Publications by authors named "Christopher K Glass"

247 Publications

Clonal hematopoiesis driven by DNMT3A and TET2 mutations: role in monocyte and macrophage biology and atherosclerotic cardiovascular disease.

Curr Opin Hematol 2022 Jan;29(1):1-7

Sulpizio Cardiovascular Center, Division of Cardiology, University of California San Diego, La Jolla, California, USA.

Purpose Of Review: Clonal hematopoiesis of indeterminate potential (CHIP), defined by the presence of somatic mutations in hematopoietic cells, is associated with advanced age and increased mortality due to cardiovascular disease. Gene mutations in DNMT3A and TET2 are the most frequently identified variants among patients with CHIP and provide selective advantage that spurs clonal expansion and myeloid skewing. Although DNMT3A and TET2 appear to have opposing enzymatic influence on DNA methylation, mounting data has characterized convergent inflammatory pathways, providing insights to how CHIP may mediate atherosclerotic cardiovascular disease (ASCVD).

Recent Findings: We review a multitude of studies that characterize aberrant inflammatory signaling as result of DNMT3A and TET2 deficiency in monocytes and macrophages, immune cells with prominent roles in atherosclerosis. Although specific DNA methylation signatures associated with these known epigenetic regulators have been identified, many studies have also characterized diverse modulatory functions of DNTM3A and TET2 that urge cell and context-specific experimental studies to further define how DNMT3A and TET2 may nonenzymatically activate inflammatory pathways with clinically meaningful consequences.

Summary: CHIP, common in elderly individuals, provides an opportunity understand and potentially modify age-related chronic inflammatory ASCVD risk.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1097/MOH.0000000000000688DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8639635PMC
January 2022

Exploiting dynamic enhancer landscapes to decode macrophage and microglia phenotypes in health and disease.

Mol Cell 2021 Oct 30;81(19):3888-3903. Epub 2021 Aug 30.

Department of Medicine, University of California, San Diego, San Diego, CA, USA; Department of Cellular and Molecular Medicine, University of California, San Diego, San Diego, CA, USA. Electronic address:

The development and functional potential of metazoan cells is dependent on combinatorial roles of transcriptional enhancers and promoters. Macrophages provide exceptionally powerful model systems for investigation of mechanisms underlying the activation of cell-specific enhancers that drive transitions in cell fate and cell state. Here, we review recent advances that have expanded appreciation of the diversity of macrophage phenotypes in health and disease, emphasizing studies of liver, adipose tissue, and brain macrophages as paradigms for other tissue macrophages and cell types. Studies of normal tissue-resident macrophages and macrophages associated with cirrhosis, obese adipose tissue, and neurodegenerative disease illustrate the major roles of tissue environment in remodeling enhancer landscapes to specify the development and functions of distinct macrophage phenotypes. We discuss the utility of quantitative analysis of environment-dependent changes in enhancer activity states as an approach to discovery of regulatory transcription factors and upstream signaling pathways.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.molcel.2021.08.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8500948PMC
October 2021

A Cdk4/6-dependent phosphorylation gradient regulates the early to late G1 phase transition.

Sci Rep 2021 07 19;11(1):14736. Epub 2021 Jul 19.

Department of Cellular and Molecular Medicine, University of California San Diego, La Jolla, CA, USA.

During early G1 phase, Rb is exclusively mono-phosphorylated by cyclin D:Cdk4/6, generating 14 different isoforms with specific binding patterns to E2Fs and other cellular protein targets. While mono-phosphorylated Rb is dispensable for early G1 phase progression, interfering with cyclin D:Cdk4/6 kinase activity prevents G1 phase progression, questioning the role of cyclin D:Cdk4/6 in Rb inactivation. To dissect the molecular functions of cyclin D:Cdk4/6 during cell cycle entry, we generated a single cell reporter for Cdk2 activation, RB inactivation and cell cycle entry by CRISPR/Cas9 tagging endogenous p27 with mCherry. Through single cell tracing of Cdk4i cells, we identified a time-sensitive early G1 phase specific Cdk4/6-dependent phosphorylation gradient that regulates cell cycle entry timing and resides between serum-sensing and cyclin E:Cdk2 activation. To reveal the substrate identity of the Cdk4/6 phosphorylation gradient, we performed whole proteomic and phospho-proteomic mass spectrometry, and identified 147 proteins and 82 phospho-peptides that significantly changed due to Cdk4 inhibition in early G1 phase. In summary, we identified novel (non-Rb) cyclin D:Cdk4/6 substrates that connects early G1 phase functions with cyclin E:Cdk2 activation and Rb inactivation by hyper-phosphorylation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-021-94200-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8290049PMC
July 2021

A Chinese hamster transcription start site atlas that enables targeted editing of CHO cells.

NAR Genom Bioinform 2021 Sep 13;3(3):lqab061. Epub 2021 Jul 13.

Novo Nordisk Foundation Center for Biosustainability at UC San Diego 92093, USA.

