Natural and cryptic peptides dominate the immunopeptidome of atypical teratoid rhabdoid tumors

BACKGROUND: Atypical teratoid/rhabdoid tumors (AT/RT) are highly aggressive CNS tumors of infancy and early childhood. Hallmark is the surprisingly simple genome with inactivating mutations or deletions in the SMARCB1 gene as the oncogenic driver. Nevertheless, AT/RTs are infiltrated by immune cells and even clonally expanded T cells. However, it is unclear which epitopes T cells might recognize on AT/RT cells. METHODS: Here, we report a comprehensive mass spectrometry (MS)-based analysis of naturally presented human leukocyte antigen (HLA) class I and class II ligands on 23 AT/RTs. MS data were validated by matching with a human proteome dataset and exclusion of peptides that are part of the human benignome. Cryptic peptide ligands were identified using Peptide-PRISM. RESULTS: Comparative HLA ligandome analysis of the HLA ligandome revealed 55 class I and 139 class II tumor-exclusive peptides. No peptide originated from the SMARCB1 region. In addition, 61 HLA class I tumor-exclusive peptide sequences derived from non-canonically translated proteins. Combination of peptides from natural and cryptic class I and class II origin gave optimal representation of tumor cell compartments. Substantial overlap existed with the cryptic immunopeptidome of glioblastomas, but no concordance was found with extracranial tumors. More than 80% of AT/RT exclusive peptides were able to successfully prime CD8(+) T cells, whereas naturally occurring memory responses in AT/RT patients could only be detected for class II epitopes. Interestingly, >50% of AT/RT exclusive class II ligands were also recognized by T cells from glioblastoma patients but not from healthy donors. CONCLUSIONS: These findings highlight that AT/RTs, potentially paradigmatic for other pediatric tumors with a low mutational load, present a variety of highly immunogenic HLA class I and class II peptides from canonical as well as non-canonical protein sources. Inclusion of such cryptic peptides into therapeutic vaccines would enable an optimized mapping of the tumor cell surface, thereby reducing the likelihood of immune evasion

The expression of nuclear and membrane estrogen receptors in the European eel throughout spermatogenesis

[EN] Estradiol (E-2) can bind to nuclear estrogen receptors (ESR) or membrane estrogen receptors (GPER). While mammals possess two nuclear ESRs and one membrane GPER, the European eel, like most other teleosts, has three nuclear ESRs and two membrane GPERs, as the result of a teleost specific genome duplication. In the current study, the expression of the three nuclear ESRs (ESR1, ESR2a and ESR2b) and the two membrane GPERs (GPERa and GPERb) in the brain-pituitary-gonad (BPG) axis of the European eel was measured, throughout spermatogenesis. The eels were first transferred from freshwater (FW) to seawater (SW), inducing parallel increases in E2 plasma levels and the expression of ESRs. This indicates that salinity has a stimulatory effect on the E-2 signalling pathway along the BPG axis. Stimulation of sexual maturation by weekly injections of human chorionic gonadotropin (hCG) induced a progressive decrease in E-2 plasma levels, and different patterns of expression of ESRs and GPERs in the BPG axis. The expression of nuclear ESRs increased in some parts of the brain, suggesting a possible upregulation due to a local production of E-2. In the testis, the highest expression levels of the nuclear ESRs were observed at the beginning of spermatogenesis, possibly mediating the role of E2 as spermatogonia renewal factor, followed by a sharply decrease in the expression of ESRs. Conversely, there was a marked increase observed in the expression of both membrane GPERs throughout spermatogenesis, suggesting they play a major role in the final stages of spermatogenesis.Funded by the Spanish Ministry of Science and Innovation (REPRO-TEMP project; AGL2013-41646-R) and IMPRESS (Marie Sklodowska-Curie Actions; Grant agreement no: 642893). M.C. Vilchez has a predoctoral grant from UPV PAID Programme (2011-S2-02-6521), M. Morini has a predoctoral grant from Generalitat Valenciana (Programa Grisolia). D.S. Penaranda was supported by MICINN and UPV (PTA2011-4948-1).Morini, M.; Peñaranda, D.; Vilchez Olivencia, MC.; Tveiten, H.; Lafont, A.; Dufour, S.; Pérez Igualada, LM.... (2017). The expression of nuclear and membrane estrogen receptors in the European eel throughout spermatogenesis. Comparative Biochemistry and Physiology Part A Molecular & Integrative Physiology. 203:91-99. doi:10.1016/j.cbpa.2016.08.020S919920

