Cytotoxic cells kill intracellular bacteria through granulysin-mediated delivery of granzymes.

When killer lymphocytes recognize infected cells, perforin delivers cytotoxic proteases (granzymes) into the target cell to trigger apoptosis. What happens to intracellular bacteria during this process is unclear. Human, but not rodent, cytotoxic granules also contain granulysin, an antimicrobial peptide.

The reproductive phenotype of mice null for transcription factor Krüppel-like factor 13 suggests compensatory function of family member Krüppel-like factor 9 in the peri-implantation uterus.

The ovarian hormones estrogen and progesterone promote uterine receptivity and successful pregnancy through their cognate receptors functioning in concert with context-dependent nuclear coregulators. Previously, we showed that the transcription factor Krüppel-like factor (KLF) 9 is a progesterone receptor (PGR) coactivator in the uterus and that mice null for Klf9 exhibit subfertility and reduced progesterone sensitivity. The highly related family member KLF13 displays increased expression in uteri of pregnant and nonpregnant Klf9 null mice and similarly regulates PGR-mediated transactivation in endometrial stromal cells.

Fbw7γ-mediated degradation of KLF13 prevents RANTES expression in resting human but not murine T lymphocytes.

RANTES (CCL5) is a chemokine implicated in many human diseases. We previously showed that the transcription factor Kruppel-like factor 13 (KLF13) controls the late (3-5 days after activation) expression of RANTES in T lymphocytes and that KLF13 itself is translationally regulated through the 5'-untranslated region of its mRNA. Here, we show that KLF13 levels are further regulated by ubiquitination and degradation.

15 kDa granulysin causes differentiation of monocytes to dendritic cells but lacks cytotoxic activity.

Granulysin is expressed as two isoforms by human cytotoxic cells: a single mRNA gives rise to 15 kDa granulysin, a portion of which is cleaved to a 9 kDa protein. Studies with recombinant 9 kDa granulysin have demonstrated its cytolytic and proinflammatory properties, but much less is known about the biologic function of the 15 kDa isoform. In this study, we show that the subcellular localization and functions of 9 and 15 kDa granulysin are largely distinct.

15 kDa Granulysin versus GM-CSF for monocytes differentiation: analogies and differences at the transcriptome level.

Background: Granulysin is an antimicrobial and proinflammatory protein with several isoforms. While the 9 kDa isoform is a well described cytolytic molecule with pro-inflammatory activity, the functions of the 15 kDa isoform is less well understood. Recently it was shown that 15 kDa Granulysin can act as an alarmin that is able to activate monocytes and immature dendritic cells.

KLF13 sustains thymic memory-like CD8(+) T cells in BALB/c mice by regulating IL-4-generating invariant natural killer T cells.

"Memory-like T cells" are a subset of thymic cells that acquire effector function through the maturation process rather than interaction with specific antigen. Disruption of genes encoding T cell signaling proteins or transcription factors have provided insights into the differentiation of such cells. In this study, we show that in BALB/c, but not C57BL/6, mice, a large portion of thymic CD4(-)CD8(+) T cells exhibit a memory-like phenotype.

Granulysin delivered by cytotoxic cells damages endoplasmic reticulum and activates caspase-7 in target cells.

Granulysin is a human cytolytic molecule present in cytotoxic granules with perforin and granzymes. Recombinant 9-kDa granulysin kills a variety of microbes, including bacteria, yeast, fungi, and parasites, and induces apoptosis in tumor cells by causing intracellular calcium overload, mitochondrial damage, and activation of downstream caspases. Reasoning that granulysin delivered by cytotoxic cells may work in concert with other molecules, we crossed granulysin transgenic (GNLY(+/-)) mice onto perforin (perf)- or granzyme B (gzmb)-deficient mice to examine granulysin-mediated killing in a more physiologic whole-cell system.

Monocyte-derived DC maturation strategies and related pathways: a transcriptional view.