Chinese hamster ovary (CHO) cells are widely used for producing biopharmaceuticals, and engineering gene expression in CHO is key to improving drug quality and affordability. However, engineering gene expression or activating silent genes requires accurate annotation of the underlying regulatory elements and transcription start sites (TSSs). Unfortunately, most TSSs in the published Chinese hamster genome sequence were computationally predicted and are frequently inaccurate. Here, we use nascent transcription start site sequencing methods to revise TSS annotations for 15 308 Chinese hamster genes and 3034 non-coding RNAs based on experimental data from CHO-K1 cells and 10 hamster tissues. We further capture tens of thousands of putative transcribed enhancer regions with this method. Our revised TSSs improves upon the RefSeq annotation by revealing core sequence features of gene regulation such as the TATA box and the Initiator and, as exemplified by targeting the glycosyltransferase gene , facilitate activating silent genes by CRISPRa. Together, we envision our revised annotation and data will provide a rich resource for the CHO community, improve genome engineering efforts and aid comparative and evolutionary studies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/nargab/lqab061DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8276764PMC
September 2021

Diet-regulated production of PDGFcc by macrophages controls energy storage.

Science 2021 07;373(6550)

Immunology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.

The mechanisms by which macrophages regulate energy storage remain poorly understood. We identify in a genetic screen a platelet-derived growth factor (PDGF)/vascular endothelial growth factor (VEGF)-family ortholog, Pvf3, that is produced by macrophages and is required for lipid storage in fat-body cells of larvae. Genetic and pharmacological experiments indicate that the mouse Pvf3 ortholog PDGFcc, produced by adipose tissue-resident macrophages, controls lipid storage in adipocytes in a leptin receptor- and C-C chemokine receptor type 2-independent manner. PDGFcc production is regulated by diet and acts in a paracrine manner to control lipid storage in adipose tissues of newborn and adult mice. At the organismal level upon PDGFcc blockade, excess lipids are redirected toward thermogenesis in brown fat. These data identify a macrophage-dependent mechanism, conducive to the design of pharmacological interventions, that controls energy storage in metazoans.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/science.abe9383DOI Listing
July 2021

Mechanisms underlying divergent responses of genetically distinct macrophages to IL-4.

Sci Adv 2021 Jun 16;7(25). Epub 2021 Jun 16.

Department of Cellular and Molecular Medicine, School of Medicine, University of California, San Diego, La Jolla, CA 92093, USA.

Mechanisms by which noncoding genetic variation influences gene expression remain only partially understood but are considered to be major determinants of phenotypic diversity and disease risk. Here, we evaluated effects of >50 million single-nucleotide polymorphisms and short insertions/deletions provided by five inbred strains of mice on the responses of macrophages to interleukin-4 (IL-4), a cytokine that plays pleiotropic roles in immunity and tissue homeostasis. Of >600 genes induced >2-fold by IL-4 across the five strains, only 26 genes reached this threshold in all strains. By applying deep learning and motif mutation analyses to epigenetic data for macrophages from each strain, we identified the dominant combinations of lineage-determining and signal-dependent transcription factors driving IL-4 enhancer activation. These studies further revealed mechanisms by which noncoding genetic variation influences absolute levels of enhancer activity and their dynamic responses to IL-4, thereby contributing to strain-differential patterns of gene expression and phenotypic diversity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/sciadv.abf9808DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8208725PMC
June 2021

Age-dependent instability of mature neuronal fate in induced neurons from Alzheimer's patients.

Cell Stem Cell 2021 09 27;28(9):1533-1548.e6. Epub 2021 Apr 27.

Laboratory of Genetics, The Salk Institute for Biological Studies, La Jolla, CA, USA. Electronic address:

Sporadic Alzheimer's disease (AD) exclusively affects elderly people. Using direct conversion of AD patient fibroblasts into induced neurons (iNs), we generated an age-equivalent neuronal model. AD patient-derived iNs exhibit strong neuronal transcriptome signatures characterized by downregulation of mature neuronal properties and upregulation of immature and progenitor-like signaling pathways. Mapping iNs to longitudinal neuronal differentiation trajectory data demonstrated that AD iNs reflect a hypo-mature neuronal identity characterized by markers of stress, cell cycle, and de-differentiation. Epigenetic landscape profiling revealed an underlying aberrant neuronal state that shares similarities with malignant transformation and age-dependent epigenetic erosion. To probe for the involvement of aging, we generated rejuvenated iPSC-derived neurons that showed no significant disease-related transcriptome signatures, a feature that is consistent with epigenetic clock and brain ontogenesis mapping, which indicate that fibroblast-derived iNs more closely reflect old adult brain stages. Our findings identify AD-related neuronal changes as age-dependent cellular programs that impair neuronal identity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.stem.2021.04.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8423435PMC
September 2021

Genome-wide screens uncover KDM2B as a modifier of protein binding to heparan sulfate.

Nat Chem Biol 2021 06 12;17(6):684-692. Epub 2021 Apr 12.

Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA, USA.