Global patterns in endemicity and vulnerability of soil fungi

Fungi are highly diverse organisms, which provide multiple ecosystem services. However, compared with charismatic animals and plants, the distribution patterns and conservation needs of fungi have been little explored. Here, we examined endemicity patterns, global change vulnerability and conservation priority areas for functional groups of soil fungi based on six global surveys using a high-resolution, long-read metabarcoding approach. We found that the endemicity of all fungi and most functional groups peaks in tropical habitats, including Amazonia, Yucatan, West-Central Africa, Sri Lanka, and New Caledonia, with a negligible island effect compared with plants and animals. We also found that fungi are predominantly vulnerable to drought, heat and land-cover change, particularly in dry tropical regions with high human population density. Fungal conservation areas of highest priority include herbaceous wetlands, tropical forests, and woodlands. We stress that more attention should be focused on the conservation of fungi, especially root symbiotic arbuscular mycorrhizal and ectomycorrhizal fungi in tropical regions as well as unicellular early-diverging groups and macrofungi in general. Given the low overlap between the endemicity of fungi and macroorganisms, but high conservation needs in both groups, detailed analyses on distribution and conservation requirements are warranted for other microorganisms and soil organisms

Global patterns in endemicity and vulnerability of soil fungi

Fungi are highly diverse organisms, which provide multiple ecosystem services. However, compared with charismatic animals and plants, the distribution patterns and conservation needs of fungi have been little explored. Here, we examined endemicity patterns, global change vulnerability and conservation priority areas for functional groups of soil fungi based on six global surveys using a high-resolution, long-read metabarcoding approach. We found that the endemicity of all fungi and most functional groups peaks in tropical habitats, including Amazonia, Yucatan, West-Central Africa, Sri Lanka, and New Caledonia, with a negligible island effect compared with plants and animals. We also found that fungi are predominantly vulnerable to drought, heat and land-cover change, particularly in dry tropical regions with high human population density. Fungal conservation areas of highest priority include herbaceous wetlands, tropical forests, and woodlands. We stress that more attention should be focused on the conservation of fungi, especially root symbiotic arbuscular mycorrhizal and ectomycorrhizal fungi in tropical regions as well as unicellular early-diverging groups and macrofungi in general. Given the low overlap between the endemicity of fungi and macroorganisms, but high conservation needs in both groups, detailed analyses on distribution and conservation requirements are warranted for other microorganisms and soil organisms

