Polycomb mediated epigenetic silencing and replication timing at the INK4a/ARF locus during senescence.

Background: The INK4/ARF locus encodes three tumor suppressor genes (p15(Ink4b), Arf and p16(Ink4a)) and is frequently inactivated in a large number of human cancers. Mechanisms regulating INK4/ARF expression are not fully characterized.

Principal Findings: Here we show that in young proliferating embryonic fibroblasts (MEFs) the Polycomb Repressive Complex 2 (PRC2) member EZH2 together with PRC1 members BMI1 and M33 are strongly expressed and localized at the INK4/ARF regulatory domain (RD) identified as a DNA replication origin.

Identification of Salmonella functions critical for bacterial cell division within eukaryotic cells.

Salmonella typhimurium multiplication inside eukaryotic host cells is critical for virulence. Salmonella typhimurium strain SL1344 appears as filaments upon growth in macrophages and MelJuSo cells, a human melanoma cell line, indicating a specific blockage in the bacterial cell division process. Several studies have investigated the host cell response impairing bacterial division.

Intracellular trafficking study of a RB51 B. abortus vaccinal strain isolated from cow milk.

Brucella is responsible for one of the major worldwide zoonoses. Over the last century, several vaccines have been used against brucellosis. Among these, the rough vaccine Brucella abortus RB51 was introduced with the idea that it would not interfere with the diagnosis of brucellosis.

An evolutionary and structural perspective on T cell antigen receptor function.

After a brief overview of the themes and variations that occur in the family of receptors containing immunoreceptor tyrosine-based activation motifs (ITAMs), and of recent structural data on the ligand-binding subunits of these receptors, we use these data to revisit how information on the state and quality of occupancy of the binding site of the T cell antigen receptor (TCR) is conveyed to the proximal components of the TCR transduction cassette.

Brucella intracellular life: from invasion to intracellular replication.

Brucella organisms are pathogens that ultimate goal is to propagate in their preferred niche, the cell. Upon cell contact the bacteria is internalized via receptor molecules by activating small GTPases of the Rho subfamily and by a moderate recruitment of actin filaments. Once inside cells, Brucella localizes in early phagosomes, where it avoids fusion with late endosomes and lysosomes.

Analysis of donor NK and T cells infused in patients undergoing MHC-matched allogeneic hematopoietic transplantation.

We retrospectively analyzed the percentages and absolute numbers of T cells, natural killer (NK) cells and NK cell subsets in cryopreserved samples of either bone marrow or blood non-T cell-depleted allogeneic MHC-matched hematopoietic grafts. Using flow cytometry, we found higher numbers of NK cells in aphereses than in bone marrow collections. We further investigated the distribution of NK cell subsets, defined by the cell surface expression of MHC class I-specific receptors, in these allogeneic grafts.

Comparative analysis of the promoter structure and genomic organization of the human and mouse ABCA7 gene encoding a novel ABCA transporter.

We report here the genomic and transcriptional characterization in mouse and man of a novel transporter of the ABCA subclass, named ABCA7. As it is the case for other ABCA genes, the predicted protein encoded by ABCA7 is a full symmetric transporter, highly conserved across species. The ABCA7 gene maps to human chromosome 19 and to the homologous region at band B4-C1 on mouse chromosome 10.

Involvement of inhibitory NKRs in the survival of a subset of memory-phenotype CD8+ T cells.

Inhibitory natural killer receptors (NKRs) such as killer cell immunoglobulin-like receptors (KIRs) in humans and Ly49 molecules in mice are expressed on NK cells and recognize multiple major histocompatibility (MHC) class I proteins. In humans and mice, a subset of CD8+ T cells also expresses NKRs and harbors a memory phenotype. Using mice that are transgenic for KIR2DL3 and its cognate HLA-Cw3 ligand, we show that engagement of inhibitory NKRs selectively drives the in vivo accumulation of a subset of memory-phenotype CD8+ T cells that express the beta chain of the interleukin 2 receptor.