Heparan sulfate (HS) proteoglycans bind extracellular proteins that participate in cell signaling, attachment and endocytosis. These interactions depend on the arrangement of sulfated sugars in the HS chains generated by well-characterized biosynthetic enzymes; however, the regulation of these enzymes is largely unknown. We conducted genome-wide CRISPR-Cas9 screens with a small-molecule ligand that binds to HS. Screening of A375 melanoma cells uncovered additional genes and pathways impacting HS formation. The top hit was the epigenetic factor KDM2B, a histone demethylase. KDM2B inactivation suppressed multiple HS sulfotransferases and upregulated the sulfatase SULF1. These changes differentially affected the interaction of HS-binding proteins. KDM2B-deficient cells displayed decreased growth rates, which was rescued by SULF1 inactivation. In addition, KDM2B deficiency altered the expression of many extracellular matrix genes. Thus, KDM2B controls proliferation of A375 cells through the regulation of HS structure and serves as a master regulator of the extracellular matrix.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41589-021-00776-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8159865PMC
June 2021

PI3Kγ inhibition suppresses microglia/TAM accumulation in glioblastoma microenvironment to promote exceptional temozolomide response.

Proc Natl Acad Sci U S A 2021 04;118(16)

Department of Neurosurgery, University of Minnesota, Minneapolis, MN 55455;

Precision medicine in oncology leverages clinical observations of exceptional response. Toward an understanding of the molecular features that define this response, we applied an integrated, multiplatform analysis of RNA profiles derived from clinically annotated glioblastoma samples. This analysis suggested that specimens from exceptional responders are characterized by decreased accumulation of microglia/macrophages in the glioblastoma microenvironment. Glioblastoma-associated microglia/macrophages secreted interleukin 11 (IL11) to activate STAT3-MYC signaling in glioblastoma cells. This signaling induced stem cell states that confer enhanced tumorigenicity and resistance to the standard-of-care chemotherapy, temozolomide (TMZ). Targeting a myeloid cell restricted an isoform of phosphoinositide-3-kinase, phosphoinositide-3-kinase gamma isoform (PI3Kγ), by pharmacologic inhibition or genetic inactivation disrupted this signaling axis by reducing microglia/macrophage-associated IL11 secretion in the tumor microenvironment. Mirroring the clinical outcomes of exceptional responders, PI3Kγ inhibition synergistically enhanced the anti-neoplastic effects of TMZ in orthotopic murine glioblastoma models. Moreover, inhibition or genetic inactivation of PI3Kγ in murine glioblastoma models recapitulated expression profiles observed in clinical specimens isolated from exceptional responders. Our results suggest key contributions from tumor-associated microglia/macrophages in exceptional responses and highlight the translational potential for PI3Kγ inhibition as a glioblastoma therapy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.2009290118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8072253PMC
April 2021

Incorporation of a nucleoside analog maps genome repair sites in postmitotic human neurons.

Science 2021 04;372(6537):91-94

Laboratory of Genetics, The Salk Institute for Biological Studies, 10010 North Torrey Pines Road, La Jolla, CA 92037-1002, USA.

Neurons are the longest-lived cells in our bodies and lack DNA replication, which makes them reliant on a limited repertoire of DNA repair mechanisms to maintain genome fidelity. These repair mechanisms decline with age, but we have limited knowledge of how genome instability emerges and what strategies neurons and other long-lived cells may have evolved to protect their genomes over the human life span. A targeted sequencing approach in human embryonic stem cell-induced neurons shows that, in neurons, DNA repair is enriched at well-defined hotspots that protect essential genes. These hotspots are enriched with histone H2A isoforms and RNA binding proteins and are associated with evolutionarily conserved elements of the human genome. These findings provide a basis for understanding genome integrity as it relates to aging and disease in the nervous system.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/science.abb9032DOI Listing
April 2021

Purification of mouse hepatic non-parenchymal cells or nuclei for use in ChIP-seq and other next-generation sequencing approaches.

STAR Protoc 2021 Mar 10;2(1):100363. Epub 2021 Mar 10.

Department of Medicine, University of California, San Diego, La Jolla, CA 92093, USA.

Significant advancements in understanding disease mechanisms can occur through combined analysis of next-generation sequencing datasets generated using purified cell populations. Here, we detail our optimized protocol for purification of mouse hepatic macrophages (or other liver non-parenchymal populations) suitable for use in various next-generation sequencing protocols. An alternative framework is described for sorting pre-fixed hepatic nuclei populations. This strategy has the advantage of rapidly preserving the nuclei and can facilitate success with ChIP-seq for more challenging molecules. For complete details on the use and execution of these protocols, please refer to Muse et al. (2018), Sakai et al. (2019), and Seidman et al. (2020).
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.xpro.2021.100363DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7960533PMC
March 2021

Epigenetic Regulation of Kupffer Cell Function in Health and Disease.

Front Immunol 2020 26;11:609618. Epub 2021 Jan 26.

Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA, United States.

