scholarly journals Glioblastoma epigenome profiling identifies SOX10 as a master regulator of molecular tumour subtype

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Yonghe Wu ◽  
Michael Fletcher ◽  
Zuguang Gu ◽  
Qi Wang ◽  
Barbara Costa ◽  
...  
Keyword(s):  
Regulatory Networks ◽  
Immune Cell ◽  
Sequencing Data ◽  
Master Regulator ◽  
Mesenchymal Transition ◽  
Functional Studies ◽  
Bona Fide ◽  
Oligodendrocyte Precursor

AbstractGlioblastoma frequently exhibits therapy-associated subtype transitions to mesenchymal phenotypes with adverse prognosis. Here, we perform multi-omic profiling of 60 glioblastoma primary tumours and use orthogonal analysis of chromatin and RNA-derived gene regulatory networks to identify 38 subtype master regulators, whose cell population-specific activities we further map in published single-cell RNA sequencing data. These analyses identify the oligodendrocyte precursor marker and chromatin modifier SOX10 as a master regulator in RTK I-subtype tumours. In vitro functional studies demonstrate that SOX10 loss causes a subtype switch analogous to the proneural–mesenchymal transition observed in patients at the transcriptomic, epigenetic and phenotypic levels. SOX10 repression in an in vivo syngeneic graft glioblastoma mouse model results in increased tumour invasion, immune cell infiltration and significantly reduced survival, reminiscent of progressive human glioblastoma. These results identify SOX10 as a bona fide master regulator of the RTK I subtype, with both tumour cell-intrinsic and microenvironmental effects.

scholarly journals Dickkopf-1: A Promising Target for Cancer Immunotherapy

2021 ◽  
Vol 12 ◽  
Author(s):  
Hang Yin Chu ◽  
Zihao Chen ◽  
Luyao Wang ◽  
Zong-Kang Zhang ◽  
Xinhuan Tan ◽  
...  
Keyword(s):  
Cancer Immunotherapy ◽  
Immune Cell ◽  
Immune Surveillance ◽  
Suppressor Cells ◽  
Functional Studies ◽  
Promising Target ◽  
Using Data ◽  
Dickkopf 1

Clinical studies in a range of cancers have detected elevated levels of the Wnt antagonist Dickkopf-1 (DKK1) in the serum or tumors of patients, and this was frequently associated with a poor prognosis. Our analysis of DKK1 gene profile using data from TCGA also proves the high expression of DKK1 in 14 types of cancers. Numerous preclinical studies have demonstrated the cancer-promoting effects of DKK1 in both in vitro cell models and in vivo animal models. Furthermore, DKK1 showed the ability to modulate immune cell activities as well as the immunosuppressive cancer microenvironment. Expression level of DKK1 is positively correlated with infiltrating levels of myeloid-derived suppressor cells (MDSCs) in 20 types of cancers, while negatively associated with CD8+ T cells in 4 of these 20 cancer types. Emerging experimental evidence indicates that DKK1 has been involved in T cell differentiation and induction of cancer evasion of immune surveillance by accumulating MDSCs. Consequently, DKK1 has become a promising target for cancer immunotherapy, and the mechanisms of DKK1 affecting cancers and immune cells have received great attention. This review introduces the rapidly growing body of literature revealing the cancer-promoting and immune regulatory activities of DKK1. In addition, this review also predicts that by understanding the interaction between different domains of DKK1 through computational modeling and functional studies, the underlying functional mechanism of DKK1 could be further elucidated, thus facilitating the development of anti-DKK1 drugs with more promising efficacy in cancer immunotherapy.

scholarly journals The emerging landscape of in vitro and in vivo epigenetic allelic effects

F1000Research ◽  
2017 ◽  
Vol 6 ◽  
pp. 2108 ◽  
Author(s):  
Christopher Gregg
Keyword(s):  
Cell Fate ◽  
Gene Regulatory Networks ◽  
Regulatory Networks ◽  
Disease Susceptibility ◽  
Monoallelic Expression ◽  
Functional Studies ◽  
Cell Type Specific ◽  
Gene Regulatory

Epigenetic mechanisms that cause maternally and paternally inherited alleles to be expressed differently in offspring have the potential to radically change our understanding of the mechanisms that shape disease susceptibility, phenotypic variation, cell fate, and gene expression. However, the nature and prevalence of these effects in vivo have been unclear and are debated. Here, I consider major new studies of epigenetic allelic effects in cell lines and primary cells and in vivo. The emerging picture is that these effects take on diverse forms, and this review attempts to clarify the nature of the different forms that have been uncovered for genomic imprinting and random monoallelic expression (RME). I also discuss apparent discrepancies between in vitro and in vivo studies. Importantly, multiple studies suggest that allelic effects are prevalent and can be developmental stage- and cell type-specific. I propose some possible functions and consider roles for allelic effects within the broader context of gene regulatory networks, cellular diversity, and plasticity. Overall, the field is ripe for discovery and is in need of mechanistic and functional studies.

