Multidimensional Single-Nuclei RNA-Seq Reconstruction of Adipose Tissue Reveals Adipocyte Plasticity Underlying Thermogenic Response

Adipose tissue has been classified based on its morphology and function as white, brown, or beige / brite. It plays an essential role as a regulator of systemic metabolism through paracrine and endocrine signals. Recently, multiple adipocyte subtypes have been revealed using RNA sequencing technology, going beyond simply defined morphology but by their cellular origin, adaptation to metabolic stress, and plasticity. Here, we performed an in-depth analysis of publicly available single-nuclei RNAseq from adipose tissue and utilized a workflow template to characterize adipocyte plasticity, heterogeneity, and secretome profiles. The reanalyzed dataset led to the identification of different subtypes of adipocytes including three subpopulations of thermogenic adipocytes and provided a characterization of distinct transcriptional profiles along the adipocyte trajectory under thermogenic challenges. This study provides a useful resource for further investigations regarding mechanisms related to adipocyte plasticity and trans-differentiation. Highlights Multidimensional transcriptome analysis at single-nucleus resolution recovers nuclei of cell types in adipose tissue Adaptative thermogenic response results in 3 distinct mature adipose cell types Single-nuclei transcriptomic-based secretome analysis reveals adipose cell-type-specific genes The in vivo trajectory of adipocyte plasticity for thermogenic response reveals sets of trans-differentiation genes Graphic Abstract

[1]  B. Spiegelman,et al.  Mitochondrial TNAP controls thermogenesis by hydrolysis of phosphocreatine , 2021, Nature.

[2]  B. Spiegelman,et al.  Creatine kinase B controls futile creatine cycling in thermogenic fat , 2021, Nature.

[3]  Joseph E. Powell,et al.  Nebulosa recovers single-cell gene expression signals by kernel density estimation , 2021, Bioinform..

[4]  S. Mandrup,et al.  Plasticity of Epididymal Adipose Tissue in Response to Diet-Induced Obesity at Single-Nucleus Resolution. , 2020, Cell metabolism.

[5]  S. Farmer,et al.  Shifts of Immune Cell Populations Differ in Response to Different Effectors of Beige Remodeling of Adipose Tissue , 2020, iScience.

[6]  A. Regev,et al.  snRNA-seq reveals a subpopulation of adipocytes that regulates thermogenesis , 2020, Nature.

[7]  Lawrence M. Lifshitz,et al.  Single-Cell RNA Profiling Reveals Adipocyte to Macrophage Signaling Sufficient to Enhance Thermogenesis , 2020, Cell reports.

[8]  J. Pessin,et al.  The Impact of Single-Cell Genomics on Adipose Tissue Research , 2020, International journal of molecular sciences.

[9]  Alice E. Pollard,et al.  Thermogenic adipocytes: lineage, function and therapeutic potential , 2020, The Biochemical journal.

[10]  Mirjana Efremova,et al.  CellPhoneDB: inferring cell–cell communication from combined expression of multi-subunit ligand–receptor complexes , 2020, Nature Protocols.

[11]  Warren A. Cheung,et al.  Single-cell analysis of human adipose tissue identifies depot- and disease-specific cell types , 2019, Nature Metabolism.

[12]  P. Scherer,et al.  Dermal adipose tissue has high plasticity and undergoes reversible dedifferentiation in mice. , 2019, The Journal of clinical investigation.

[13]  D. Cho,et al.  Refining the adipose progenitor cell landscape in healthy and obese visceral adipose tissue using single-cell gene expression profiling , 2019, Life Science Alliance.

[14]  V. Gradinaru,et al.  Low- and high-thermogenic brown adipocyte subpopulations coexist in murine adipose tissue. , 2019, The Journal of clinical investigation.

[15]  Gennady Korotkevich,et al.  Fast gene set enrichment analysis , 2019, bioRxiv.

[16]  A. Tanay,et al.  MetaCell: analysis of single-cell RNA-seq data using K-nn graph partitions , 2019, Genome Biology.