Kupffer cells, the resident macrophages of the liver, comprise the largest pool of tissue macrophages in the body. Within the liver sinusoids Kupffer cells perform functions common across many tissue macrophages including response to tissue damage and antigen presentation. They also engage in specialized activities including iron scavenging and the uptake of opsonized particles from the portal blood. Here, we review recent studies of the epigenetic pathways that establish Kupffer cell identity and function. We describe a model by which liver-environment specific signals induce lineage determining transcription factors necessary for differentiation of Kupffer cells from bone-marrow derived monocytes. We conclude by discussing how these lineage determining transcription factors (LDTFs) drive Kupffer cell behavior during both homeostasis and disease, with particular focus on the relevance of Kupffer cell LDTF pathways in the setting of non-alcoholic fatty liver disease and non-alcoholic steatohepatitis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fimmu.2020.609618DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7870864PMC
June 2021

Heterogeneity of HSCs in a Mouse Model of NASH.

Hepatology 2021 Aug 10;74(2):667-685. Epub 2021 Aug 10.

Department of Medicine, University of California, San Diego, La Jolla, CA.

Background And Aims: In clinical and experimental NASH, the origin of the scar-forming myofibroblast is the HSC. We used foz/foz mice on a Western diet to characterize in detail the phenotypic changes of HSCs in a NASH model.

Approach And Results: We examined the single-cell expression profiles (scRNA sequencing) of HSCs purified from the normal livers of foz/foz mice on a chow diet, in NASH with fibrosis of foz/foz mice on a Western diet, and in livers during regression of NASH after switching back to a chow diet. Selected genes were analyzed using immunohistochemistry, quantitative real-time PCR, and short hairpin RNA knockdown in primary mouse HSCs. Our analysis of the normal liver identified two distinct clusters of quiescent HSCs that correspond to their acinar position of either pericentral vein or periportal vein. The NASH livers had four distinct HSC clusters, including one representing the classic fibrogenic myofibroblast. The three other HSC clusters consisted of a proliferating cluster, an intermediate activated cluster, and an immune and inflammatory cluster. The livers with NASH regression had one cluster of inactivated HSCs, which was similar to, but distinct from, the quiescent HSCs.

Conclusions: Analysis of single-cell RNA sequencing in combination with an interrogation of previous studies revealed an unanticipated heterogeneity of HSC phenotypes under normal and injured states.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/hep.31743DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8346581PMC
August 2021

FOXO1 constrains activation and regulates senescence in CD8 T cells.

Cell Rep 2021 01;34(4):108674

Division of Biological Sciences, Molecular Biology Section, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0377, USA; Department of Cellular and Molecular Medicine, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0377, USA. Electronic address:

Naive and memory T cells are maintained in a quiescent state, yet capable of rapid response and differentiation to antigen challenge via molecular mechanisms that are not fully understood. In naive cells, the deletion of Foxo1 following thymic development results in the increased expression of multiple AP-1 family members, rendering T cells less able to respond to antigenic challenge. Similarly, in the absence of FOXO1, post-infection memory T cells exhibit the characteristics of extended activation and senescence. Age-based analysis of human peripheral T cells reveals that levels of FOXO1 and its downstream target, TCF7, are inversely related to host age, whereas the opposite is found for AP-1 factors. These characteristics of aging also correlate with the formation of T cells manifesting features of cellular senescence. Our work illustrates a role for FOXO1 in the active maintenance of stem-like properties in T cells at the timescales of acute infection and organismal life span.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.celrep.2020.108674DOI Listing
January 2021

Nuclei isolation of multiple brain cell types for omics interrogation.

Nat Protoc 2021 03 25;16(3):1629-1646. Epub 2021 Jan 25.

Department of Cellular and Molecular Medicine, University of California San Diego, La Jolla, CA, USA.

We present a nuclei isolation protocol for genomic and epigenomic interrogation of multiple cell type populations in the human and rodent brain. The nuclei isolation protocol allows cell type-specific profiling of neurons, microglia, oligodendrocytes, and astrocytes, being compatible with fresh and frozen samples obtained from either resected or postmortem brain tissue. This 2-day procedure consists of tissue homogenization with fixation, nuclei extraction, and antibody staining followed by fluorescence-activated nuclei sorting (FANS) and does not require specialized skillsets. Cell type-specific nuclei populations can be used for downstream omic-scale sequencing applications with an emphasis on epigenomic interrogation such as histone modifications, transcription factor binding, chromatin accessibility, and chromosome architecture. The nuclei isolation protocol enables translational examination of archived healthy and diseased brain specimens through utilization of existing medical biorepositories.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41596-020-00472-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7969463PMC
March 2021

Monocyte Regulation in Homeostasis and Malignancy.

Trends Immunol 2021 02 11;42(2):104-119. Epub 2021 Jan 11.