The molecular characterization of antibody binding to a superantigen-like protein from a commensal microbe

2021 ◽  
Vol 118 (39) ◽  
pp. e2023898118
Author(s):  
Marta T. Borowska ◽  
Christoph Drees ◽  
Alexander E. Yarawsky ◽  
Mayuri Viswanathan ◽  
Sean M. Ryan ◽  
...  
Keyword(s):  
Heavy Chain ◽  
Immune Cell ◽  
Targeted Mutagenesis ◽  
Functional Studies ◽  
Immunoglobulin Binding Protein ◽  
Contact Residues ◽  
Stimulation Experiments ◽  
Immunoglobulin Binding

Microorganisms have coevolved diverse mechanisms to impair host defenses. A major one, superantigens, can result in devastating effects on the immune system. While all known superantigens induce vast immune cell proliferation and come from opportunistic pathogens, recently, proteins with similar broad specificity to antibody variable (V) domain families were identified in a commensal microbiota. These proteins, identified in the human commensal Ruminococcus gnavus, are called immunoglobulin-binding protein (Ibp) A and B and have been shown to activate B cells in vitro expressing either human VH3 or murine VH5/6/7. Here, we provide molecular and functional studies revealing the basis of this Ibp/immunoglobulin (Ig) interaction. The crystal structure and biochemical assays of a truncated IbpA construct in complex with mouse VH5 antigen-binding fragment (Fab) shows a binding of Ig heavy chain framework residues to the Ibp Domain D and the C-terminal heavy chain binding domain (HCBD). We used targeted mutagenesis of contact residues and affinity measurements and performed studies of the Fab-IbpA complex to determine the stoichiometry between Ibp and VH domains, suggesting Ibp may serve to cluster full-length IgA antibodies in vivo. Furthermore, in vitro stimulation experiments indicate that binding of the Ibp HCBD alone is sufficient to activate responsive murine B cell receptors. The presence of these proteins in a commensal microbe suggest that binding a broad repertoire of immunoglobulins, particularly in the gut/microbiome environment, may provide an important function in the maintenance of host/microbiome homeostasis contrasting with the pathogenic role of structurally homologous superantigens expressed by pathogens.

scholarly journals scATAC-Seq reveals epigenetic heterogeneity associated with an EMT-like process in male germline stem cells and its regulation by G9a

2020 ◽  
Author(s):  
Jinyue Liao ◽  
Hoi Ching Suen ◽  
Alfred Chun Shui Luk ◽  
Annie Wing Tung Lee ◽  
Judy Kin Wing Ng ◽  
...  
Keyword(s):  
Stem Cells ◽  
Single Cell ◽  
Regulatory Networks ◽  
Epithelial Mesenchymal Transition ◽  
Stem Cell Differentiation ◽  
Germline Stem Cells ◽  
Gene Markers ◽  
Mesenchymal Transition

AbstractBackgroundEpithelial-mesenchymal transition (EMT) is a phenomenon in which epithelial cells acquire mesenchymal traits. It contributes to organogenesis and tissue homeostasis, as well as stem cell differentiation. Emerging evidence indicates that heterogeneous expression of EMT gene markers presents in sub-populations of germline stem cells (GSCs). However, the functional implications of such heterogeneity are largely elusive.ResultsWe unravelled an EMT-like process in GSCs by in vitro extracellular matrix (ECM) model and single-cell genomics approaches. We found that histone methyltransferase G9a regulated an EMT-like program in GSC in vitro and contributed to neonatal germ cell migration in vivo. Through modulating ECM, we demonstrated that GSCs exist in interconvertible epithelial-like and mesenchymal-like cell states. GSCs gained higher migratory ability after transition to a mesenchymal-like cell state, which was largely mediated by the TGF-β signaling pathway. Dynamics of epigenetic regulation at the single-cell level was also found to align with the EMT-like process. Chromatin accessibility profiles generated by single-cell sequencing assay for transposase-accessible chromatin (scATAC-seq) clustered GSCs into epithelial-like and mesenchymal-like states, which were associated with differentiation status. The high-resolution data revealed regulators in the EMT-like process, including transcription factors Zeb1. We further identified putative enhancer-promoter interactions and cis-co-accessibility networks at loci such as Tgfb1, Notch1 and Lin28a. Lastly, we identified HES1 as the putative target underlying G9a’s regulation.ConclusionOur work provides the foundation for understanding the EMT-like process and a comprehensive resource for future investigation of epigenetic regulatory networks in GSCs.

