Cytotoxic CD4+ T cells use granulysin to kill Cryptococcus neoformans, and activation of this pathway is defective in HIV patients

Abstract

IntroductionCD4 ϩ T cells function as both regulators and effectors in the immune response. 1 CD4 ϩ T cells produce cytokines, providing help in the immune response to pathogens, and have often been referred to as Th cells. 2 However, over the past 2 decades, the ability of CD4 ϩ T cells to become cytotoxic has been reported in both mice and humans against alloantigens, microbe-infected cells, and cancer. 1,[3][4][5][6][7][8] Previous studies in mice demonstrate that CD4 ϩ cytotoxic T lymphocytes (CTLs) cause target cell lysis primarily via Fas/FasL interactions. [9][10][11][12] Later, studies on human CD4 ϩ T cells demonstrated that CD4 ϩ CTLs also use perforin and granzymebased granule exocytosis. [13][14][15] This pathway involves disruption of the target cell membrane through extracellular Ca 2ϩ -dependent polymerization of perforin, accompanied by uptake of granzymes, which are responsible for downstream activation of caspases and subsequent DNA fragmentation, resulting in target cell apoptosis.Although the importance of CTL-mediated killing of tumor or microbe-infected cells is well established, direct CTL-mediated microbial killing has been underinvestigated. Evidence has suggested that CTLs have very potent direct antimicrobial activity. This activity requires cell contact and involves regulated exocytosis of granules from the cytoplasm of the effector cells. 16 Perforin and granzymes are well-known cytolytic molecules 17 ; however, granulysin is less well recognized. 16,18 The human granulysin cDNA was originally isolated by subtractive hybridization in a search for T-cell-specific molecules expressed "late" (3-5 days) after activation. 19 Granulysin, a member of the saposin-like protein family of lipid binding proteins, colocalizes with perforin and granzymes in the cytolytic granules of human CTLs and natural killer (NK) cells. [20][21][22] Two major protein products of granulysin, 15 and 9 kDa, are detected in CTL and NK cells. 22 It is apparent that the 9-kDa form is processed from the larger, precursor 15-kDa form, 22 and recombinant 9-kDa granulysin disrupts artificial liposomes and cell membranes, damages mitochondria, and activates caspase 9 to induce apoptosis. 23,24 Additionally, granulysin exhibits potent cytotoxic activity against a broad panel of microbial pathogens and tumor cells. 18,20 Our previous observation, using Cryptococcus neoformans, one of the most important microbial pathogens in patients with compromised CD4 ϩ T-cell-dependent immunity, such as the acquired immunodeficiency syndrome (AIDS), [25][26][27] indicated that granulysin is required for the CD8 ϩ T-cell-mediated direct antifungal activity. 28 By contrast, NK cells use perforin, rather than granulysin, as the effector molecule to kill C neoformans. 29 Although CD4 ϩ and CD8 ϩ T cells and NK cells are important in cryptococcal host defense, 30-32 defective CD4 ϩ T cells are clearly the major risk factor for disease in human immunodeficiency virus (HIV)-infected patients. 33 Previous studies have demonstrated that IL-2-ac...

Granulysin Production and Anticryptococcal Activity Is Dependent upon a Far Upstream Enhancer That Binds STAT5 in Human Peripheral Blood CD4+ T Cells

“…Within this cytokine family, IL-2, IL-15, and IL-21 promote granulysin and cytotoxic activity of CTLs (19,20). We have demonstrated that IL-2 and IL-15 regulate the granulysindependent killing of the fungus C. neoformans by human peripheral blood CD4 + and CD8 + T cells (12,18). The role of g c was extended by the observation that IL-21 and IL-15 could activate granulysin expression within CD8 + CD45RO + T cells (21).…”

“…We have demonstrated that granulysin production by IL-2-treated CD4 + T cells is dependent on activation of the Jak3 and STAT5 signaling pathways (23). Moreover, defects in this pathway lead to impaired anticryptococcal activity in HIV-infected patients (12). Additionally, granulysin induction occurred in CD8 + T cells via Jak/STATdependent signaling pathways after activation with IL-15 or IL-21 (21).…”

“…However, the genetic regulation of granulysin expression, especially in human CTLs, is still elusive. Previous studies have demonstrated that IL-2 activated human peripheral blood CD4 + T cell signals via Jak3/STAT5, downstream of the IL-2/IL-2R signaling pathway, leading to production of granulysin (12,23). Because the signaling pathway and the resultant microbicidal activity is defective in HIV-infected patients, the mechanism by which STAT5 leads to granulysin expression is of great interest.…”

“…It is apparent that the 9-kDa form is processed from the larger, precursor 15-kDa form (3), and recombinant 9-kDa granulysin disrupts artificial liposomes and cell membranes, damages mitochondria, and activates caspase 9 to induce apoptosis (4,5). The 9-kDa granulysin exhibits potent cytotoxic activity against a variety of tumors (6)(7)(8) and microbes (9,10), including Cryptococcus neoformans (11,12), Mycobacterium tuberculosis (13), and Plasmodium falciparum (14).…”

Perforin level in CD4+ T cells from patients with systemic lupus erythematosus

“…The PRF1 gene is expressed primarily in NK cells and effector CD8 + T cells [31,32]. PRF1 is also expressed in a subpopulation of cytotoxic CD4 + T cells, which may combat various pathogens (33)(34)(35)(36)(37)(38)(39). An increased number of cytotoxic CD4 + T cells has been observed in patients with multiple sclerosis, rheumatoid arthritis, inflammatory bowel disease, and acute coronary syndromes [20][21][22][23].…”