Simian Immunodeficiency Virus-Producing Cells in Follicles Are Partially Suppressed by CD8+ Cells In Vivo.

Unlabelled: Human immunodeficiency virus (HIV)- and simian immunodeficiency virus (SIV)-specific CD8 T cells are typically largely excluded from lymphoid B cell follicles, where HIV- and SIV-producing cells are most highly concentrated, indicating that B cell follicles are somewhat of an immunoprivileged site. To gain insights into virus-specific follicular CD8 T cells, we determined the location and phenotype of follicular SIV-specific CD8 T cells in situ, the local relationship of these cells to Foxp3 cells, and the effects of CD8 depletion on levels of follicular SIV-producing cells in chronically SIV-infected rhesus macaques. We found that follicular SIV-specific CD8 T cells were able to migrate throughout follicular areas, including germinal centers.

Comparison of Vibratome and Compresstome sectioning of fresh primate lymphoid and genital tissues for in situ MHC-tetramer and immunofluorescence staining.

Background: For decades, the Vibratome served as a standard laboratory resource for sectioning fresh and fixed tissues. In skilled hands, high quality and consistent fresh unfixed tissue sections can be produced using a Vibratome but the sectioning procedure is extremely time consuming. In this study, we conducted a systematic comparison between the Vibratome and a new approach to section fresh unfixed tissues using a Compresstome.

Single domain intrabodies against WASP inhibit TCR-induced immune responses in transgenic mice T cells.

Intrabody technology provides a novel approach to decipher the molecular mechanisms of protein function in cells. Single domains composed of only the variable regions (V(H) or V(L)) of antibodies are the smallest recombinant antibody fragments to be constructed thus far. In this study, we developed transgenic (Tg) mice expressing the V(H) or V(L) single domains derived from a monoclonal antibody raised against the N-terminal domain of Wiskott-Aldrich syndrome protein (WASP), which is an adaptor molecule in immune cells.

Identification of Fyn as the binding partner for the WASP N-terminal domain in T cells.

Wiskott-Aldrich syndrome protein (WASP) plays important roles in TCR signaling. In transgenic (Tg) mice, over-expression of the WASP N-terminal region (exons 1-5) including the enabled/vasodilator-stimulated phosphoprotein (Ena/VASP) homology 1 (EVH1) domain and anti-WASP-EVH1 single-chain variable fragment (scFv) intracellular expressed antibodies (intrabodies) impairs IL-2 production in activated T cells. However, it largely remains unknown that how this domain transduces TCR signaling.

Failure of mycoplasma lipoprotein MALP-2 to induce NK cell activation through dendritic cell TLR2.

Macrophage-activating lipopeptide 2 (MALP-2), a mycoplasmal diacylated lipopeptide with palmitic acid moiety (Pam2), activates Toll-like receptor (TLR) 2 to induce inflammatory cytokines. TLR2 is known to mature myeloid dendritic cells (mDC) to drive mDC contact-mediated natural killer (NK) cell activation. Here we tested if MALP-2 activates NK cells through stimulation of TLR2 on mDC.

The peptide sequence of diacyl lipopeptides determines dendritic cell TLR2-mediated NK activation.

Natural killer (NK) cells are lymphocyte effectors that are activated to control certain microbial infections and tumors. Many NK-activating and regulating receptors are involved in regulating NK cell function. In addition, activation of naïve NK cells is fundamentally triggered by cytokines or myeloid dendritic cells (mDC) in various modes.

Cytoplasmic expression and specific binding of the VH/VL single domain intrabodies in transfected NIH3T3 cells.

Intracellularly expressed antibody fragments (intrabodies) have been utilized as powerful tools not only for clinical applications but also for the functional analysis of proteins inside the cell. Among several types of intrabodies developed so far, single domain types composed of only the variable regions (V(H) or V(L)) of antibodies are the smallest and thus the easiest to design. In this study, four types of single domain intrabodies were evaluated against a cytosolic protein, Wiskott-Aldrich syndrome protein (WASP), in gene-transfected NIH3T3 cells.

Impaired LPS-induced signaling in microglia overexpressing the Wiskott-Aldrich syndrome protein N-terminal domain.

Wiskott-Aldrich syndrome protein (WASP) plays important roles in TCR signaling, but its roles in signal transduction in innate immune cells have not been well characterized. As microglia are the primary immune effector cells in the brain, WASP may possibly have important roles in microglial activation, such as production of inflammatory and anti-inflammatory cytokines and neurotoxic factors. Here, we established a microglial cell line from WASP dominant-negative transgenic (Tg) mice overexpressing the N-terminal enabled/vasodilator-stimulated phosphoprotein homology 1 (EVH1) domain.

Intrabodies against the EVH1 domain of Wiskott-Aldrich syndrome protein inhibit T cell receptor signaling in transgenic mice T cells.

Intracellularly expressed antibodies (intrabodies) have been used to inhibit the function of various kinds of protein inside cells. However, problems with stability and functional expression of intrabodies in the cytosol remain unsolved. In this study, we show that single-chain variable fragment (scFv) intrabodies constructed with a heavy chain variable (V(H)) leader signal sequence at the N-terminus were translocated from the endoplasmic reticulum into the cytosol of T lymphocytes and inhibited the function of the target molecule, Wiskott-Aldrich syndrome protein (WASP).

Scarabaecin, a novel cysteine-containing antifungal peptide from the rhinoceros beetle, Oryctes rhinoceros.

A novel antifungal peptide, scarabaecin (4080Da), was isolated from the coconut rhinoceros beetle, Oryctes rhinoceros. Scarabaecin cDNA was cloned by reverse transcriptase-polymerase chain reactions (RT-PCR) using a primer based on the N-terminal amino acid sequence. The amino acid sequence deduced from scarabaecin cDNA showed no significant similarity to those of reported proteins.