Difference between revisions of "Part:BBa K4040015"
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The function of CAR-MEGF10 is the same as the CARγ, which increases the phagocytosis of the macrophages. And the structure is also highly similar to the CARγ, with the only difference lying in the intracellular domain of the CAR, in which the common γ subunit of Fc receptors is replaced by MEGF10.
 
The function of CAR-MEGF10 is the same as the CARγ, which increases the phagocytosis of the macrophages. And the structure is also highly similar to the CARγ, with the only difference lying in the intracellular domain of the CAR, in which the common γ subunit of Fc receptors is replaced by MEGF10.
 
===The mechanism of CAR-MEGF10===
 
===The mechanism of CAR-MEGF10===
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After the extracellular αS-scFv domain binds to the S protein of SARS-CoV-2, the intracellular cytosolic Immunoreceptor Tyrosine-based Activation Motifs (ITAMs) are activated with the help of the Src family kinases. And the SH2 binding domains for the protein expressing SH2 matching domains will be further exposed due to the space conformation change. The protein expressing SH2 matching domains, protein Syk, a phagocytic signaling effector can then active the downstream signaling transduction cascades and initiate the phagocytosis in our CARγ-macrophages eventually.
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To assay the library of CAR-Ps, previous study has introduced each CAR-P into J774A.1 murine macrophages by lentiviral infection. As an engulfment target, they used 5 mm diameter silica beads coated with a supported lipid bilayer. A His8-tagged extracellular domain of CD19 was bound to a NiNTA-lipid incorporated into the supported lipid bilayers. Macrophages expressing a CAR-P with the Megf10 (CAR�P Megf10) or FcRV (CAR-PFcRV ) intracellular domain promoted significant engulfment of CD19 beads compared to macrophages with no CAR (Figure 1, 2).
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[[File:T--NMU_China--fig3b.png|thumb|center|600px|<b>Figure 1.</b> J774A.1 macrophages expressing aCD19 CAR-P with the indicated intracellular signaling domain (green) engulf 5 mm silica beads covered with a supported lipid bilayer containing His-tagged CD19 extracellular domain. The beads were visualized with atto390-labeled lipid incorporated into the supported lipid bilayer (magenta). Cells infected with the cell membrane marker, mCherry-CAAX, were used as a control (no CAR, top left). To the right of each image is a histogram depicting the frequency of cells engulfing the indicated number of beads.]]
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[[File:T--NMU_China--fig3c.png|thumb|center|350px|<b>Figure 2.</b>  The average number of beads eaten per cell.]]
  
 
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Revision as of 13:01, 15 October 2021


CAR-MEGF10

Background and structure of CAR-MEGF10

The function of CAR-MEGF10 is the same as the CARγ, which increases the phagocytosis of the macrophages. And the structure is also highly similar to the CARγ, with the only difference lying in the intracellular domain of the CAR, in which the common γ subunit of Fc receptors is replaced by MEGF10.

The mechanism of CAR-MEGF10

To assay the library of CAR-Ps, previous study has introduced each CAR-P into J774A.1 murine macrophages by lentiviral infection. As an engulfment target, they used 5 mm diameter silica beads coated with a supported lipid bilayer. A His8-tagged extracellular domain of CD19 was bound to a NiNTA-lipid incorporated into the supported lipid bilayers. Macrophages expressing a CAR-P with the Megf10 (CAR�P Megf10) or FcRV (CAR-PFcRV ) intracellular domain promoted significant engulfment of CD19 beads compared to macrophages with no CAR (Figure 1, 2).

Figure 1. J774A.1 macrophages expressing aCD19 CAR-P with the indicated intracellular signaling domain (green) engulf 5 mm silica beads covered with a supported lipid bilayer containing His-tagged CD19 extracellular domain. The beads were visualized with atto390-labeled lipid incorporated into the supported lipid bilayer (magenta). Cells infected with the cell membrane marker, mCherry-CAAX, were used as a control (no CAR, top left). To the right of each image is a histogram depicting the frequency of cells engulfing the indicated number of beads.
Figure 2. The average number of beads eaten per cell.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 344
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 952
    Illegal NgoMIV site found at 1288
    Illegal NgoMIV site found at 1825
    Illegal NgoMIV site found at 1891
    Illegal NgoMIV site found at 2050
    Illegal AgeI site found at 440
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 1168
    Illegal BsaI.rc site found at 1432
    Illegal SapI.rc site found at 1989
    Illegal SapI.rc site found at 2619