Ex vivo production of highly stimulator mature dendritic cells (DCs) for cellular therapy has been used to treat different pathological conditions with the aim of inducing a specific immune response. In the last decade, several protocols have been developed to mature monocyte-derived DCs: each one has led to the generation of DCs showing different phenotypes and stimulatory abilities, but it is not yet known which one is the best for inducing effective immune responses. We grouped several different maturation protocols according to the downstream pathways they activated and reviewed the shared features at a transcriptomic level to reveal the potential of DCs matured by each protocol to develop Th-polarized immune responses.

Induction of granulysin and perforin cytolytic mediator expression in 10-week-old infants vaccinated with BCG at birth.

Background: While vaccination at birth with Mycobacterium bovis Bacilli Calmette-Guérin (BCG) protects against severe childhood tuberculosis, there is no consensus as to which components of the BCG-induced immune response mediate this protection. However, granulysin and perforin, found in the granules of cytotoxic T lymphocytes and Natural Killer (NK) cells, can kill intracellular mycobacteria and are implicated in protection against Mycobacterium tuberculosis.

Methods: We compared the cellular expression of granulysin and perforin cytolytic molecules in cord blood and peripheral blood from 10-week-old infants vaccinated at birth with either Japanese or Danish BCG, administered either intradermally or percutaneously.

Granulysin production and anticryptococcal activity is dependent upon a far upstream enhancer that binds STAT5 in human peripheral blood CD4+ T cells.

Previous studies have demonstrated that STAT5 is critical for expression of granulysin and antimicrobial activity. Because the signaling pathway and the resultant microbicidal activity are defective in HIV-infected patients, the mechanism by which STAT5 leads to granulysin expression is of great interest. In the current study, IL-2-stimulated CRL-2105 CD4(+) T cells expressed granulysin and killed Cryptococcus neoformans similar to primary CD4(+) T cells.

Expression and purification of 15 kDa granulysin utilizing an insect cell secretion system.

Granulysin is an antimicrobial and proinflammatory protein expressed in activated human T cells and natural killer cells. A single mRNA produces the 15 kDa isoform which is then cleaved at the amino and carboxy termini to produce the 9 kDa isoform. Recombinant 9 kDa granulysin has been studied in detail but little is known about the function of the 15 kDa isoform, and no protocol has been published describing expression and purification of this form.

Granulysin activates antigen-presenting cells through TLR4 and acts as an immune alarmin.

Granulysin (GNLY), an antimicrobial protein present in the granules of human cytotoxic T lymphocytes and natural killer (NK) cells, is produced as an intact 15-kDa form that is cleaved to yield a 9-kDa form. Alarmins are endogenous mediators that can induce recruitment and activation of antigen-presenting cells (APCs) and consequently promote the generation of immune response. We hypothesized that GNLY might function as an alarmin.

Ten years of the Immune Tolerance Network: an integrated clinical research organization.

The U.S. National Institutes of Health Roadmap and the U.

XCL1 enhances regulatory activities of CD4+ CD25(high) CD127(low/-) T cells in human allergic asthma.

Chemokine-mediated recruitment of regulatory cell subsets to the airway during inflammation and enhancement of their activities are potential strategies for therapeutic development in allergic asthma (AA). In this study, we aim to explore the role of XCL1, a chemokine associated with immune suppression and allergy, on CD4(+)CD25(high)CD127(low/-) regulatory T cell (Treg) function in AA. Flow cytometry and PCR analysis showed a reduction in XCL1 and XCR1 expression in AA Treg compared with healthy control and nonallergic asthmatic counterparts.

Late expression of granulysin by microbicidal CD4+ T cells requires PI3K- and STAT5-dependent expression of IL-2Rbeta that is defective in HIV-infected patients.

Granulysin is a cytolytic effector molecule used by lymphocytes to kill tumor and microbial cells. Regulation of granulysin production is complex. A significant delay (5 days) following stimulation of CD4(+) T cells with IL-2 occurs before granulysin is produced.