Medical Research Council (MRC) Centre for Reproductive Health, Queen's Medical Research Institute, University of Edinburgh, Edinburgh EH16 4TJ, UK. Electronic address:

Monocytes are progenitors to macrophages and a subclass of dendritic cells (monocyte-derived dendritic cells, MoDCs), but they also act as circulating sensors that respond to environmental changes and disease. Technological advances have defined the production of classical monocytes in the bone marrow through the identification of lineage-determining transcription factors (LDTFs) and have proposed alternative routes of differentiation. Monocytes released into the circulation can be recruited to tissues by specific chemoattractants where they respond to sequential niche-specific signals that determine their differentiation into terminal effector cells. New aspects of monocyte biology in the circulation are being revealed, exemplified by the influence of cancer on the systemic alteration of monocyte subset abundance and transcriptional profiles. These changes can act to enhance the metastatic spread of primary cancers and may offer therapeutic opportunities.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.it.2020.12.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7877795PMC
February 2021

Microanatomy of the Human Atherosclerotic Plaque by Single-Cell Transcriptomics.

Circ Res 2020 11 28;127(11):1437-1455. Epub 2020 Sep 28.

Laboratory of Clinical Chemistry and Haematology, University Medical Center, Heidelberglaan 100, Utrecht, the Netherlands (L.S., A.B., F.W.A., S.W.v.d.L., M.M., G.P.).

Rationale: Atherosclerotic lesions are known for their cellular heterogeneity, yet the molecular complexity within the cells of human plaques has not been fully assessed.

Objective: Using single-cell transcriptomics and chromatin accessibility, we gained a better understanding of the pathophysiology underlying human atherosclerosis.

Methods And Results: We performed single-cell RNA and single-cell ATAC sequencing on human carotid atherosclerotic plaques to define the cells at play and determine their transcriptomic and epigenomic characteristics. We identified 14 distinct cell populations including endothelial cells, smooth muscle cells, mast cells, B cells, myeloid cells, and T cells and identified multiple cellular activation states and suggested cellular interconversions. Within the endothelial cell population, we defined subsets with angiogenic capacity plus clear signs of endothelial to mesenchymal transition. CD4 and CD8 T cells showed activation-based subclasses, each with a gradual decline from a cytotoxic to a more quiescent phenotype. Myeloid cells included 2 populations of proinflammatory macrophages showing IL (interleukin) 1B or TNF (tumor necrosis factor) expression as well as a foam cell-like population expressing TREM2 (triggering receptor expressed on myeloid cells 2) and displaying a fibrosis-promoting phenotype. ATACseq data identified specific transcription factors associated with the myeloid subpopulation and T cell cytokine profiles underlying mutual activation between both cell types. Finally, cardiovascular disease susceptibility genes identified using public genome-wide association studies data were particularly enriched in lesional macrophages, endothelial, and smooth muscle cells.

Conclusions: This study provides a transcriptome-based cellular landscape of human atherosclerotic plaques and highlights cellular plasticity and intercellular communication at the site of disease. This detailed definition of cell communities at play in atherosclerosis will facilitate cell-based mapping of novel interventional targets with direct functional relevance for the treatment of human disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1161/CIRCRESAHA.120.316770DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7641189PMC
November 2020

MAGGIE: leveraging genetic variation to identify DNA sequence motifs mediating transcription factor binding and function.

Bioinformatics 2020 07;36(Suppl_1):i84-i92

Department of Cellular and Molecular Medicine, School of Medicine.

Motivation: Genetic variation in regulatory elements can alter transcription factor (TF) binding by mutating a TF binding motif, which in turn may affect the activity of the regulatory elements. However, it is unclear which motifs are prone to impact transcriptional regulation if mutated. Current motif analysis tools either prioritize TFs based on motif enrichment without linking to a function or are limited in their applications due to the assumption of linearity between motifs and their functional effects.

Results: We present MAGGIE (Motif Alteration Genome-wide to Globally Investigate Elements), a novel method for identifying motifs mediating TF binding and function. By leveraging measurements from diverse genotypes, MAGGIE uses a statistical approach to link mutations of a motif to changes of an epigenomic feature without assuming a linear relationship. We benchmark MAGGIE across various applications using both simulated and biological datasets and demonstrate its improvement in sensitivity and specificity compared with the state-of-the-art motif analysis approaches. We use MAGGIE to gain novel insights into the divergent functions of distinct NF-κB factors in pro-inflammatory macrophages, revealing the association of p65-p50 co-binding with transcriptional activation and the association of p50 binding lacking p65 with transcriptional repression.

Availability And Implementation: The Python package for MAGGIE is freely available at https://github.com/zeyang-shen/maggie. The accession number for the NF-κB ChIP-seq data generated for this study is Gene Expression Omnibus: GSE144070.

Supplementary Information: Supplementary data are available at Bioinformatics online.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/bioinformatics/btaa476DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7355228PMC
July 2020

Enhancer reprogramming driven by high-order assemblies of transcription factors promotes phenotypic plasticity and breast cancer endocrine resistance.

Nat Cell Biol 2020 06 18;22(6):701-715. Epub 2020 May 18.

Department of Molecular Medicine, Mays Cancer Center, University of Texas Health Science Center at San Antonio, San Antonio, TX, USA.

