Identification of phagocytosis regulators using magnetic genome-wide CRISPR screens

Haney, Michael S.; Bohlen, Christopher J.; Morgens, David W.; Ousey, James; Barkal, Amira; Tsui, C. Kimberly; Ego, Braeden K.; Levin, Roni; Kamber, Roarke A.; Collins, Hannah Y.; Tucker, Andrew F.; Li, Amy; Vorselen, Daan; Labitigan, Lorenzo; Crane, Emily; Boyle, Evan; Jiang, Lihua; Chan, Joanne; Rincón, Esther; Greenleaf, William J.; Li, Billy; Snyder, M; Weissman, Irving L.; Theriot, Julie A.; Collins, Sean R.; Barres, Ben A.; Bassik, Michael C.

doi:10.1038/s41588-018-0254-1

Cited by 157 publications

(178 citation statements)

References 63 publications

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“…Importantly, phagocytosis is the process responsible for the induction of the pro-to anti-phenotypical switch in MPs. While numerous studies investigated the mechanisms involved in the progression or inhibition of phagocytosis, how apoptotic cells attract MPs and how MPs recognize them is still unknown [10,14,15,50,51]. In our study, phagocytosis of LysM CRE :Nfix fl/fl MPs was not impaired compared to control cells.…”

Section: Discussioncontrasting

confidence: 43%

The Transcription Factor Nfix Requires RhoA-ROCK1 Dependent Phagocytosis to Mediate Macrophage Skewing during Skeletal Muscle Regeneration

Saclier

Lapi

Bonfanti

et al. 2020

Cells

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Macrophages (MPs) are immune cells which are crucial for tissue repair. In skeletal muscle regeneration, pro-inflammatory cells first infiltrate to promote myogenic cell proliferation, then they switch into an anti-inflammatory phenotype to sustain myogenic cells differentiation and myofiber formation. This phenotypical switch is induced by dead cell phagocytosis. We previously demonstrated that the transcription factor Nfix, a member of the nuclear factor I (Nfi) family, plays a pivotal role during muscle development, regeneration and in the progression of muscular dystrophies. Here, we show that Nfix is mainly expressed by anti-inflammatory macrophages. Upon acute injury, mice deleted for Nfix in myeloid line displayed a significant defect in the process of muscle regeneration. Indeed, Nfix is involved in the macrophage phenotypical switch and macrophages lacking Nfix failed to adopt an anti-inflammatory phenotype and interact with myogenic cells. Moreover, we demonstrated that phagocytosis induced by the inhibition of the RhoA-ROCK1 pathway leads to Nfix expression and, consequently, to acquisition of the anti-inflammatory phenotype. Our study identified Nfix as a link between RhoA-ROCK1-dependent phagocytosis and the MP phenotypical switch, thus establishing a new role for Nfix in macrophage biology for the resolution of inflammation and tissue repair.

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Section: Discussioncontrasting

confidence: 43%

The Transcription Factor Nfix Requires RhoA-ROCK1 Dependent Phagocytosis to Mediate Macrophage Skewing during Skeletal Muscle Regeneration

Saclier

Lapi

Bonfanti

et al. 2020

Cells

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“…Jakobsdottir et al () identified a rare missense (p.P155L) variant affecting the C‐Type Lectin‐like domain (CTLD) of TM2D3 as a risk allele for late‐onset Alzheimer's disease in an Icelandic population, and proposed that Amx/TM2D3 may contribute directly or indirectly to the γ‐secretase cleavage of Notch and APP, a hypothesis that needs to be tested biochemically. More recently, Haney et al () identified TM2D1–3 as phagocytosis regulators based on CRISPR‐based screens using human phagocytic cell lines and proposed that these proteins may be involved in Alzheimer's disease by regulating amyloid clearance in the brain. This hypothesis should also be tested by biochemical analysis.…”

Section: Introductionmentioning

confidence: 99%

Maternal almondex, a neurogenic gene, is required for proper subcellular Notch distribution in early Drosophila embryogenesis

et al. 2019

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Notch signaling plays crucial roles in the control of cell fate and physiology through local cell–cell interactions. The core processes of Notch signal transduction are well established, but the mechanisms that fine‐tune the pathway in various developmental and post‐developmental contexts are less clear. Drosophila almondex, which encodes an evolutionarily conserved double‐pass transmembrane protein, was identified in the 1970s as a maternal‐effect gene that regulates Notch signaling in certain contexts, but its mechanistic function remains obscure. In this study, we examined the role of almondex in Notch signaling during early Drosophila embryogenesis. We found that in addition to being required for lateral inhibition in the neuroectoderm, almondex is also partially required for Notch signaling‐dependent single‐minded expression in the mesectoderm. Furthermore, we found that almondex is required for proper subcellular Notch receptor distribution in the neuroectoderm, specifically during mid‐stage 5 development. The absence of maternal almondex during this critical window of time caused Notch to accumulate abnormally in cells in a mesh‐like pattern. This phenotype did not include any obvious change in subcellular Delta ligand distribution, suggesting that it does not result from a general vesicular‐trafficking defect. Considering that dynamic Notch trafficking regulates signal output to fit the specific context, we speculate that almondex may facilitate Notch activation by regulating intracellular Notch receptor distribution during early embryogenesis.

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“…Comparatively, this route is difficult, expensive, and time-consuming (Sugimura et al, 2017). Another approach is to edit fully differentiated macrophages, which has shown some functional success in the literature (Barkal et al, 2017;Haney et al, 2018), though this also has limitations. The approach is specific to macrophages, rather than allowing for flexible differentiation, and is limited by the lifespan of mature macrophages in culture.…”

Section: Discussionmentioning

confidence: 99%

Efficient Generation of Isogenic Primary Human Myeloid Cells using CRISPR-Cas9 Ribonucleoproteins

Hiatt¹,

Cavero

McGregor

et al. 2020

Preprint

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Genome engineering of primary human cells with CRISPR-Cas9 has revolutionized experimental and therapeutic approaches to cell biology, but human myeloidlineage cells have remained largely genetically intractable. We present a method for delivery of CRISPR-Cas9 ribonucleoprotein (RNP) complexes by nucleofection directly into CD14+ human monocytes purified from peripheral blood, leading to high rates of precise gene knockout. These cells can be efficiently differentiated into monocyte-derived macrophages or dendritic cells. This process yields genetically-edited cells that retain critical markers of both myeloid differentiation and phagocytic function. Genetic ablation of the restriction factor SAMHD1 increased HIV-1 infection more than fifty-fold, demonstrating the power of this system for genotype-phenotype interrogation. This fast, flexible and scalable platform can be used for genetic studies of human myeloid cells in immune signaling, inflammation, cancer immunology, host-pathogen interactions, and beyond, and could facilitate development of novel myeloid cellular therapies.

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Identification of phagocytosis regulators using magnetic genome-wide CRISPR screens

Cited by 157 publications

References 63 publications

The Transcription Factor Nfix Requires RhoA-ROCK1 Dependent Phagocytosis to Mediate Macrophage Skewing during Skeletal Muscle Regeneration

The Transcription Factor Nfix Requires RhoA-ROCK1 Dependent Phagocytosis to Mediate Macrophage Skewing during Skeletal Muscle Regeneration

Maternal almondex, a neurogenic gene, is required for proper subcellular Notch distribution in early Drosophila embryogenesis

Efficient Generation of Isogenic Primary Human Myeloid Cells using CRISPR-Cas9 Ribonucleoproteins

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