Cancer cells are commonly more resistant to cell death activated by the membranolytic protein complex C5b-9. Several surface-expressed and intracellular proteins that protect cells from ...complement-dependent cytotoxicity (CDC) have been identified. In this study, we investigated the function of heat shock protein 90 (Hsp90), an essential and ubiquitously expressed chaperone, overexpressed in cancer cells, in C5b-9-induced cell death. As shown, inhibition of Hsp90 with geldanamycin or radicicol is enhancing sensitivity of K562 erythroleukemia cells to CDC. Similarly, Hsp90 inhibition confers in Ramos B cell lymphoma cells elevated sensitivity to treatment with rituximab and complement. C5b-9 deposition is elevated on geldanamycin-treated cells. Purified Hsp90 binds directly to C9 and inhibits zinc-induced C9 polymerization, indicating that Hsp90 may act directly on the C5b-9 complex. Mortalin, also known as stress protein 70 or GRP75, is a mitochondrial chaperone that confers resistance to CDC. The postulated cooperation between Hsp90 and mortalin in protection from CDC was tested. Geldanamycin failed to sensitize toward CDC cells with knocked down mortalin. Direct binding of Hsp90 to mortalin was shown by co-immunoprecipitation in cell extracts after triggering with complement as well as by using purified recombinant proteins. These results provide an insight into the protective mechanisms utilized by cancer cells to evade CDC. They suggest that Hsp90 protects cells from CDC by inhibiting, together with mortalin, C5b-9 assembly and/or stability at the plasma membrane.
Mortalin/GRP75 is a ubiquitous mitochondrial chaperone related to the cytosolic heat shock protein 70 (HSP70). It protects cells from senescence and apoptosis and is overexpressed in cancer cells. ...Cell resistance to complement‐dependent cytotoxicity depends on mortalin and during complement attack mortalin is released from cells. Our goal was to determine whether cancer patients have circulating mortalin in blood. The significance of mortalin in blood to survival prospects of colorectal cancer patients was evaluated. Occurrence of extracellular soluble HSP70 (sHSP70) is documented. We developed a sensitive ELISA for mortalin. The association between mortalin level and survival was subjected to the Cox proportional hazards analysis (univariate and multivariate analyses). Mortalin concentration in serum of colorectal cancer patients was 10–214 ng/ml. Survival data of the patients were known from an earlier study of sHSP70 in these samples. Cox regression analysis indicated that high mortalin (>60 ng/ml) is a risk factor for shorter survival. Serum levels of sHSP70 and mortalin in patients were independent variables. Concurrence of high sHSP70 and mortalin was associated with rapid disease progression (HR = 4, 2.04–8.45, p < 0.001). Addition of high sHSP70 and mortalin to a baseline model of age, sex and TNM stage, significantly (p < 0.001) enhanced the risk score to 8 (3.26–20.46). This is the first demonstration of circulating mortalin in cancer patients. Analysis of mortalin in blood, and even more so of mortalin and sHSP70, adds a high prognostic value to the TNM stage and will identify colorectal cancer patients at high risk of poor survival.
What's new?
Mortalin is a ubiquitous mitochondrial chaperone protein that protects cells from apoptosis and is overexpressed in cancer cells. Here, a novel ELISA developed for the detection and quantification of circulating mortalin in sera of colorectal cancer patients is described. The assay revealed that high circulating levels of both mortalin and the closely related cytosolic protein HSP70 are indicative of short patient survival. The results suggest that mortalin, HSP70, and TNM staging are independent survival determinants for colorectal cancer.
IL-13 and IL-4 are potent mediators of type 2-associated inflammation such as those found in atopic dermatitis (AD). IL-4 shares overlapping biological functions with IL-13, a finding that is mainly ...explained by their ability to signal via the type 2 IL-4 receptor (R), which is composed of IL-4Rα in association with IL-13Rα1. Nonetheless, the role of the type 2 IL-4R in AD remains to be clearly defined. Induction of two distinct models of experimental AD in
mice, which lack the type 2 IL-4R, revealed that dermatitis, including ear and epidermal thickening, was dependent on type 2 IL-4R signaling. Expression of TNF-α was dependent on the type 2 IL-4R, whereas induction of IL-4, IgE, CCL24, and skin eosinophilia was dependent on the type 1 IL-4R. Neutralization of IL-4, IL-13, and TNF-α as well as studies in bone marrow-chimeric mice revealed that dermatitis, TNF-α, CXCL1, and CCL11 expression were exclusively mediated by IL-13 signaling via the type 2 IL-4R expressed by nonhematopoietic cells. Conversely, induction of IL-4, CCL24, and eosinophilia was dependent on IL-4 signaling via the type 1 IL-4R expressed by hematopoietic cells. Last, we pharmacologically targeted IL-13Rα1 and established a proof of concept for therapeutic targeting of this pathway in AD. Our data provide mechanistic insight into the differential roles of IL-4, IL-13, and their receptor components in allergic skin and highlight type 2 IL-4R as a potential therapeutic target in AD and other allergic diseases such as asthma and eosinophilic esophagitis.