Effects of Anacetrapib in Patients with Atherosclerotic Vascular Disease

BACKGROUND: Patients with atherosclerotic vascular disease remain at high risk for cardiovascular events despite effective statin-based treatment of low-density lipoprotein (LDL) cholesterol levels. The inhibition of cholesteryl ester transfer protein (CETP) by anacetrapib reduces LDL cholesterol levels and increases high-density lipoprotein (HDL) cholesterol levels. However, trials of other CETP inhibitors have shown neutral or adverse effects on cardiovascular outcomes. METHODS: We conducted a randomized, double-blind, placebo-controlled trial involving 30,449 adults with atherosclerotic vascular disease who were receiving intensive atorvastatin therapy and who had a mean LDL cholesterol level of 61 mg per deciliter (1.58 mmol per liter), a mean non-HDL cholesterol level of 92 mg per deciliter (2.38 mmol per liter), and a mean HDL cholesterol level of 40 mg per deciliter (1.03 mmol per liter). The patients were assigned to receive either 100 mg of anacetrapib once daily (15,225 patients) or matching placebo (15,224 patients). The primary outcome was the first major coronary event, a composite of coronary death, myocardial infarction, or coronary revascularization. RESULTS: During the median follow-up period of 4.1 years, the primary outcome occurred in significantly fewer patients in the anacetrapib group than in the placebo group (1640 of 15,225 patients [10.8%] vs. 1803 of 15,224 patients [11.8%]; rate ratio, 0.91; 95% confidence interval, 0.85 to 0.97; P=0.004). The relative difference in risk was similar across multiple prespecified subgroups. At the trial midpoint, the mean level of HDL cholesterol was higher by 43 mg per deciliter (1.12 mmol per liter) in the anacetrapib group than in the placebo group (a relative difference of 104%), and the mean level of non-HDL cholesterol was lower by 17 mg per deciliter (0.44 mmol per liter), a relative difference of -18%. There were no significant between-group differences in the risk of death, cancer, or other serious adverse events. CONCLUSIONS: Among patients with atherosclerotic vascular disease who were receiving intensive statin therapy, the use of anacetrapib resulted in a lower incidence of major coronary events than the use of placebo. (Funded by Merck and others; Current Controlled Trials number, ISRCTN48678192 ; ClinicalTrials.gov number, NCT01252953 ; and EudraCT number, 2010-023467-18 .)

Induction of fibroblast growth factor 21 does not require activation of the hepatic X-box binding protein 1 in mice

Objective: Fibroblast growth factor 21 (FGF21), a key regulator of the metabolic response to fasting, is highly induced by endoplasmic reticulum (ER) stress. The X-box binding protein 1 (Xbp1) is one of several ER stress proteins that has been shown to directly activate the FGF21 promoter. We aimed to determine whether hepatic Xbp1 is required for induction of hepatic FGF21 in vivo. Methods: Mice bearing a hepatocyte-specific deletion of Xbp1 (Xbp1LKO) were subjected to fasting, pharmacologic ER stress, or a ketogenic diet, all potent stimuli of Fgf21 expression. Results: Hepatocyte-specific Xbp1 knockout mice demonstrated normal induction of FGF21 in response to fasting or pharmacologic ER stress and enhanced induction of FGF21 in response to a ketogenic diet. Consistent with preserved induction of FGF21, Xbp1LKO mice exhibited normal induction of FGF21 target genes and normal ketogenesis in response to fasting or a ketogenic diet. Conclusion: Hepatic Xbp1 is not required for induction of FGF21 under physiologic or pathophysiologic conditions in vivo. Keywords: Unfolded protein response, Endoplasmic reticulum stress, Fasting, Fatty acid oxidation, Ketogenic die

Endoplasmic Reticulum Stress Regulates Hepatic Bile Acid Metabolism in MiceSummary

Background & Aims: Cholestasis promotes endoplasmic reticulum (ER) stress in the liver, however, the effect of ER stress on hepatic bile acid metabolism is unknown. We aim to determine the effect of ER stress on hepatic bile acid synthesis and transport in mice. Methods: ER stress was induced pharmacologically in C57BL/6J mice and human hepatoma (HepG2) cells. The hepatic expression of genes controlling bile acid synthesis and transport was determined. To measure the activity of the primary bile acid synthetic pathway, the concentration of 7α-hydroxy-4-cholesten-3-1 was measured in plasma. Results: Induction of ER stress in mice and HepG2 cells rapidly suppressed the hepatic expression of the primary bile acid synthetic enzyme, cholesterol 7α-hydroxylase. Plasma levels of 7α-hydroxy-4-cholesten-3-1 were reduced in mice subjected to ER stress, indicating impaired bile acid synthesis. Induction of ER stress in mice and HepG2 cells increased expression of the bile salt export pump (adenosine triphosphate binding cassette [Abc]b11) and a bile salt efflux pump (Abcc3). The observed regulation of Cyp7a1, Abcb11, and Abcc3 occurred in the absence of hepatic inflammatory cytokine activation and was not dependent on activation of hepatic small heterodimer partner or intestinal fibroblast growth factor 15. Consistent with suppressed bile acid synthesis and enhanced bile acid export from hepatocytes, prolonged ER stress decreased the hepatic bile acid content in mice. Conclusions: Induction of ER stress in mice suppresses bile acid synthesis and enhances bile acid removal from hepatocytes independently of established bile acid regulatory pathways. These data show a novel function of the ER stress response in regulating bile acid metabolism. Keywords: Unfolded Protein Response, Cyp7a1, 7α-Hydroxy-4-Cholesten-3-1, Bile Acid Synthesi