Identification of endogenous peptides recognized by in vivo or in vitro generated alloreactive cytotoxic T lymphocytes: distinct characteristics correlated with CD8 dependence.

We studied the molecular basis for CD8 independence of in vivo generated (BM3.3) versus CD8 dependence of in vitro sensitized (KB5.C20/Des) alloreactive H-2K(b)-specific cytotoxic T lymphocytes (CTL).

The lack of NF-kappa B transactivation and PKC epsilon expression in CD4(+)CD8(+) thymocytes correlates with negative selection.

Deletion of autoreactive thymocytes at the DP stage is the basis for tolerance to thymus-expressed self antigens. In this study we investigated whether distinct signalling pathways are induced in DP thymocytes as compared to mature T cells upon stimulation with antigen. Using triple transgenic mice expressing a TCR transgene, dominant negative ras/Mek proteins and a reporter gene construct with AP-1 or NF-kappa B binding sites, we showed a complete lack of transcriptional activity of NF-kappa B but not AP-1 in DP thymocytes, whereas both were transcriptionally active in mature T cells after antigenic stimulation.

Specific docking of apolipoprotein A-I at the cell surface requires a functional ABCA1 transporter.

The identification of defects in ABCA1 as the molecular basis of Tangier disease has highlighted its crucial role in the loading with phospholipids and cholesterol of nascent apolipoprotein particles. Indeed the expression of ABCA1 affects apolipoprotein A-I (apoA-I)-mediated removal of lipids from cell membranes, and the possible role of ABCA1 as an apoA-I surface receptor has been recently suggested. In the present study, we have investigated the role of the ABCA1 transporter as an apoA-I receptor with the analysis of a panel of transfectants expressing functional or mutant forms of ABCA1.

Function of Rho family proteins in actin dynamics during phagocytosis and engulfment.

Phagocytosis is the uptake of large particles by cells by a mechanism that is based on local rearrangement of the actin microfilament cytoskeleton. In higher organisms, phagocytic cells are essential for host defence against invading pathogens, and phagocytosis contributes to inflammation and the immune response. In addition, engulfment, defined as the phagocytic clearance of cell corpses generated by programmed cell death or apoptosis, has an essential role in tissue homeostasis.

Combined natural killer cell and dendritic cell functional deficiency in KARAP/DAP12 loss-of-function mutant mice.

KARAP/DAP12 is a transmembrane polypeptide with an intracytoplasmic immunoreceptor tyrosine-based activation motif (ITAM). KARAP/DAP12 is associated with several activating cell surface receptors in hematopoietic cells. Here, we report that knockin mice bearing a nonfunctional KARAP/DAP12 ITAM present altered innate immune responses.

The Pim-1 kinase stimulates maturation of TCRbeta-deficient T cell progenitors: implications for the mechanism of Pim-1 action.

We demonstrate that overexpression of Pim-1, a cytoplasmic serine/threonine kinase of poorly defined function, results in the development of substantial numbers of CD4(+)CD8(+) double-positive thymocytes in two independent knock-out mouse models (i.e. the RAG-1-deficient and TCRbeta gene enhancer-deleted mice) in which production of a functionally rearranged TCRbeta gene (hence the pre-TCR) is impaired.

Association of signal-regulatory proteins beta with KARAP/DAP-12.

The signal-regulatory proteins (SIRP) are Ig-like cell surface receptors detected in hematopoietic and non-hematopoietic cells. SIRP are classified as SIRPalpha molecules, containing a 110- to 113-amino acid long, or SIRPbeta molecules, with a 5-amino acid long intracytoplasmic domain. SIRPalpha molecules belong to inhibitory immunoreceptor tyrosine-based inhibition motif (ITIM)-bearing molecules.

Membrane recruitment of Rac1 triggers phagocytosis.

