ABSTRACT
Ceramide can be converted into sphingomyelin by sphingomyelin synthases (SMS) 1 and 2. In this study, we show that in human leukemia Jurkat cells, which express mainly SMS1, Fas ligand (FasL) treatment inhibited SMS activity in a dose- and time-dependent manner before nuclear fragmentation. The SMS inhibition elicited by FasL (1) was abrogated by benzyloxycarbonyl valyl-alanyl-aspartyl-(O-methyl)-fluoromethylketone (zVAD-fmk), a broad-spectrum caspase inhibitor; (2) did not occur in caspase-8-deficient cells and (3) was not affected in caspase-9-deficient cells. Western blot experiments showed SMS1 cleavage in a caspase-dependent manner upon FasL treatment. In a cell-free system, caspase-2, -7, -8 and -9, but not caspase-3 and -10, cleaved SMS1. In HeLa cells, SMS1 was Golgi localized and relocated throughout the cytoplasm in cells exhibiting an early apoptotic phenotype on FasL treatment. zVAD-fmk prevented FasL-induced SMS1 relocation. Thus, FasL-mediated SMS1 inhibition and relocation depend on caspase activation and likely represent proximal events in Fas signaling. FasL-induced ceramide production and cell death were enhanced in cells stably expressing an siRNA against SMS1. Conversely, in cells stably overexpressing SMS1, FasL neither increased ceramide generation nor efficiently induced cell death. Altogether, our data show that SMS1 is a novel caspase target that is functionally involved in the regulation of FasL-induced apoptosis.
Subject(s)
Apoptosis/physiology , Caspases/metabolism , Fas Ligand Protein/metabolism , Leukemia/metabolism , Membrane Proteins/metabolism , Nerve Tissue Proteins/metabolism , Sphingomyelins/metabolism , Transferases (Other Substituted Phosphate Groups)/metabolism , Amino Acid Chloromethyl Ketones/pharmacology , Ceramides/metabolism , Dose-Response Relationship, Drug , Enzyme Inhibitors/pharmacology , Fas Ligand Protein/pharmacology , Golgi Apparatus/enzymology , HeLa Cells , Humans , Jurkat Cells , Membrane Proteins/antagonists & inhibitors , Membrane Proteins/genetics , Nerve Tissue Proteins/antagonists & inhibitors , Nerve Tissue Proteins/genetics , RNA Interference/physiology , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , Signal Transduction/physiology , Transferases (Other Substituted Phosphate Groups)/antagonists & inhibitors , Transferases (Other Substituted Phosphate Groups)/geneticsABSTRACT
The aim of this study was to clarify the nature of the clonal lymphocyte infiltration in Sjögren's syndrome (SS) patients associated with lymphoproliferative disorders. We examined B cell clonality in lymphoproliferative tissues from six primary SS patients associated with lymphoproliferative disorders or lymphoma by cloning and sequencing of the gene rearrangement of the immunoglobulin heavy chain complementarity determining region 3 (IgVH-CDR3). Three patients with sequential observation showed progressional clonal expansion with the presence of the same subclone in different tissues during the course of disease. Among them, one patient developed mucosa-associated lymphoid tissue (MALT) lymphoma in glandular parotid. The other three SS patients concomitant with malignant B cells lymphomas showed different clonal expansion of B cells between nodal sites and salivary glands. The cloanality analysis indicated that monoclonal B cell population could spread from one glandular site to another site during the course of SS, suggesting that the malignant clone may arise from the general abnormal microenvironment, not restricted to the glandular tissue, in some SS patients.
Subject(s)
B-Lymphocytes/pathology , Lymphoproliferative Disorders/pathology , Neoplastic Stem Cells/pathology , Sjogren's Syndrome/pathology , Aged , Aged, 80 and over , Amino Acid Sequence , Complementarity Determining Regions/genetics , Disease Progression , Female , Gene Rearrangement, B-Lymphocyte, Heavy Chain , Humans , Immunoglobulin Heavy Chains/genetics , Lymphoma, B-Cell/pathology , Lymphoma, B-Cell, Marginal Zone/pathology , Male , Middle Aged , Polymerase Chain Reaction/methodsABSTRACT
AIMS: To isolate and characterize an oxalate-degrading Pandoraea sp. OXJ-11. METHODS AND RESULTS: A new bacterium Pandoraea sp. OXJ-11 was isolated from soil samples, which can grow in the medium with oxalate as the sole carbon and energy source. The isolate OXJ-11 is Gram-negative straight rod. It occurs singly and is motile by means of a double polar flagellum. Catalase is positive and nitrate is not reduced. It grows aerobically and the optimum growth temperature and the optimum pH are at 30 degrees C and pH 6.0, respectively. The polyphasic taxonomic data along with 16S rRNA sequence comparison demonstrate that the isolate OXJ-11 should belong to the genus Pandoraea and represent a new member in this family. CONCLUSIONS: Oxalate could be degraded and the oxalate-degrading enzyme activity was detected when the isolate OXJ-11 grew in the medium with oxalate as carbon source. SIGNIFICANCE AND IMPACT OF THE STUDY: Oxalate-degrading Pandoraea sp. OXJ-11 would be beneficial to the potential application in the control of sclerotinia stem rot in economically important plants caused by fungus Sclerotinia sclerotiorum, and in making plants resistant to the white mold disease by oxalate-degrading enzyme transgene.
