Family of CD93 and Recently Discovered Groups of CTLDs

Efferocytosis is essential for homeostasis and prevention of the inflammatory and autoimmune diseases resulting from apoptotic cell lysis. CD93 is a transmembrane glycoprotein previously implicated in efferocytosis, with mutations in CD93 predisposing patients to efferocytosis‐associated diseases. CD93 is a cell surface protein, which is proteolytically shed under inflammatory conditions, but it is unknown how CD93 mediates efferocytosis or whether its efferocytic activity is mediated by the soluble or membrane‐bound form. Herein, using cell lines and human monocytes and macrophages, we demonstrate that soluble CD93 (sCD93) potently opsonizes apoptotic cells but not a broad range of microorganisms, whereas membrane‐bound CD93 has no phagocytic, efferocytic, or tethering activity. Using mass spectrometry, we identified αxβ2 as the receptor that recognizes sCD93, and via deletion mutagenesis determined that sCD93 binds to apoptotic cells via its C‐type lectin‐like domain and to αxβ2 by its EGF‐like repeats. The bridging of apoptotic cells to αxβ2 markedly enhanced efferocytosis by macrophages and was abrogated by αxβ2 knockdown. Combined, these data elucidate the mechanism by which CD93 regulates efferocytosis and identifies a previously unreported opsonin‐receptor system utilized by phagocytes for the efferocytic clearance of apoptotic cells.

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“…5C). CTLD activity is often dependent on calcium [32], but unexpectedly, sCD93 binding was Ca 2+ independent ( Fig. 5D).…”

Section: Identification Of the Ligand And Receptor Binding Domains Inmentioning

confidence: 92%

Soluble CD93 is an apoptotic cell opsonin recognized by α_xβ₂

et al. 2019

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“…CD93 is a member of the group XIV C-type lectin family, which also includes thrombomodulin, CLEC14A and CD248 (53). While all members of this family contain a C-type lectin-like domain, carbohydrate binding has not been reported for any of these receptors.…”

Section: Discussionmentioning

confidence: 99%

Soluble CD93 is an Apoptotic Cell Opsonin Recognized by the α_xβ₂Integrin

Ellins

et al. 2018

Preprint

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Abbreviations: ADAM -a disintegrin and metalloproteinase. BSA -bovine serum albumin. CHO -Chinese hamster ovary cells. CTLD -C-type lectin-like domain. FBS -fetal bovine serum. MMPmatrix metalloproteinase. PBMC -peripheral blood mononuclear cells. PFA -paraformaldehyde. sCD93 -soluble CD93. ABSTRACTEfferocytosis -the phagocytic removal of apoptotic cells -is essential for the maintenance of homeostasis and prevention of the inflammatory and autoimmune diseases which can follow the lysis of uncleared apoptotic cells. CD93 is a transmembrane glycoprotein previously implicated in efferocytosis and angiogenesis, and upon mutation, results in the onset of efferocytosis-associated diseases such as atherosclerosis and rheumatoid arthritis. CD93 is produced as a cell surface protein which is shed as soluble CD93, but it is unknown how CD93 mediates efferocytosis or whether its efferocytic activity is mediated by the soluble or membrane-bound form. Herein, we demonstrate that the membrane bound form of CD93 has no phagocytic, efferocytic, or tethering activity, whereas soluble CD93 potently opsonizes apoptotic cells but not a broad range of Gram-Negative, Gram-Positive or fungal microorganisms. Using mass spectrometry, we identified the αxβ2 integrin as the receptor required for soluble CD93-mediated efferocytosis, and via deletion mutagenesis determined that soluble CD93 binds to apoptotic cells via its C-Type Lectin-Like domain, and to αxβ2 by its EGF-like repeats. This bridging of apoptotic cells to the αxβ2 integrin markedly enhanced efferocytosis by macrophages, and could be abrogated by knockdown of αxβ2 integrin. Combined, these data elucidate the mechanism by which CD93 regulates efferocytosis and identify a previously unreported opsonin-receptor system utilized by the immune system for the efferocytic clearance of apoptotic cells.Clearly, there is data supporting three distinct and mutually exclusive models of CD93 efferocytic function. In this study, we directly asses these three models of CD93 function, using in vitro models of binding, efferocytosis and opsonization. These experiments disfavor the efferocytic receptor and tethering receptor models of CD93 function, and instead demonstrate a strong opsonic effect of sCD93 against apoptotic cells but not against a panel of microorganisms. Using mass-spectrometry we then identified αxβ2 integrin as the efferocytic receptor which recognizes sCD93 opsonized targets, with apoptotic cells "bridged" by the CD93 CTLD domain to αxβ2 integrin on phagocytes via the CD93 EGFlike domain. This represents a previously undescribed opsonin-receptor system which aids in the efferocytosis of apoptotic cell by phagocytes such as macrophages. MATERIALS AND METHODS MaterialsDH5a Escherichia coli was a gift from Dr. John McCormick (University of Western Ontario), Rhodotorula minuta and Candida glabrate were gifts from Dr. Andre Lachance (University of Western Ontario). Escherichia coli K29 + and K29 -, and Burkholderia cenocepacia were gifts from Dr. Susan Koval (University of Wester...

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Family of CD93 and Recently Discovered Groups of CTLDs

Cited by 2 publications

References 224 publications

Soluble CD93 is an apoptotic cell opsonin recognized by α_xβ₂

Soluble CD93 is an apoptotic cell opsonin recognized by α_xβ₂

Soluble CD93 is an Apoptotic Cell Opsonin Recognized by the α_xβ₂Integrin

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Family of CD93 and Recently Discovered Groups of CTLDs

Cited by 2 publications

References 224 publications

Soluble CD93 is an apoptotic cell opsonin recognized by αxβ2

Soluble CD93 is an apoptotic cell opsonin recognized by αxβ2

Soluble CD93 is an Apoptotic Cell Opsonin Recognized by the αxβ2Integrin

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Soluble CD93 is an apoptotic cell opsonin recognized by α_xβ₂

Soluble CD93 is an apoptotic cell opsonin recognized by α_xβ₂

Soluble CD93 is an Apoptotic Cell Opsonin Recognized by the α_xβ₂Integrin