Endoplasmic Reticulum Stress Activates the Hepatic Activator Protein 1 Complex via Mitogen Activated Protein Kinase-Dependent Signaling Pathways

<div><p>Background and Aims</p><p>Endoplasmic reticulum (ER) stress is induced in many forms of chronic liver disease and may promote the development of hepatocellular carcinoma. The activator protein 1 (AP-1) complex is a transcription factor that promotes hepatic carcinogenesis in response to cellular stress. The aim of this study was to determine the role of ER stress in the regulation of the hepatic AP-1 complex.</p><p>Methods</p><p>Human hepatocellular carcinoma (HepG2) cells and C57BL/6J mice were subjected to pharmacologic ER stress and the expression of AP-1-associated genes and proteins was assessed. To determine the role of MAPK signaling in ER stress-induced AP-1 activation, ER stress was induced in JNK- and ERK-inhibited HepG2 cells.</p><p>Results</p><p>Induction of ER stress promoted the activation of both Jun- and Fos-related genes and proteins of the AP-1 complex in HepG2 cells and murine liver. Inhibition of ERK phosphorylation in HepG2 cells completely prevented ER stress-induced activation of the fos-related components of AP-1 whereas activation of Jun-related components was only partially attenuated. Conversely, inhibition of JNK phosphorylation in HepG2 cells reduced ER stress-induced activation of Jun-related components but did not prevent activation of fos-related components.</p><p>Conclusions</p><p>ER stress activates the hepatic AP-1 complex via MAPK-dependent signaling pathways. ER stress-induced activation of Fos-related components is dependent primarily on ERK activation whereas ER stress-induced activation of Jun-related components is dependent primarily on JNK activation, although there is interplay between these regulatory pathways. These data implicate a novel signaling pathway by which sustained ER stress, as observed in many chronic liver diseases, may promote hepatic carcinogenesis.</p></div

Induction of ER stress in mice activates the hepatic AP-1 complex.

<p>Relative hepatic mRNA expression of G<i>rp78/Bip</i>, <i>Chop</i>, <i>cFos</i>, <i>Fra-1</i>, <i>cJun</i>, and <i>JunD</i> in mice treated with tunicamycin (Tm) at doses of A) 2 mg/kg for 6 hours, B) 2 mg/kg for 72 hours, or C) 0.5 mg/kg over 5 days. Hepatic protein expression of total and phosphorylated CFOS and CJUN in mice treated with Tm at doses of D) 2 mg/kg for 6 hours, E) 2 mg/kg for 72 hours, or F) 0.5 mg/kg over 5 days. Quantitative PCR shown as mean (n = 6) ± SD. * p<0.05 vs vehicle-treated. Representative Western blot of pooled samples, n = 2–3.</p

Effects of ERK- and JNK-inhibition on tunicamycin-induced cytotoxicity and cellular proliferation.

<p>Cytoxicity as measured by LDH release (A,C) and cellular proliferation as measured by BrdU incorporation (B,D) in HepG2 cells pretreated with MEK1 inhibitor (PD184352) or JNK inhibitor (SP600125) followed by treatment with tunicamycin or vehicle (control) for 6 hours. Mean (n = 6) ± SD. *p<0.05.</p