Functional Studies on the C-Type Lectin Receptor CD302 Present on Dendritic Cells and Macrophages

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2198-2198
Author(s):  
Derek NJ Hart ◽  
Pablo Silveira ◽  
Tsun Ho Lo ◽  
Nirupama Verma ◽  
Ai Vu ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Dendritic Cells ◽  
Immune Cell ◽  
Blood Monocytes ◽  
Lectin Receptors ◽  
Functional Studies ◽  
Cell Functions ◽  
Equity Ownership

Abstract Introduction: C-type lectin receptors (CLR) play an important role in the immune system by recognising molecular patterns expressed by exogenous and endogenous threats. They have been shown to capture and internalise antigens and to mediate other important immune cell functions. DEC205 and CLEC9A are being actively investigated as targets for clinical therapeutic cancer vaccines. We discovered CD302 as a new CLR expressed on human dendritic cells (DC), monocytes and macrophages (J Immunol 2007;179:6052). Our initial studies suggested the molecule could play a role in cell adhesion or migration due to its co-localisation with migratory structures on macrophages. Our study set out to investigate the potential immunological function of CD302 using mouse models and to define its wider tissue expression in man. Methods: We generated CD302 knockout (KO) mice lacking exon 1 of its gene, abrogating transcription, for functional studies. We characterised the transcriptional expression of CD302 in mouse immune cells using real-time PCR. We developed monoclonal mAb to mCD302. Human studies utilized the anti-CD302 mAbs, MMRI-20 & 21 in flow cytometry and confocal microscopy studies of human immune cell populations. Results: CD302 was primarily expressed in mouse liver, lungs, lymph nodes (LN) and spleen. In spleen, macrophages, granulocytes and dendritic cells (DC) expressed CD302. Analysis of LN DC subsets revealed 2.5-fold higher CD302 mRNA expression in migratory compared to resident DC populations. Enumeration of various immune populations in lymphoid organs by flow cytometry uncovered a modest deficiency in migratory DC number and proportion within LN of CD302 KO mice compared to wild-type (WT) mice. In vitro studies showed CD302 KO and WT DC had an equivalent capacity to be activated by various stimuli, prime T cells and migrate towards the lymphoid-homing chemokines CCL19/CCL21. CD302 KO migratory DC exhibited a reduced in vivo migratory capacity to LN after FITC skin-painting. However, CD302 KO macrophages migrated similarly to WT macrophages in vivo in response to thioglycollate. In man, CD302 was present in high density in liver and peripheral blood monocytes and myeloid but not plasmacytoid DC. Current studies are aimed at clarifying its distribution on tissue DC and macrophage subsets. Anti-CD302 coated microbeads were taken up by human monocyte derived macrophages and anti-CD302 mAb was also internalized by DC. Confocal studies showed that CD302 co-localized with F-actin structures at the near basal surface such as filopodia and lamellipodia and podosomes of human macrophages and EGFP tagged CD302 expressed in COS-1 cells associated with F-actin. Conclusion: Our data suggests that CD302 may play a specialist role in DC and macrophage membrane functions. This appears to relate to its ability to associate with F-actin and may contribute to the membrane interactions required for DC to migrate towards the draining LN. Disclosures Hart: DendroCyte BioTech Pty Ltd: Equity Ownership. Clark:DendroCyte BioTech Pty Ltd: Equity Ownership.

scholarly journals Bona fide alveolar macrophage generation from long-term in vitro culture of fetal precursors provides a novel platform for gene therapy and functional studies

2021 ◽  
Author(s):  
Fengqi Li ◽  
Katarzyna Maria Okreglicka ◽  
Lea Maria Pohlmeier ◽  
Christoph Schneider ◽  
Manfred Kopf
Keyword(s):  
Gene Therapy ◽  
Alveolar Macrophage ◽  
Genetic Manipulation ◽  
Fetal Liver ◽  
Influenza Virus Infection ◽  
Gm Csf ◽  
Functional Studies ◽  
Bona Fide