Acquired therapy resistance is a major problem for anticancer treatment, yet the underlying molecular mechanisms remain unclear. Using an established breast cancer cellular model, we show that endocrine resistance is associated with enhanced phenotypic plasticity, indicated by a general downregulation of luminal/epithelial differentiation markers and upregulation of basal/mesenchymal invasive markers. Consistently, similar gene expression changes are found in clinical breast tumours and patient-derived xenograft samples that are resistant to endocrine therapies. Mechanistically, the differential interactions between oestrogen receptor α and other oncogenic transcription factors, exemplified by GATA3 and AP1, drive global enhancer gain/loss reprogramming, profoundly altering breast cancer transcriptional programs. Our functional studies in multiple culture and xenograft models reveal a coordinated role of GATA3 and AP1 in re-organizing enhancer landscapes and regulating cancer phenotypes. Collectively, our study suggests that differential high-order assemblies of transcription factors on enhancers trigger genome-wide enhancer reprogramming, resulting in transcriptional transitions that promote tumour phenotypic plasticity and therapy resistance.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41556-020-0514-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7737911PMC
June 2020

Niche-Specific Reprogramming of Epigenetic Landscapes Drives Myeloid Cell Diversity in Nonalcoholic Steatohepatitis.

Immunity 2020 06 1;52(6):1057-1074.e7. Epub 2020 May 1.

Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA, USA; Department of Medicine, University of California, San Diego, La Jolla, CA, USA. Electronic address:

Tissue-resident and recruited macrophages contribute to both host defense and pathology. Multiple macrophage phenotypes are represented in diseased tissues, but we lack deep understanding of mechanisms controlling diversification. Here, we investigate origins and epigenetic trajectories of hepatic macrophages during diet-induced non-alcoholic steatohepatitis (NASH). The NASH diet induced significant changes in Kupffer cell enhancers and gene expression, resulting in partial loss of Kupffer cell identity, induction of Trem2 and Cd9 expression, and cell death. Kupffer cell loss was compensated by gain of adjacent monocyte-derived macrophages that exhibited convergent epigenomes, transcriptomes, and functions. NASH-induced changes in Kupffer cell enhancers were driven by AP-1 and EGR that reprogrammed LXR functions required for Kupffer cell identity and survival to instead drive a scar-associated macrophage phenotype. These findings reveal mechanisms by which disease-associated environmental signals instruct resident and recruited macrophages to acquire distinct gene expression programs and corresponding functions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.immuni.2020.04.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7305990PMC
June 2020

ZNF263 is a transcriptional regulator of heparin and heparan sulfate biosynthesis.

Proc Natl Acad Sci U S A 2020 04 10;117(17):9311-9317. Epub 2020 Apr 10.

Department of Cellular and Molecular Medicine, University of California San Diego, La Jolla, CA 92093-0687;

Heparin is the most widely prescribed biopharmaceutical in production globally. Its potent anticoagulant activity and safety makes it the drug of choice for preventing deep vein thrombosis and pulmonary embolism. In 2008, adulterated material was introduced into the heparin supply chain, resulting in several hundred deaths and demonstrating the need for alternate sources of heparin. Heparin is a fractionated form of heparan sulfate derived from animal sources, predominantly from connective tissue mast cells in pig mucosa. While the enzymes involved in heparin biosynthesis are identical to those for heparan sulfate, the factors regulating these enzymes are not understood. Examination of the promoter regions of all genes involved in heparin/heparan sulfate assembly uncovered a transcription factor-binding motif for , a C2H2 zinc finger protein. CRISPR-mediated targeting and siRNA knockdown of in mammalian cell lines and human primary cells led to dramatically increased expression levels of , a key enzyme involved in imparting anticoagulant activity to heparin, and another glucosaminyl 3--sulfotransferase expressed in cells. Enhanced 3--sulfation increased binding to antithrombin, which enhanced Factor Xa inhibition, and binding of neuropilin-1. Analysis of transcriptomics data showed distinctively low expression of in mast cells compared with other (non-heparin-producing) immune cells. These findings demonstrate a novel regulatory factor in heparan sulfate modification that could further advance the possibility of bioengineering anticoagulant heparin in cultured cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1920880117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7196839PMC
April 2020

Author Correction: A monocyte gene expression signature in the early clinical course of Parkinson's disease.

Sci Rep 2020 Apr 7;10(1):6261. Epub 2020 Apr 7.

Department of Molecular Neurology, University Hospital Erlangen, Friedrich-Alexander Universität (FAU) Erlangen-Nürnberg, 91054, Erlangen, Germany.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-020-62928-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7136204PMC
April 2020

Chronic Peripheral Inflammation Causes a Region-Specific Myeloid Response in the Central Nervous System.

