Glucose-induced CRL4COP1-p53 axis amplifies glycometabolism to drive tumorigenesis.

The diabetes-cancer association remains underexplained. Here, we describe a glucose-signaling axis that reinforces glucose uptake and glycolysis to consolidate the Warburg effect and overcome tumor suppression. Specifically, glucose-dependent CK2 O-GlcNAcylation impedes its phosphorylation of CSN2, a modification required for the deneddylase CSN to sequester Cullin RING ligase 4 (CRL4). Glucose, therefore, elicits CSN-CRL4 dissociation to assemble the CRL4COP1 E3 ligase, which targets p53 to derepress glycolytic enzymes. A genetic or pharmacologic disruption of the O-GlcNAc-CK2-CSN2-CRL4COP1 axis abrogates glucose-induced p53 degradation and cancer cell proliferation. Diet-induced overnutrition upregulates the CRL4COP1-p53 axis to promote PyMT-induced mammary tumorigenesis in wild type but not in mammary-gland-specific p53 knockout mice. These effects of overnutrition are reversed by P28, an investigational peptide inhibitor of COP1-p53 interaction. Thus, glycometabolism self-amplifies via a glucose-induced post-translational modification cascade culminating in CRL4COP1-mediated p53 degradation. Such mutation-independent p53 checkpoint bypass may represent the carcinogenic origin and targetable vulnerability of hyperglycemia-driven cancer.

H2A.Z facilitates licensing and activation of early replication origins.

DNA replication is a tightly regulated process that ensures the precise duplication of the genome during the cell cycle1. In eukaryotes, the licensing and activation of replication origins are regulated by both DNA sequence and chromatin features2. However, the chromatin-based regulatory mechanisms remain largely uncharacterized. Here we show that, in HeLa cells, nucleosomes containing the histone variant H2A.Z are enriched with histone H4 that is dimethylated on its lysine 20 residue (H4K20me2) and with bound origin-recognition complex (ORC). In vitro studies show that H2A.Z-containing nucleosomes bind directly to the histone lysine methyltransferase enzyme SUV420H1, promoting H4K20me2 deposition, which is in turn required for ORC1 binding. Genome-wide studies show that signals from H4K20me2, ORC1 and nascent DNA strands co-localize with H2A.Z, and that depletion of H2A.Z results in decreased H4K20me2, ORC1 and nascent-strand signals throughout the genome. H2A.Z-regulated replication origins have a higher firing efficiency and early replication timing compared with other origins. Our results suggest that the histone variant H2A.Z epigenetically regulates the licensing and activation of early replication origins and maintains replication timing through the SUV420H1-H4K20me2-ORC1 axis.

Author Correction: H2A.Z facilitates licensing and activation of early replication origins.

An Amendment to this paper has been published and can be accessed via a link at the top of the paper.

miR-22 promotes immunosuppression via activating JAK/STAT3 signaling in cutaneous squamous cell carcinoma.

Immunotherapy is the only approved systemic therapy for advanced cutaneous squamous cell carcinoma (cSCC), however, roughly 50% of patients do not respond to the therapy and resistance often occurs over time to those who initially respond. Immunosuppression could have a critical role in developing treatment resistance, thus, understanding the mechanisms of how immunosuppression is developed and regulated may be the key to improving clinical diagnosis and treatment strategies for cSCC. Here, through using a series of immunocompetent genetically engineered mouse models, we demonstrate that miR-22 promotes cSCC development by establishing regulatory T cells (Tregs)-mediated immunosuppressive tumor microenvironment (TME) in a tumor cell autonomous manner. Mechanism investigation revealed that miR-22 elicits the constitutive activation of JAK/STAT3 signaling by directly targeting its suppressor SOCS3, which augments cancer cell-derived chemokine secretion and Tregs recruitment. Epithelial-specific and global knockouts of miR-22 repress papilloma and cSCC development and progression, manifested with reduced Tregs infiltration and elevated CD8+ T cell activation. Transcriptomic analysis and functional rescue study confirmed CCL17, CCL20 and CCL22 as the main affected chemokines that mediate the chemotaxis between miR-22 highly expressing keratinocyte tumor cells and Tregs. Conversely, overexpression of SOCS3 reversed miR-22-induced Tregs recruitment toward tumor cells. Clinically, gradually increasing Tregs infiltration during cSCC progression was negatively correlated with SOCS3 abundance, supported by previously documented elevated miR-22 levels. Thus, our study uncovers a novel miR-22-SOCS3-JAK/STAT3-chemokines regulatory mechanism in defining the immunosuppressive TME and highlights the promising clinical application value of miR-22 as a common targeting molecule against JAK/STAT3 signaling and immune escape in cSCC.