[17]  C. Schéele,et al.  Human brown adipose tissue is phenocopied by classical brown adipose tissue in physiologically humanized mice , 2019, Nature Metabolism.

[18]  Qingbo Xu,et al.  Single-Cell RNA-Sequencing and Metabolomics Analyses Reveal the Contribution of Perivascular Adipose Tissue Stem Cells to Vascular Remodeling , 2019, Arteriosclerosis, thrombosis, and vascular biology.

[19]  K. Moore,et al.  Single-Cell RNA Sequencing of Visceral Adipose Tissue Leukocytes Reveals that Caloric Restriction Following Obesity Promotes the Accumulation of a Distinct Macrophage Population with Features of Phagocytic Cells , 2019, Immunometabolism.

[20]  I. Amit,et al.  Lipid-Associated Macrophages Control Metabolic Homeostasis in a Trem2-Dependent Manner , 2019, Cell.

[21]  Luis C. Santos,et al.  Single cell analysis reveals immune cell–adipocyte crosstalk regulating the transcription of thermogenic adipocytes , 2019, bioRxiv.

[22]  Paul J. Hoffman,et al.  Comprehensive Integration of Single-Cell Data , 2018, Cell.

[23]  C. Benoist,et al.  Distinct immunocyte-promoting and adipocyte-generating stromal components coordinate adipose tissue immune and metabolic tenors , 2019, Science Immunology.

[24]  P. O’Brien,et al.  Identification of Metabolically Distinct Adipocyte Progenitor Cells in Human Adipose Tissues. , 2019, Cell reports.

[25]  Ivona Percec,et al.  Identification of a mesenchymal progenitor cell hierarchy in adipose tissue , 2019, Science.

[26]  Samantha Riesenfeld,et al.  EmptyDrops: distinguishing cells from empty droplets in droplet-based single-cell RNA sequencing data , 2019, Genome Biology.

[27]  R. Satija,et al.  Normalization and variance stabilization of single-cell RNA-seq data using regularized negative binomial regression , 2019, Genome Biology.

[28]  Konstantinos D. Tsirigos,et al.  SignalP 5.0 improves signal peptide predictions using deep neural networks , 2019, Nature Biotechnology.

[29]  B. Spiegelman,et al.  New Advances in Adaptive Thermogenesis: UCP1 and Beyond. , 2019, Cell metabolism.

[30]  Damian Szklarczyk,et al.  STRING v11: protein–protein association networks with increased coverage, supporting functional discovery in genome-wide experimental datasets , 2018, Nucleic Acids Res..

[31]  The UniProt Consortium,et al.  UniProt: a worldwide hub of protein knowledge , 2018, Nucleic Acids Res..

[32]  S. B. Peres,et al.  Toll-Like Receptor-4 Disruption Suppresses Adipose Tissue Remodeling and Increases Survival in Cancer Cachexia Syndrome , 2018, Scientific Reports.

[33]  Kerstin B. Meyer,et al.  Single-cell reconstruction of the early maternal–fetal interface in humans , 2018, Nature.

[34]  S. Kajimura,et al.  Thermal stress induces glycolytic beige fat formation via a myogenic state , 2018, Nature.

[35]  James T. Webber,et al.  Single-cell transcriptomics of 20 mouse organs creates a Tabula Muris , 2018, Nature.

[36]  M. Batista,et al.  Physical Exercise-Induced Myokines and Muscle-Adipose Tissue Crosstalk: A Review of Current Knowledge and the Implications for Health and Metabolic Diseases , 2018, Front. Physiol..

[37]  Quy H. Nguyen,et al.  Experimental Considerations for Single-Cell RNA Sequencing Approaches , 2018, Front. Cell Dev. Biol..

[38]  Y. Kluger,et al.  Zero-preserving imputation of scRNA-seq data using low-rank approximation , 2018, bioRxiv.

[39]  Mengle Shao,et al.  Identification of functionally distinct fibro-inflammatory and adipogenic stromal subpopulations in visceral adipose tissue of adult mice , 2018, eLife.