Cell Rep 2020 03;30(12):4082-4095.e6

Department of Molecular Neurology, Friedrich Alexander University Erlangen-Nürnberg, University Hospital Erlangen, 91054 Erlangen, Germany; Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA. Electronic address:

Systemic immune dysregulation contributes to the development of neuropsychiatric and neurodegenerative diseases. The precise effect of chronic peripheral immune stimulation on myeloid cells across anatomical brain regions is unclear. Here, we demonstrate brain-region-specific differences in myeloid responses induced by chronic peripheral inflammation. This shift in the myeloid compartment is associated with the appearance of an inflammatory myeloid subpopulation in the cortex, striatum, and thalamus accompanied by regional transcriptomic fingerprints that include induction of chemokines, complement factors, and endothelial adhesion molecules. In contrast, myeloid immune responses within the hippocampus and cerebellum are subtle or absent. Treatment with the anti-tumor necrosis factor α (anti-TNF-α) antibody infliximab ablates the region-specific inflammatory response. A region-specific myeloid cell response to chronic peripheral inflammation is observed in postmortem brains from individuals with rheumatoid arthritis. Our data suggest that chronic peripheral inflammation has heterogeneous effects on the brain, as evidenced by the spectrum of myeloid cell responses observed across brain regions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.celrep.2020.02.109DOI Listing
March 2020

Coordinated demethylation of H3K9 and H3K27 is required for rapid inflammatory responses of endothelial cells.

EMBO J 2020 04 3;39(7):e103949. Epub 2020 Mar 3.

Isotope Science Center, The University of Tokyo, Tokyo, Japan.

Histone H3 lysine-9 di-methylation (H3K9me2) and lysine-27 tri-methylation (H3K27me3) are linked to repression of gene expression, but the functions of repressive histone methylation dynamics during inflammatory responses remain enigmatic. Here, we report that lysine demethylases 7A (KDM7A) and 6A (UTX) play crucial roles in tumor necrosis factor (TNF)-α signaling in endothelial cells (ECs), where they are regulated by a novel TNF-α-responsive microRNA, miR-3679-5p. TNF-α rapidly induces co-occupancy of KDM7A and UTX at nuclear factor kappa-B (NF-κB)-associated elements in human ECs. KDM7A and UTX demethylate H3K9me2 and H3K27me3, respectively, and are both required for activation of NF-κB-dependent inflammatory genes. Chromosome conformation capture-based methods furthermore uncover increased interactions between TNF-α-induced super enhancers at NF-κB-relevant loci, coinciding with KDM7A and UTX recruitments. Simultaneous pharmacological inhibition of KDM7A and UTX significantly reduces leukocyte adhesion in mice, establishing the biological and potential translational relevance of this mechanism. Collectively, these findings suggest that rapid erasure of repressive histone marks by KDM7A and UTX is essential for NF-κB-dependent regulation of genes that control inflammatory responses of ECs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.15252/embj.2019103949DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7110136PMC
April 2020

Blockade of IL-17 signaling reverses alcohol-induced liver injury and excessive alcohol drinking in mice.

JCI Insight 2020 02 13;5(3). Epub 2020 Feb 13.

Department of Surgery, and.

Chronic alcohol abuse has a detrimental effect on the brain and liver. There is no effective treatment for these patients, and the mechanism underlying alcohol addiction and consequent alcohol-induced damage of the liver/brain axis remains unresolved. We compared experimental models of alcoholic liver disease (ALD) and alcohol dependence in mice and demonstrated that genetic ablation of IL-17 receptor A (IL-17ra-/-) or pharmacological blockade of IL-17 signaling effectively suppressed the increased voluntary alcohol drinking in alcohol-dependent mice and blocked alcohol-induced hepatocellular and neurological damage. The level of circulating IL-17A positively correlated with the alcohol use in excessive drinkers and was further increased in patients with ALD as compared with healthy individuals. Our data suggest that IL-17A is a common mediator of excessive alcohol consumption and alcohol-induced liver/brain injury, and targeting IL-17A may provide a novel strategy for treatment of alcohol-induced pathology.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1172/jci.insight.131277DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7098802PMC
February 2020

Macrophage Syk-PI3Kγ Inhibits Antitumor Immunity: SRX3207, a Novel Dual Syk-PI3K Inhibitory Chemotype Relieves Tumor Immunosuppression.

Mol Cancer Ther 2020 03 23;19(3):755-764. Epub 2020 Jan 23.

UCSD Department of Pediatrics, University of California, San Diego, San Diego, California.

Macrophages (MΦ) play a critical role in tumor growth, immunosuppression, and inhibition of adaptive immune responses in cancer. Hence, targeting signaling pathways in MΦs that promote tumor immunosuppression will provide therapeutic benefit. PI3Kγ has been recently established by our group and others as a novel immuno-oncology target. Herein, we report that an MΦ Syk-PI3K axis drives polarization of immunosuppressive MΦs that establish an immunosuppressive tumor microenvironment in syngeneic tumor models. Genetic or pharmacologic inhibition of Syk and/or PI3Kγ in MΦs promotes a proinflammatory MΦ phenotype, restores CD8 T-cell activity, destabilizes HIF under hypoxia, and stimulates an antitumor immune response. Assay for transposase-accessible Chromatin using Sequencing (ATAC-seq) analyses on the bone marrow-derived macrophages (BMDM) show that inhibition of Syk kinase promotes activation and binding of NF-κB motif in Syk BMDMs, thus stimulating immunostimulatory transcriptional programming in MΦs to suppress tumor growth. Finally, we have developed the "first-in-class" dual Syk/PI3K inhibitor, SRX3207, for the combinatorial inhibition of Syk and PI3K in one small molecule. This chemotype demonstrates efficacy in multiple tumor models and represents a novel combinatorial approach to activate antitumor immunity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/1535-7163.MCT-19-0947DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7450492PMC
March 2020

Transcriptomic and epigenetic mechanisms underlying myeloid diversity in the lung.