In vitro and in vivo antimicrobial activity of granulysin-derived peptides against Vibrio cholerae.

Objectives: To determine the antibacterial activity of synthetic peptides derived from the cationic antimicrobial peptide granulysin against Vibrio cholerae.

Methods: The antibacterial activity of granulysin-derived peptides was assessed in vitro by microtitre and cfu assays. Toxicity against human peripheral blood mononuclear cells (PBMCs) was measured by propidium iodide uptake and haemolysis by measuring the levels of haemoglobin released after incubation of red blood cells (RBCs) with granulysin peptides.

IL-24 modulates IFN-gamma expression in patients with tuberculosis.

IL-24 is a newly described member of the IL-10 family. We previously demonstrated that PBMC from TB patients exhibited low levels of IL-24 and IFN-gamma compared to subjects with latent tuberculosis infection (LTBI). In order to investigate the role of IL-24 in IFN-gamma expression in TB patients, we stimulated PBMC from individuals with LTBI or TB patients with the Mtb-specific antigen, early secretory antigenic target-6 (ESAT-6) and measured cytokine expression using quantitative real-time PCR (qPCR).

IL-9 is associated with an impaired Th1 immune response in patients with tuberculosis.

Although a defective Th1 response has been demonstrated in patients infected with Mycobacterium tuberculosis (Mtb), the mechanisms leading to this defect are not well understood. To study the immune response to Mtb infection, we stimulated PBMC from individuals with latent tuberculosis infection (LTBI) or patients with tuberculosis (TB) with the Mtb specific antigen early secretory antigenic target-6 (ESAT-6). mRNAs for a panel of cytokines were measured using quantitative real-time PCR (qPCR).

Altered phosphorylated signal transducer and activator of transcription profile of CD4+CD161+ T cells in asthma: modulation by allergic status and oral corticosteroids.

Background: Asthma is a complex immunologic disorder linked to altered cytokine signaling.

Objective: We tested whether asthmatic patients showed any change in cytokine-dependent signal transducer and activator of transcription (STAT) levels, focusing on the central/effector-memory CD4(+)CD161(+) subset, which represents 15% to 25% of circulating T cells.

Methods: We quantified intracellular levels of active phosphorylated STAT (phospho-STAT) 1, 3, 5, and 6 by means of flow cytometry, without any activation or expansion.

CD96 is a leukemic stem cell-specific marker in human acute myeloid leukemia.

Permanent cure of acute myeloid leukemia (AML) by chemotherapy alone remains elusive for most patients because of the inability to effectively eradicate leukemic stem cells (LSCs), the self-renewing component of the leukemia. To develop therapies that effectively target LSC, one potential strategy is to identify cell surface markers that can distinguish LSC from normal hematopoietic stem cells (HSCs). In this study, we employ a signal sequence trap strategy to isolate cell surface molecules expressed on human AML-LSC and find that CD96, which is a member of the Ig gene superfamily, is a promising candidate as an LSC-specific antigen.

Biophysical analysis of the interaction of granulysin-derived peptides with enterobacterial endotoxins.

To combat infections by Gram-negative bacteria, it is not only necessary to kill the bacteria but also to neutralize pathogenicity factors such as endotoxin (lipopolysaccharide, LPS). The development of antimicrobial peptides based on mammalian endotoxin-binding proteins is a promising tool in the fight against bacterial infections, and septic shock syndrome. Here, synthetic peptides derived from granulysin (Gra-pep) were investigated in microbiological and biophysical assays to understand their interaction with LPS.

Interaction of PRP4 with Kruppel-like factor 13 regulates CCL5 transcription.

Activation of resting T lymphocytes initiates differentiation into mature effector cells over 3-7 days. The chemokine CCL5 (RANTES) and its major transcriptional regulator, Krüppel-like factor 13 (KLF13), are expressed late (3-5 days) after activation in T lymphocytes. Using yeast two-hybrid screening of a human thymus cDNA library, PRP4, a serine/threonine protein kinase, was identified as a KLF13-binding protein.