[40]  R. Pique-Regi,et al.  Deconstructing Adipogenesis Induced by β3-Adrenergic Receptor Activation with Single-Cell Expression Profiling. , 2018, Cell metabolism.

[41]  John C Marioni,et al.  Detection and removal of barcode swapping in single-cell RNA-seq data , 2017, Nature Communications.

[42]  Petra C. Schwalie,et al.  A stromal cell population that inhibits adipogenesis in mammalian fat depots , 2018, Nature.

[43]  Denis Torre,et al.  eXpression2Kinases (X2K) Web: linking expression signatures to upstream cell signaling networks , 2018, Nucleic Acids Res..

[44]  M. Kumari,et al.  Warming Induces Significant Reprogramming of Beige, but Not Brown, Adipocyte Cellular Identity. , 2018, Cell metabolism.

[45]  Paul Hoffman,et al.  Integrating single-cell transcriptomic data across different conditions, technologies, and species , 2018, Nature Biotechnology.

[46]  J. B. Kim,et al.  Adipocyte CD1d determines adipose inflammation and insulin resistance in obesity , 2018, Adipocyte.

[47]  Fabian J Theis,et al.  SCANPY: large-scale single-cell gene expression data analysis , 2018, Genome Biology.

[48]  S. Joost,et al.  Single cell transcriptomics suggest that human adipocyte progenitor cells constitute a homogeneous cell population , 2017, Stem Cell Research & Therapy.

[49]  J. Graff,et al.  Distinct cellular and molecular mechanisms for β3 adrenergic receptor-induced beige adipocyte formation , 2017, eLife.

[50]  Hannah A. Pliner,et al.  Reversed graph embedding resolves complex single-cell trajectories , 2017, Nature Methods.

[51]  F. Coulpier,et al.  Egr1 deficiency induces browning of inguinal subcutaneous white adipose tissue in mice , 2017, bioRxiv.

[52]  M. Zoratti,et al.  Now UCP(rotein), Now You Don't: UCP1 Is Not Mandatory for Thermogenesis. , 2017, Cell metabolism.

[53]  I. Hellmann,et al.  Comparative Analysis of Single-Cell RNA Sequencing Methods , 2016, bioRxiv.

[54]  Andrew J. Hill,et al.  Single-cell mRNA quantification and differential analysis with Census , 2017, Nature Methods.

[55]  J. Granneman,et al.  Metabolic heterogeneity of activated beige/brite adipocytes in inguinal adipose tissue , 2017, Scientific Reports.

[56]  Grace X. Y. Zheng,et al.  Massively parallel digital transcriptional profiling of single cells , 2016, Nature Communications.

[57]  S. Farmer,et al.  Browning of White Adipose Tissue with Roscovitine Induces a Distinct Population of UCP1+ Adipocytes. , 2016, Cell metabolism.

[58]  Roland Eils,et al.  Complex heatmaps reveal patterns and correlations in multidimensional genomic data , 2016, Bioinform..

[59]  Andrew D. Rouillard,et al.  Enrichr: a comprehensive gene set enrichment analysis web server 2016 update , 2016, Nucleic Acids Res..

[60]  Charles H. Yoon,et al.  Dissecting the multicellular ecosystem of metastatic melanoma by single-cell RNA-seq , 2016, Science.

[61]  B. Spiegelman,et al.  A Creatine-Driven Substrate Cycle Enhances Energy Expenditure and Thermogenesis in Beige Fat , 2015, Cell.

[62]  Bruce M. Spiegelman,et al.  Brown and Beige Fat: Physiological Roles beyond Heat Generation. , 2015, Cell metabolism.

[63]  Juancarlos Chan,et al.  Gene Ontology Consortium: going forward , 2014, Nucleic Acids Res..

[64]  Yi Pan,et al.  CytoNCA: A cytoscape plugin for centrality analysis and evaluation of protein interaction networks , 2015, Biosyst..