Tissue-resident macrophage (MΦTR)-based immune therapies have been proposed for various diseases. However, it is still a challenge to generate a considerable amount of MΦTR that possess tissue-specific functions. Here we show that fetal liver monocytes (FLiMo) cultured with GM-CSF (also known as CSF2) rapidly differentiate into a long-lived, homogeneous alveolar macrophage (AM)-like population in vitro. CSF2-cultured FLiMo (CSF2-cFLiMo) remain the capacity to develop into bona fide AM upon transfer into Csf2ra-/- neonates and prevent development of alveolar proteinosis for at least 1 year in vivo. Compared to bone marrow-derived macrophages (BMM), CSF2-cFliMo more efficiently engraft empty AM niches in the lung and protect mice from influenza virus infection. Notably, to harness the potential of this approach for gene therapy, we restored a disrupted Csf2ra gene in FLiMo and their capacity to develop into AM in vivo, by retroviral expression of a wild-type gene. Taken together, we provide a novel platform for high-throughput genetic manipulation of AM that can be used for biological and therapeutic studies.

scholarly journals Pluripotent stem cell-derived epithelium misidentified as brain microvascular endothelium requires ETS factors to acquire vascular fate

2021 ◽  
Vol 118 (8) ◽  
pp. e2016950118 ◽  
Author(s):  
Tyler M. Lu ◽  
Sean Houghton ◽  
Tarig Magdeldin ◽  
José Gabriel Barcia Durán ◽  
Andrew P. Minotti ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Neurological Diseases ◽  
Microvascular Endothelial Cells ◽  
Brain Microvascular Endothelial Cells ◽  
Microvascular Endothelium ◽  
Functional Studies ◽  
Bona Fide ◽  
Microvascular Endothelial

Cells derived from pluripotent sources in vitro must resemble those found in vivo as closely as possible at both transcriptional and functional levels in order to be a useful tool for studying diseases and developing therapeutics. Recently, differentiation of human pluripotent stem cells (hPSCs) into brain microvascular endothelial cells (ECs) with blood–brain barrier (BBB)-like properties has been reported. These cells have since been used as a robust in vitro BBB model for drug delivery and mechanistic understanding of neurological diseases. However, the precise cellular identity of these induced brain microvascular endothelial cells (iBMECs) has not been well described. Employing a comprehensive transcriptomic metaanalysis of previously published hPSC-derived cells validated by physiological assays, we demonstrate that iBMECs lack functional attributes of ECs since they are deficient in vascular lineage genes while expressing clusters of genes related to the neuroectodermal epithelial lineage (Epi-iBMEC). Overexpression of key endothelial ETS transcription factors (ETV2, ERG, and FLI1) reprograms Epi-iBMECs into authentic endothelial cells that are congruent with bona fide endothelium at both transcriptomic as well as some functional levels. This approach could eventually be used to develop a robust human BBB model in vitro that resembles the human brain EC in vivo for functional studies and drug discovery.

scholarly journals HDAC4 promotes nasopharyngeal carcinoma progression and serves as a therapeutic target

2021 ◽  
Vol 12 (2) ◽  
Author(s):  
Chun Cheng ◽  
Jun Yang ◽  
Si-Wei Li ◽  
Guofu Huang ◽  
Chenxi Li ◽  
...  
Keyword(s):  
Nasopharyngeal Carcinoma ◽  
Tumor Growth ◽  
Therapeutic Target ◽  
Histone Deacetylases ◽  
Migration And Invasion ◽  
Poor Overall Survival ◽  
Mesenchymal Transition ◽  
E Cadherin

AbstractHistone deacetylases (HDACs) are involved in tumor progression, and some have been successfully targeted for cancer therapy. The expression of histone deacetylase 4 (HDAC4), a class IIa HDAC, was upregulated in our previous microarray screen. However, the role of HDAC4 dysregulation and mechanisms underlying tumor growth and metastasis in nasopharyngeal carcinoma (NPC) remain elusive. Here, we first confirmed that the HDAC4 levels in primary and metastatic NPC tissues were significantly increased compared with those in normal nasopharyngeal epithelial tissues and found that high HDAC4 expression predicted a poor overall survival (OS) and progression-free survival (PFS). Functionally, HDAC4 accelerated cell cycle G1/S transition and induced the epithelial-to-mesenchymal transition to promote NPC cell proliferation, migration, and invasion in vitro, as well as tumor growth and lung metastasis in vivo. Intriguingly, knockdown of N-CoR abolished the effects of HDAC4 on the invasion and migration abilities of NPC cells. Mechanistically, HDAC3/4 binds to the E-cadherin promoter to repress E-cadherin transcription. We also showed that the HDAC4 inhibitor tasquinimod suppresses tumor growth in NPC. Thus, HDAC4 may be a potential diagnostic marker and therapeutic target in patients with NPC.