Targeting miR-31 represses tumourigenesis and dedifferentiation of BRAFV600E-associated thyroid carcinoma.

BACKGROUND: BRAFV600E is the most common genetic mutation in differentiated thyroid cancer (DTC) occurring in 60% of patients and drives malignant tumour cell phenotypes including proliferation, metastasis and immune-escape. BRAFV600E-mutated papillary thyroid cancer (PTC) also displays greatly reduced expression of thyroid differentiation markers, thus tendency to radioactive iodine (RAI) refractory and poor prognosis. Therefore, understanding the molecular mechanisms and main oncogenic events underlying BRAFV600E will guide future therapy development. METHODS: Bioinformatics and clinical specimen analyses, genetic manipulation of BRAFV600E-induced PTC model, functional and mechanism exploration guided with transcriptomic screening, as well as systematic rescue experiments were applied to investigate miR-31 function within BRAFV600E-induced thyroid cancer development. Besides, nanoparticles carrying miR-31 antagomirs were testified to alleviate 131I iodide therapy on PTC models. RESULTS: We identify miR-31 as a significantly increased onco-miR in BRAFV600E-associated PTC that promotes tumour progression, metastasis and RAI refractoriness via sustained Wnt/ß-catenin signalling. Mechanistically, highly activated BRAF/MAPK pathway induces miR-31 expression via c-Jun-mediated transcriptional regulation across in vitro and transgenic mouse models. MiR-31 in turn facilitates ß-catenin stabilisation via directly repressing tumour suppressors CEBPA and DACH1, which direct the expression of multiple essential Wnt/ß-catenin pathway inhibitors. Genetic functional assays showed that thyroid-specific knockout of miR-31 inhibited BRAFV600E-induced PTC progression, and strikingly, enhanced expression of sodium-iodide symporter and other thyroid differentiation markers, thus promoted 131I uptake. Nanoparticle-mediated application of anti-miR-31 antagomirs markedly elevated radio-sensitivity of BRAFV600E-induced PTC tumours to 131I therapy, and efficiently suppressed tumour progression in the pre-clinical mouse model. CONCLUSIONS: Our findings elucidate a novel BRAF/MAPK-miR-31-Wnt/ß-catenin regulatory mechanism underlying clinically BRAFV600E-associated DTC tumourigenesis and dedifferentiation, also highlight a potential adjuvant therapeutic strategy for advanced DTC.

Modeling and insights into the structural characteristics of drug-induced autoimmune diseases.