IL-4 receptor (R) α, the common receptor chain for IL-4 and IL-13, is a critical component in IL-4- and IL-13-mediated signaling and subsequent effector functions such as those observed in type 2 ...inflammatory responses. Nonetheless, the existence of intrinsic pathways capable of amplifying IL-4Rα-induced responses remains unknown. In this study, we identified the myeloid-associated Ig receptor CD300f as an IL-4-induced molecule in macrophages. Subsequent analyses demonstrated that CD300f was colocalized and physically associated with IL-4Ra. Using ... cells and receptor cross-linking experiments, we established that CD300f amplified IL-4Rα-induced responses by augmenting IL-4/IL-13-induced signaling, mediator release, and priming. Consistently, IL-4- and aeroallergen-treated ... mice displayed decreased IgE production, chemokine expression, and inflammatory cell recruitment. Impaired responses in ... mice were not due to the inability to generate a proper Th2 response, because IL-4/IL-13 levels were markedly increased in allergen-challenged ... mice, a finding that is consistent with decreased cytokine consumption. Finally, CD300f expression was increased in monocytes and eosinophils obtained from allergic rhinitis patients. Collectively, our data highlight a previously unidentified role for CD300f in IL-4Rα-induced immune cell responses. These data provide new insights into the molecular mechanisms governing IL-4Ra-induced responses, and may provide new therapeutic tools to target IL-4 in allergy and asthma. (ProQuest: ... denotes formulae/symbols omitted.)
Significance IL-4 receptor (R) α is a critical component in IL-4â and IL-13âmediated signaling and subsequent effector functions such as those observed in allergy. Thus, it is a primary ...therapeutic target in diseases such as atopic dermatitis and asthma. Despite extensive studies, it is unknown whether an additional receptor system exists that may act to amplify IL-4Rα signaling and subsequent IL-4/IL-13âinduced responses. We now report that CD300f is physically associated with IL-4Rα and potently amplifies IL-4Rαâinduced responses in vitro and in vivo. Our results establish CD300f as a previously unidentified IL-4Rα coreceptor. To the best of our knowledge, this is the first report of an additional receptor that serves to amplify the IL-4 signaling pathway.
IL-4 receptor (R) α, the common receptor chain for IL-4 and IL-13, is a critical component in IL-4â and IL-13âmediated signaling and subsequent effector functions such as those observed in type 2 inflammatory responses. Nonetheless, the existence of intrinsic pathways capable of amplifying IL-4Rαâinduced responses remains unknown. In this study, we identified the myeloid-associated Ig receptor CD300f as an IL-4âinduced molecule in macrophages. Subsequent analyses demonstrated that CD300f was colocalized and physically associated with IL-4Rα. Using Cd300f â»/â» cells and receptor cross-linking experiments, we established that CD300f amplified IL-4Rαâinduced responses by augmenting IL-4/IL-13âinduced signaling, mediator release, and priming. Consistently, IL-4â and aeroallergen-treated Cd300f â»/â» mice displayed decreased IgE production, chemokine expression, and inflammatory cell recruitment. Impaired responses in Cd300f â»/â» mice were not due to the inability to generate a proper Th2 response, because IL-4/IL-13 levels were markedly increased in allergen-challenged Cd300f â»/â» mice, a finding that is consistent with decreased cytokine consumption. Finally, CD300f expression was increased in monocytes and eosinophils obtained from allergic rhinitis patients. Collectively, our data highlight a previously unidentified role for CD300f in IL-4Rαâinduced immune cell responses. These data provide new insights into the molecular mechanisms governing IL-4Rαâinduced responses, and may provide new therapeutic tools to target IL-4 in allergy and asthma.
The chemokine CCL5 (RANTES) plays active promalignancy roles in breast malignancy. The secretion of CCL5 by breast tumor cells is an important step in its tumor-promoting activities; therefore, ...inhibition of CCL5 secretion may have antitumorigenic effects. We demonstrate that, in breast tumor cells, CCL5 secretion necessitated the trafficking of CCL5-containing vesicles on microtubules from the endoplasmic reticulum (ER) to the post-Golgi stage, and CCL5 release was regulated by the rigidity of the actin cytoskeleton. Focusing on the 40s loop of CCL5, we found that the
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sequence of CCL5 was indispensable for its inclusion in motile vesicles, and for its secretion. The TRKN-mutated chemokine reached the Golgi, but trafficked along the ER-to-post-Golgi route differently than the wild-type (WT) chemokine. Based on the studies showing that the 40s loop of CCL5 mediates its binding to glycosaminoglycans (GAG), we analyzed the roles of GAG in regulating CCL5 secretion. TRKN-mutated CCL5 had lower propensity for colocalization with GAG in the Golgi compared to the WT chemokine. Secretion of WT CCL5 was significantly reduced in CHO mutant cells deficient in GAG synthesis, and the WT chemokine acquired an ER-like distribution in these cells, similar to that of TRKN-mutated CCL5 in GAG-expressing cells. The release of WT CCL5 was also reduced after inhibition of GAG presence/synthesis by intracellular expression of heparanase, inhibition of GAG sulfation, and sulfate deprivation. The need for a
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motif and for a GAG-mediated process in CCL5 secretion may enable the future design of modalities that prevent CCL5 release by breast tumor cells.
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