[65]  Pirjo Nuutila,et al.  [Human brown adipose tissue]. , 2015, Duodecim; laaketieteellinen aikakauskirja.

[66]  Zhimin Chen,et al.  The brown fat-enriched secreted factor Nrg 4 preserves metabolic homeostasis through attenuating hepatic lipogenesis , 2016 .

[67]  B. Spiegelman,et al.  Tumour-derived PTH-related protein triggers adipose tissue browning and cancer cachexia , 2014, Nature.

[68]  Philippe Bardou,et al.  jvenn: an interactive Venn diagram viewer , 2014, BMC Bioinformatics.

[69]  Cole Trapnell,et al.  Pseudo-temporal ordering of individual cells reveals dynamics and regulators of cell fate decisions , 2014, Nature Biotechnology.

[70]  G. Montana,et al.  Brown and white adipose tissues: intrinsic differences in gene expression and response to cold exposure in mice , 2014, American journal of physiology. Endocrinology and metabolism.

[71]  Walter de Back,et al.  Morpheus: a user-friendly modeling environment for multiscale and multicellular systems biology , 2014, Bioinform..

[72]  C. Wolfrum,et al.  The origin and definition of brite versus white and classical brown adipocytes , 2013, Adipocyte.

[73]  J. Granneman,et al.  Identification of an adipogenic niche for adipose tissue remodeling and restoration. , 2013, Cell metabolism.

[74]  T. Rülicke,et al.  Bi-directional interconversion of brite and white adipocytes , 2013, Nature Cell Biology.

[75]  Edward Y. Chen,et al.  Enrichr: interactive and collaborative HTML5 gene list enrichment analysis tool , 2013, BMC Bioinformatics.

[76]  Kyoung-Jae Won,et al.  EBF2 determines and maintains brown adipocyte identity. , 2013, Cell metabolism.

[77]  Yuanyuan Wu,et al.  Adipose tissue heterogeneity: implication of depot differences in adipose tissue for obesity complications. , 2013, Molecular aspects of medicine.

[78]  B. Spiegelman,et al.  Beige Adipocytes Are a Distinct Type of Thermogenic Fat Cell in Mouse and Human , 2012, Cell.

[79]  D. Jacobsen,et al.  Ethanol-induced oxidative stress via the CYP2E1 pathway disrupts adiponectin secretion from adipocytes. , 2012, Alcoholism, clinical and experimental research.

[80]  Richard J. Simpson,et al.  ExoCarta as a resource for exosomal research , 2012, Journal of extracellular vesicles.

[81]  Avi Ma'ayan,et al.  Expression2Kinases: mRNA profiling linked to multiple upstream regulatory layers , 2012, Bioinform..

[82]  B. Spiegelman,et al.  Transcriptional control of brown fat development. , 2010, Cell metabolism.

[83]  B. Spiegelman,et al.  Transcriptional control of brown adipocyte development and physiological function--of mice and men. , 2009, Genes & development.

[84]  Sean R. Davis,et al.  GEOquery: a bridge between the Gene Expression Omnibus (GEO) and BioConductor , 2007, Bioinform..

[85]  P. Scherer.  Adipose Tissue , 2006, Diabetes.

[86]  N. Blom,et al.  Feature-based prediction of non-classical and leaderless protein secretion. , 2004, Protein engineering, design & selection : PEDS.

[87]  P. Shannon,et al.  Cytoscape: a software environment for integrated models of biomolecular interaction networks. , 2003, Genome research.

[88]  A. Krogh,et al.  Predicting transmembrane protein topology with a hidden Markov model: application to complete genomes. , 2001, Journal of molecular biology.

[89]  P. Puigserver,et al.  A Cold-Inducible Coactivator of Nuclear Receptors Linked to Adaptive Thermogenesis , 1998, Cell.

[90]  B. Spiegelman,et al.  Adipose expression of tumor necrosis factor-alpha: direct role in obesity-linked insulin resistance. , 1993, Science.

[91]  J. Friedman,et al.  Molecular mapping of the mouse ob mutation. , 1991, Genomics.