The incidence and complexity of drug-induced autoimmune diseases (DIAD) have been on the rise in recent years, which may lead to serious or fatal consequences. Besides, many environmental and industrial chemicals can also cause DIAD. However, there are few effective approaches to estimate the DIAD potential of drugs and other chemicals currently, and the structural characteristics and mechanism of action of DIAD compounds have not been clarified. In this study, we developed the in silico models for chemical DIAD prediction and investigated the structural characteristics of DIAD chemicals based on the reliable drug data on human autoimmune diseases. We collected 148 medications which were reported can cause DIAD clinically and 450 medications that clearly do not cause DIAD. Several different machine learning algorithms and molecular fingerprints were combined to develop the in silico models. The best performed model provided the good overall accuracy on validation set with 76.26%. The model was made freely available on the website http://diad.sapredictor.cn/. To further investigate the differences in structural characteristics between DIAD chemicals and non-DIAD chemicals, several key physicochemical properties were analyzed. The results showed that AlogP, molecular polar surface area (MPSA), and the number of hydrogen bond donors (nHDon) were significantly different between the DIAD and non-DIAD structures. They may be related to the DIAD toxicity of chemicals. In addition, 14 structural alerts (SA) for DIAD toxicity were detected from predefined substructures. The SAs may be helpful to explain the mechanism of action of drug induced autoimmune disease, and can used to identify the chemicals with potential DIAD toxicity. The structural alerts have been integrated in a structural alert-based web server SApredictor (http://www.sapredictor.cn). We hope the results could provide useful information for the recognition of DIAD chemicals and the insights of structural characteristics for chemical DIAD toxicity.

HSA-MnO2-131I Combined Imaging and Treatment of Anaplastic Thyroid Carcinoma.

Purpose Compelling evidence suggests that nanoparticles (NPs) play a crucial role in cancer therapy. NPs templated with human serum albumin (HSA) has good retention in tumors. Manganese dioxide (MnO2) has been used to enhance the effect of radiotherapy. In this study, synthesized NPs using HSA-MnO2 labeled 131I to perform both imaging and therapy for anaplastic thyroid carcinoma (ATC). Method HSA-MnO2 was synthesized via HSA using a simple biomineralization method, and then labeled with Na131I by the chloramine T method. The cytotoxicity and biosafety of HSA-MnO2 were evaluated by the MTT test. The proliferation-inhibiting effect of HSA-MnO2-131I was evaluated in papillary thyroid cancer cell lines (K1, BCPAP, and KTC) and anaplastic thyroid carcinoma cell lines (Cal62, THJ16T, and ARO). For further translational application in medicine, we established a model of transplantable subcutaneously tumors in BALB\c-nu mice to assess the anti-tumor effect of HSA-MnO2-131I. The imaging effects of NPs were evaluated by MRI and SPECT/CT. Results The MTT test proved that the HSA-MnO2 had low toxicity. HSA-MnO2-131I significantly inhibited the proliferation of PTC and ATC cell lines. In addition, the results unveiled that HSA-MnO2-131I exhibited dual-modality MR/SPECT imaging for thyroid cancer visualization. In particular, HSA-MnO2-131I had an enhanced T1 signal in MR. Using SPECT/CT, we observed that HSA-MnO2-131I had good retention in tumor tissue, which was helpful for the diagnosis and treatment of tumor. In vivo assays indicated that the NPs led to a reduction in radioresistance in the tumor hypoxic microenvironment. Conclusion The nanomaterial had a simple synthesis method, good water solubility and biosafety, and good retention in tumor tissue. Hence, it could be used for SPECT/CT and MR dual mode imaging and therapy with radioiodine of tumor cells. The experimental results provided a feasible solution for combining radiotherapy and dual-model imaging by NPs for cancer diagnosis and treatment.

Targeting myeloid derived suppressor cells reverts immune suppression and sensitizes BRAF-mutant papillary thyroid cancer to MAPK inhibitors.

MAPK signaling inhibitor (MAPKi) therapies show limited efficacy for advanced thyroid cancers despite constitutive activation of the signaling correlates with disease recurrence and persistence. Understanding how BRAF pathway stimulates tumorigenesis could lead to new therapeutic targets. Here, through genetic and pathological approaches, we demonstrate that BRAFV600E promotes thyroid cancer development by increasing myeloid-derived suppressor cells (MDSCs) penetrance. This BRAFV600E-induced immune suppression involves re-activation of the developmental factor TBX3, which in turn up-regulates CXCR2 ligands in a TLR2-NFκB dependent manner, leading to MDSCs recruitment into the tumor microenvironment. CXCR2 inhibition or MDSCs repression improves MAPKi therapy effect. Clinically, high TBX3 expression correlates with BRAFV600E mutation and increased CXCR2 ligands, along with abundant MDSCs infiltration. Thus, our study uncovers a BRAFV600E-TBX3-CXCLs-MDSCs axis that guides patient stratification and could be targeted to improve the efficacy of MAPKi therapy in advanced thyroid cancer patients.