Nat Immunol 2020 02 20;21(2):221-231. Epub 2020 Jan 20.

Department of Cellular and Molecular Medicine, University of California San Diego, La Jolla, CA, USA.

The lung is inhabited by resident alveolar and interstitial macrophages as well as monocytic cells that survey lung tissues. Each cell type plays distinct functional roles under homeostatic and inflammatory conditions, but mechanisms establishing their molecular identities and functional potential remain poorly understood. In the present study, systematic evaluation of transcriptomes and open chromatin of alveolar macrophages (AMs), interstitial macrophages (IMs) and lung monocytes from two mouse strains enabled inference of common and cell-specific transcriptional regulators. We provide evidence that these factors drive selection of regulatory landscapes that specify distinct phenotypes of AMs and IMs and entrain qualitatively different responses to toll-like receptor 4 signaling in vivo. These studies reveal a striking divergence in a fundamental innate immune response pathway in AMs and establish a framework for further understanding macrophage diversity in the lung.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41590-019-0582-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7667722PMC
February 2020

deficient microglia exhibit a premature aging transcriptome.

Life Sci Alliance 2019 12 2;2(6). Epub 2019 Dec 2.

Acute Neurology, Biogen, Cambridge, MA, USA

CX3CR1, one of the highest expressed genes in microglia in mice and humans, is implicated in numerous microglial functions. However, the molecular mechanisms underlying signaling are not well understood. Here, we analyzed transcriptomes of deficient microglia under varying conditions by RNA-sequencing (RNA-seq). In 2-mo-old mice, deletion resulted in the down-regulation of a subset of immune-related genes, without substantial epigenetic changes in markers of active chromatin. Surprisingly, deficient microglia from young mice exhibited a transcriptome consistent with that of aged sufficient animals, suggesting a premature aging transcriptomic signature. Immunohistochemical analysis of microglia in young and aged mice revealed that loss of modulates microglial morphology in a comparable fashion. Our results suggest that CX3CR1 may regulate microglial function in part by modulating the expression levels of a subset of inflammatory genes during chronological aging, making -deficient mice useful for studying aged microglia.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.26508/lsa.201900453DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6892408PMC
December 2019

Neutralization of Oxidized Phospholipids Ameliorates Non-alcoholic Steatohepatitis.

Cell Metab 2020 01 21;31(1):189-206.e8. Epub 2019 Nov 21.

Department of Medicine, University of California, San Diego, La Jolla, CA 92093, USA. Electronic address:

Oxidized phospholipids (OxPLs), which arise due to oxidative stress, are proinflammatory and proatherogenic, but their roles in non-alcoholic steatohepatitis (NASH) are unknown. Here, we show that OxPLs accumulate in human and mouse NASH. Using a transgenic mouse that expresses a functional single-chain variable fragment of E06, a natural antibody that neutralizes OxPLs, we demonstrate the causal role of OxPLs in NASH. Targeting OxPLs in hyperlipidemic Ldlr mice improved multiple aspects of NASH, including steatosis, inflammation, fibrosis, hepatocyte death, and progression to hepatocellular carcinoma. Mechanistically, we found that OxPLs promote ROS accumulation to induce mitochondrial dysfunction in hepatocytes. Neutralizing OxPLs in AMLN-diet-fed Ldlr mice reduced oxidative stress, improved hepatic and adipose-tissue mitochondrial function, and fatty-acid oxidation. These results suggest targeting OxPLs may be an effective therapeutic strategy for NASH.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cmet.2019.10.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7028360PMC
January 2020

Brain cell type-specific enhancer-promoter interactome maps and diseaserisk association.

Science 2019 11 14;366(6469):1134-1139. Epub 2019 Nov 14.

Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA.

Noncoding genetic variation is a major driver of phenotypic diversity, but functional interpretation is challenging. To better understand common genetic variation associated with brain diseases, we defined noncoding regulatory regions for major cell types of the human brain. Whereas psychiatric disorders were primarily associated with variants in transcriptional enhancers and promoters in neurons, sporadic Alzheimer's disease (AD) variants were largely confined to microglia enhancers. Interactome maps connecting disease-risk variants in cell-type-specific enhancers to promoters revealed an extended microglia gene network in AD. Deletion of a microglia-specific enhancer harboring AD-risk variants ablated expression in microglia, but not in neurons or astrocytes. These findings revise and expand the list of genes likely to be influenced by noncoding variants in AD and suggest the probable cell types in which they function.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/science.aay0793DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7028213PMC
November 2019
-->