title: The nature and nurture of cell heterogeneity: accounting for macrophage gene-environment interactions with single-cell RNA-Seq.
abstract: Single-cell RNA-Seq can be a valuable and unbiased tool to dissect cellular heterogeneity, despite the transcriptome's limitations in describing higher functional phenotypes and protein events. Perhaps the most important shortfall with transcriptomic 'snapshots' of cell populations is that they risk being descriptive, only cataloging heterogeneity at one point in time, and without microenvironmental context. Studying the genetic ('nature') and environmental ('nurture') modifiers of heterogeneity, and how cell population dynamics unfold over time in response to these modifiers is key when studying highly plastic cells such as macrophages.
  
  
  
  We introduce the programmable Polaris™ microfluidic lab-on-chip for single-cell sequencing, which performs live-cell imaging while controlling for the culture microenvironment of each cell. Using gene-edited macrophages we demonstrate how previously unappreciated knockout effects of SAMHD1, such as an altered oxidative stress response, have a large paracrine signaling component. Furthermore, we demonstrate single-cell pathway enrichments for cell cycle arrest and APOBEC3G degradation, both associated with the oxidative stress response and altered proteostasis. Interestingly, SAMHD1 and APOBEC3G are both HIV-1 inhibitors ('restriction factors'), with no known co-regulation.
  
  
  
  As single-cell methods continue to mature, so will the ability to move beyond simple 'snapshots' of cell populations towards studying the determinants of population dynamics. By combining single-cell culture, live-cell imaging, and single-cell sequencing, we have demonstrated the ability to study cell phenotypes and microenvironmental influences. It's these microenvironmental components - ignored by standard single-cell workflows - that likely determine how macrophages, for example, react to inflammation and form treatment resistant HIV reservoirs.
authors.affiliation: Wellcome Trust Centre for Human Genetics (WTCHG), University of Oxford, Oxford, OX3 7BN, UK. qilin@well.ox.ac.uk.|Weatherall Institute of Molecular Medicine (WIMM), University of Oxford, Oxford, OX3 9DS, UK. qilin@well.ox.ac.uk.
authors.forename: Quin F
authors.initials: QF
authors.lastname: Wills
authors.affiliation: Wellcome Trust Centre for Human Genetics (WTCHG), University of Oxford, Oxford, OX3 7BN, UK.
authors.forename: Esther
authors.initials: E
authors.lastname: Mellado-Gomez
authors.affiliation: Wellcome Trust Centre for Human Genetics (WTCHG), University of Oxford, Oxford, OX3 7BN, UK.|Division of Structural Biology, University of Oxford, Oxford, OX3 7BN, UK.
authors.forename: Rory
authors.initials: R
authors.lastname: Nolan
authors.affiliation: Sir William Dunn School of Pathology, University of Oxford, Oxford, OX1 3RE, UK.
authors.forename: Damien
authors.initials: D
authors.lastname: Warner
authors.affiliation: Wellcome Trust Centre for Human Genetics (WTCHG), University of Oxford, Oxford, OX3 7BN, UK.
authors.forename: Eshita
authors.initials: E
authors.lastname: Sharma
authors.affiliation: Wellcome Trust Centre for Human Genetics (WTCHG), University of Oxford, Oxford, OX3 7BN, UK.
authors.forename: John
authors.initials: J
authors.lastname: Broxholme
authors.affiliation: Wellcome Trust Centre for Human Genetics (WTCHG), University of Oxford, Oxford, OX3 7BN, UK.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Wright
authors.affiliation: Wellcome Trust Centre for Human Genetics (WTCHG), University of Oxford, Oxford, OX3 7BN, UK.
authors.forename: Helen
authors.initials: H
authors.lastname: Lockstone
authors.affiliation: Sir William Dunn School of Pathology, University of Oxford, Oxford, OX1 3RE, UK.
authors.forename: William
authors.initials: W
authors.lastname: James
authors.affiliation: Fluidigm Corporation, 7000 Shoreline Ct Ste 100, South San Francisco, CA, 94080-7603, USA.
authors.forename: Mark
authors.initials: M
authors.lastname: Lynch
authors.affiliation: Fluidigm Corporation, 7000 Shoreline Ct Ste 100, South San Francisco, CA, 94080-7603, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Gonzales
authors.affiliation: Fluidigm Corporation, 7000 Shoreline Ct Ste 100, South San Francisco, CA, 94080-7603, USA.
authors.forename: Jay
authors.initials: J
authors.lastname: West
authors.affiliation: Fluidigm Corporation, 7000 Shoreline Ct Ste 100, South San Francisco, CA, 94080-7603, USA.
authors.forename: Anne
authors.initials: A
authors.lastname: Leyrat
authors.affiliation: Wellcome Trust Centre for Human Genetics (WTCHG), University of Oxford, Oxford, OX3 7BN, UK.|Division of Structural Biology, University of Oxford, Oxford, OX3 7BN, UK.
authors.forename: Sergi
authors.initials: S
authors.lastname: Padilla-Parra
authors.affiliation: Department of Statistics, University of Oxford, Oxford, OX3 3LB, UK.
authors.forename: Sarah
authors.initials: S
authors.lastname: Filippi
authors.affiliation: Wellcome Trust Centre for Human Genetics (WTCHG), University of Oxford, Oxford, OX3 7BN, UK.|Department of Statistics, University of Oxford, Oxford, OX3 3LB, UK.
authors.forename: Chris
authors.initials: C
authors.lastname: Holmes
authors.affiliation: Sir William Dunn School of Pathology, University of Oxford, Oxford, OX1 3RE, UK. kenny.moore@path.ox.ac.uk.
authors.forename: Michael D
authors.initials: MD
authors.lastname: Moore
authors.affiliation: Wellcome Trust Centre for Human Genetics (WTCHG), University of Oxford, Oxford, OX3 7BN, UK. rbowden@well.ox.ac.uk.
authors.forename: Rory
authors.initials: R
authors.lastname: Bowden
mesh_terms: D055786:Gene Knockout Techniques; D059647:Gene-Environment Interaction; D006801:Humans; D008264:Macrophages / Q000166:cytology* / Q000378:metabolism; D010641:Phenotype; D000076106:SAM Domain and HD Domain-Containing Protein 1 / Q000172:deficiency / Q000235:genetics; D017423:Sequence Analysis, RNA; D059010:Single-Cell Analysis
chemical_list: D000076106:SAM Domain and HD Domain-Containing Protein 1
keywords: Macrophage heterogeneity; Signaling microenvironment; Single-cell culture; Single-cell imaging; Single-cell sequencing