Terpene Synthases in Rice Pan-Genome and Their Responses to Chilo suppressalis Larvae Infesting.

Terpene synthase (TPS) catalyzes the synthesis of terpenes and plays an important role in plant defense. This study identified 45 OsTPS genes (32 core genes and 13 variable genes) based on the high-quality rice gene-based pan-genome. This indicates limitations in OsTPS gene studies based on a single reference genome. In the present study, through collinearity between multiple rice genomes, one OsTPS gene absent in the reference (Nipponbare) genome was found and two TPS genes in the reference genome were found to have atypical structures, which would have been ignored in single genome analysis. OsTPS genes were divided into five groups and TPS-b was lost according to the phylogenetic tree. OsTPSs in TPS-c and TPS-g were all core genes indicating these two groups were stable during domestication. In addition, through the analysis of transcriptome data, some structural variations were found to affect the expression of OsTPS genes. Through the Ka/Ks calculation of OsTPS genes, we found that different OsTPS genes were under different selection pressure during domestication; for example, OsTPS22 and OsTPS29 experienced stronger positive selection than the other OsTPS genes. After Chilo suppressalis larvae infesting, 25 differentially expressed OsTPS genes were identified, which are involved in the diterpene phytoalexins precursors biosynthesis and ent-kaurene biosynthesis pathways. Overall, the present study conducted a bioinformatics analysis of OsTPS genes using a high-quality rice pan-genome, which provided a basis for further study of OsTPS genes.

Pan-Genome Analysis Reveals the Abundant Gene Presence/Absence Variations Among Different Varieties of Melon and Their Influence on Traits.

Melon (Cucumismelo L.) is an important vegetable crop that has been subjected to domestication and improvement. Several varieties of melons with diverse phenotypes have been produced. In this study, we constructed a melon pan-genome based on 297 accessions comprising 168 Mb novel sequences and 4,325 novel genes. Based on the results, there were abundant genetic variations among different melon groups, including 364 unfavorable genes in the IMP_A vs. LDR_A group, 46 favorable genes, and 295 unfavorable genes in the IMP_M vs. LDR_M group. The distribution of 709 resistance gene analogs (RGAs) was also characterized across 297 melon lines, of which 603 were core genes. Further, 106 genes were found to be variable, 55 of which were absent in the reference melon genome. Using gene presence/absence variation (PAV)-based genome-wide association analysis (GWAS), 13 gene PAVs associated with fruit length, fruit shape, and fruit width were identified, four of which were located in pan-genome additional contigs.

miR-22 promotes stem cell traits via activating Wnt/ß-catenin signaling in cutaneous squamous cell carcinoma.

Emerging evidence suggests that the cancer stem cells (CSCs) are key culprits of cancer metastasis and drug resistance. Understanding mechanisms regulating the critical oncogenic pathways and CSCs function could reveal new diagnostic and therapeutic strategies. We now report that miR-22, a miRNA critical for hair follicle stem/progenitor cell differentiation, promotes tumor initiation, progression, and metastasis by maintaining Wnt/ß-catenin signaling and CSCs function. Mechanistically, we find that miR-22 facilitates ß-catenin stabilization through directly repressing citrullinase PAD2. Moreover, miR-22 also relieves DKK1-mediated repression of Wnt/ß-catenin signaling by targeting a FosB-DDK1 transcriptional axis. miR-22 knockout mice showed attenuated Wnt/ß-catenin activity and Lgr5+ CSCs penetrance, resulting in reduced occurrence, progression, and metastasis of chemically induced cutaneous squamous cell carcinoma (cSCC). Clinically, miR-22 is abundantly expressed in human cSCC. Its expression is even further elevated in the CSCs proportion, which negatively correlates with PAD2 and FosB expression. Inhibition of miR-22 markedly suppressed cSCC progression and increased chemotherapy sensitivity in vitro and in xenograft mice. Together, our results revealed a novel miR-22-WNT-CSCs regulatory mechanism in cSCC and highlight the important clinical application prospects of miR-22, a common target molecule for Wnt/ß-catenin signaling and CSCs, for patient stratification and therapeutic intervention.

