytoskeleton. To identify a role for GDE3 in osteoblast differentiation, MC3T3-E1 cells stably expressing the full-length protein were constructed. Expression of GDE3 showed morphological changes, resulting in dramatic increases in alkaline phosphatase activity and calcium deposit. These results suggest that GDE3 might be a novel seven-transmembrane protein with a GP-PDE-like extracellular motif expressed during the osteoblast differentiation that dramatically accelerates the program of osteoblast differentiation and is involved in the morphological change of cells.

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We have developed a device that enables direct measurement of food texture. The device inserts a probe into a food sample and detects the vibration caused by the sample's fracture. A piezoelectric sensor was used to detect that vibration. The frequency response of the piezoelectric sensor was measured. Results showed that the sensor covered the full audio frequency range up to 20 kHz. The device probe was designed so that its resonance was not in the signal detection band. An octave multi-filter was used to analyze the obtained signals. Preliminary data were obtained and used to quantify the sample texture. Quantification of food texture was possible by using the device together with the analytical tool. We also discuss the application of the device to investigation of physical and structural aspects of food.