Rac1 is a &Rgr;-family GTP-binding protein that controls lamellipodia formation and membrane ruffling in fibroblasts. Recently, Rac1 and Cdc42, another member of the &Rgr;-family, have been shown to regulate Fc receptor-mediated phagocytosis in macrophages by controlling different steps of membrane and actin dynamics leading to particle engulfment. Here, we investigated the function of Rac1 using a membrane recruitment system that mimics phagocytosis.

Signal transduction. FasL binds preassembled Fas.

The binding of a ligand to its receptor has always been viewed as the trigger for signal transduction to ensue. However, as Golstein explains in his Perspective, new findings (Chan et al. and Siegel et al.

Altered lymphoid development in mice deficient for the mAF4 proto-oncogene.

Some chromosomal translocations in acute leukemias involve the fusion of the trithorax-related protein Mll (also called HRX, All1, Htrx,) with a variety of heterologous proteins. In acute lymphoblastic leukemia associated with the t(4;11)(q21;q23) translocation, the 4q21 gene that fuses with Mll is AF4. To gain insight into the potential role of AF4 in leukemogenesis and development, this gene was inactivated by homologous recombination in mice.

ABC1 promotes engulfment of apoptotic cells and transbilayer redistribution of phosphatidylserine.

ATP-binding-cassette transporter 1 (ABC1) has been implicated in processes related to membrane-lipid turnover. Here, using in vivo loss-of-function and in vitro gain-of-function models, we show that ABC1 promotes Ca2+-induced exposure of phosphatidylserine at the membrane, as determined by a prothrombinase assay, membrane microvesiculation and measurement of transbilayer redistribution of spin-labelled phospholipids. That ABC1 promotes engulfment of dead cells is shown by the impaired ability of ABC1-deficient macrophages to engulf apoptotic preys and by the acquisition of phagocytic behaviour by ABC1 transfectants.

TCRalpha enhancer activation occurs via a conformational change of a pre-assembled nucleo-protein complex.

The TCR alpha enhancer (Ealpha) has served as a paradigm for studying how enhancers organize trans-activators into nucleo-protein complexes thought to recruit and synergistically stimulate the transcriptional machinery. Little is known, however, of either the extent or dynamics of Ealpha occupancy by nuclear factors during T cell development. Using dimethyl sulfate (DMS) in vivo footprinting, we demonstrate extensive Ealpha occupancy, encompassing both previously identified and novel sites, not only in T cells representing a developmental stage where Ealpha is known to be active (CD4(+)CD8(+)-DP cells), but surprisingly, also in cells at an earlier developmental stage where Ealpha is not active (CD4(-)CD8(-)-DN cells).

Signaling pathways engaged by NK cell receptors: double concerto for activating receptors, inhibitory receptors and NK cells.

Despite the absence of antigen-specific receptors at their surface, NK cells can selectively eliminate virus-infected cells, tumor cells and allogenic cells. A dynamic and precisely coordinated balance between activating and inhibitory receptors governs NK cell activation programs. Multiple activating and inhibitory NK cell surface molecules have been described, a group of them acting as receptors for MHC class I molecules.

Class I-restricted presentation of exogenous antigen acquired by Fcgamma receptor-mediated endocytosis is regulated in dendritic cells.

We evaluated MHC class I- and II-restricted presentation of exogenous antigen by mouse bone marrow-derived dendritic cells (DC) and splenic B cells. DC presented to class I-restricted transgenic T cells femtomolar concentrations of antigens from liposomes targeted to the IgG Fc receptor. Targeting these liposomes to surface immunoglobulin did not permit B cells to stimulate class I-restricted responses.

VPREB3: cDNA characterization and expression in human and chromosome mapping in human and mouse.

The pre-B cell receptor (pre-BCR) regulates pre-B cell expansion and allelic exclusion at the immunoglobulin (Ig) heavy chain locus and mediates the selection of Ig heavy chain variable gene segments. During the early phase of pre-BCR assembly in the mouse, the membrane Ig mu heavy chain transiently associates with the VPREB3 protein in the endoplasmic reticulum. Here, we present the human VPREB3 cDNA sequence and its B cell-specific expression in hematopoietic cell lines.