scholarly journals GCH1 induces immunosuppression through metabolic reprogramming and IDO1 upregulation in triple-negative breast cancer

2021 ◽  
Vol 9 (7) ◽  
pp. e002383
Author(s):  
Jin-Li Wei ◽  
Si-Yu Wu ◽  
Yun-Song Yang ◽  
Yi Xiao ◽  
Xi Jin ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Triple Negative Breast Cancer ◽  
Triple Negative ◽  
Molecular Taxonomy ◽  
Underlying Mechanism ◽  
Metabolic Reprogramming ◽  
Sequencing Data ◽  
Aryl Hydrocarbon

PurposeRegulatory T cells (Tregs) heavily infiltrate triple-negative breast cancer (TNBC), and their accumulation is affected by the metabolic reprogramming in cancer cells. In the present study, we sought to identify cancer cell-intrinsic metabolic modulators correlating with Tregs infiltration in TNBC.Experimental designUsing the RNA-sequencing data from our institute (n=360) and the Molecular Taxonomy of Breast Cancer International Consortium TNBC cohort (n=320), we calculated the abundance of Tregs in each sample and evaluated the correlation between gene expression levels and Tregs infiltration. Then, in vivo and in vitro experiments were performed to verify the correlation and explore the underlying mechanism.ResultsWe revealed that GTP cyclohydrolase 1 (GCH1) expression was positively correlated with Tregs infiltration and high GCH1 expression was associated with reduced overall survival in TNBC. In vivo and in vitro experiments showed that GCH1 increased Tregs infiltration, decreased apoptosis, and elevated the programmed cell death-1 (PD-1)-positive fraction. Metabolomics analysis indicated that GCH1 overexpression reprogrammed tryptophan metabolism, resulting in L-5-hydroxytryptophan (5-HTP) accumulation in the cytoplasm accompanied by kynurenine accumulation and tryptophan reduction in the supernatant. Subsequently, aryl hydrocarbon receptor, activated by 5-HTP, bound to the promoter of indoleamine 2,3-dioxygenase 1 (IDO1) and thus enhanced the transcription of IDO1. Furthermore, the inhibition of GCH1 by 2,4-diamino-6-hydroxypyrimidine (DAHP) decreased IDO1 expression, attenuated tumor growth, and enhanced the tumor response to PD-1 blockade immunotherapy.ConclusionsTumor-cell-intrinsic GCH1 induced immunosuppression through metabolic reprogramming and IDO1 upregulation in TNBC. Inhibition of GCH1 by DAHP serves as a potential immunometabolic strategy in TNBC.

scholarly journals Operative ubiquitin-specific protease 22 deubiquitination confers a more invasive phenotype to cholangiocarcinoma

2021 ◽  
Vol 12 (7) ◽  
Author(s):  
Yu Tian ◽  
Bo Tang ◽  
Chengye Wang ◽  
Yan Wang ◽  
Jiakai Mao ◽  
...  
Keyword(s):  
Tumour Growth ◽  
Molecular Mechanisms ◽  
Epithelial To Mesenchymal Transition ◽  
Sirtuin 1 ◽  
Mesenchymal Transition ◽  
Specific Protease ◽  
Ubiquitin Specific Protease ◽  
Paired Tumour

AbstractOncogenic ubiquitin-specific protease 22 (USP22) is implicated in a variety of tumours; however, evidence of its role and underlying molecular mechanisms in cholangiocarcinoma (CCA) development remains unknown. We collected paired tumour and adjacent non-tumour tissues from 57 intrahepatic CCA (iCCA) patients and evaluated levels of the USP22 gene and protein by qPCR and immunohistochemistry. Both the mRNA and protein were significantly upregulated, correlated with the malignant invasion and worse OS of iCCA. In cell cultures, USP22 overexpression increased CCA cell proliferation and mobility, and induced epithelial-to-mesenchymal transition (EMT). Upon an interaction, USP22 deubiquitinated and stabilized sirtuin-1 (SIRT1), in conjunction with Akt/ERK activation. In implantation xenografts, USP22 overexpression stimulated tumour growth and metastasis to the lungs of mice. Conversely, the knockdown by USP22 shRNA attenuated the tumour growth and invasiveness in vitro and in vivo. Furthermore, SIRT1 overexpression reversed the USP22 functional deficiency, while the knockdown acetylated TGF-β-activated kinase 1 (TAK1) and Akt. Our present study defines USP22 as a poor prognostic predictor in iCCA that cooperates with SIRT1 and facilitates tumour development.

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