Inhibition of BRAF Sensitizes Thyroid Carcinoma to Immunotherapy by Enhancing tsMHCII-mediated Immune Recognition.

CONTEXT: Multiple mechanisms play roles in restricting the ability of T-cells to recognize and eliminate tumor cells. OBJECTIVE: To identify immune escape mechanisms involved in papillary thyroid carcinoma (PTC) to optimize immunotherapy. SETTING AND DESIGN: iTRAQ analysis was conducted to identify proteins differentially expressed in PTC samples with or without BRAFV600E mutation. Molecular mechanisms regulating tumor cell evasion were investigated by in vitro modulations of BRAF/MAPK and related pathways. The pathological significance of identified tumor-specific major histocompatibility complex class II (tsMHCII) molecules in mediating tumor cell immune escape and targeted immune therapy was further evaluated in a transgenic mouse model of spontaneous thyroid cancer. RESULTS: Proteomic analysis showed that tsMHCII level was significantly lower in BRAFV600E-associated PTCs and negatively correlated with BRAF mutation status. Constitutive activation of BRAF decreased tsMHCII surface expression on tumor cells, which inhibited activation of CD4+ T-cells and led to immune escape. Pathway analysis indicated that the transforming growth factor (TGF)-ß1/SMAD3-mediated repression of tsMHCII, which could be reversed by BRAF inhibition (BRAFi). Targeting this pathway with a combined therapy of BRAF inhibitor PLX4032 and anti-PD-1 antibody efficiently blocked tumor growth by increasing CD4+ T-cell infiltration in a transgenic PTC mouse model. CONCLUSIONS: Our results suggest that BRAFV600E mutation in PTC impairs the expression of tsMHCII through the TGF-ß1/SMAD3 pathway to enhance immune escape. Combined treatment with PLX4032 and anti-PD-1 antibody promotes recognition and elimination of PTC by the immune system in a pre-clinical mouse model, and therefore offers an effective therapeutic strategy for patients with advanced PTC.

TBX3 promotes proliferation of papillary thyroid carcinoma cells through facilitating PRC2-mediated p57KIP2 repression.

The T-box transcription factor TBX3 has been implicated in the patterning and differentiation of a number of tissues during embryonic development, and is overexpressed in a variety of cancers; however, the precise function of TBX3 in papillary thyroid carcinoma (PTC) development remains to be determined. In the current study, we report downregulation of TBX3 in PTC cells delays the G1/S-phase transition, decreases cell growth in vitro, and inhibits tumor formation in vivo. We identified p57KIP2 as a novel downstream target that serves as the key mediator of TBX3's control over PTC cell proliferation. Reduced expression of TBX3 resulted in increased p57KIP2 level, while knockdown of p57KIP2 rescues the cell-cycle arrest phenotype. In clinical PTC specimens, the expression of TBX3 is markedly upregulated and significantly correlated with advanced tumor grade, but negatively correlated with the expression of p57KIP2. Mechanism investigation revealed that TBX3 directly binds to the CDKN1C gene promoter region, the coding gene of p57KIP2, and represses its transcription. Furthermore, recruitment of main components of the PRC2 complex as well as class I histone deacetylases, HDAC1 and HDAC2, is required for TBX3 to fulfill the transcriptional repression function. Our findings illustrate the previously unknown function and mechanism in cell proliferation regulation by the TBX3-p57KIP2 axis and provide evidence for the contribution of the PRC2 complex and HDAC1/2. Targeting of this pathway may present a novel and molecular defined strategy against PTC development.