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. 2021 Aug 9;22(9):e52262. doi: 10.15252/embr.202052262

Figure EV1. NimB4 is induced by clean injury.

Figure EV1

  • A
    Representative confocal images from confocal stacks of NimB4‐sGFP third‐instar larval brains. Tissues were stained with anti‐GFP (green, corresponding to NimB4‐sGFP) and anti‐Draper (red, corresponding to glial cells). The arrows indicate the presence of NimB4‐sGFP in the glial cells of third‐instar larval brain. Scale bars = 100 μm.
  • B, C
    Quantification of the NimB4 transcripts, relative to RpL32 expression from macrophages (B) and fat body (C) extracts of wild‐type (w1118 ) animals in unchallenged conditions (UC) and 1, 2, 6 h after clean injury. Statistics compare a challenged sample and its own unchallenged sample. Data are represented as mean ± SD from three independent experiments with five animals each (*P < 0.05 by ANOVA test followed by post hoc Dunnett's multiple comparison tests. ns: not significant).
  • D
    Western blot analysis of hemolymph from L3 larvae expressing NimB4‐RFP, NimB5‐RFP, or SPvkg‐RFP from the fat body with the driver Lpp‐Gal4 driver. (NimB4: 77.4 kDa, NimB5: 62.3 kDa and RFP: 27 kDa, Tubulin: 55 kDa).
  • E
    Representative images of L3 larvae overexpressing NimB4‐RFP, SPvkg‐RFP (a secreted RFP), or CD8‐GFP in the fat body (CD8‐GFP is a membrane‐addressed GFP used to confirm that the Gal4 driver is not expressed in the nephrocytes) in the fat body. NimB4‐RFP and SPvkg‐RFP accumulate in the pericardial nephrocytes indicating that both proteins are secreted into the hemolymph. CD8‐GFP is absent from the pericardial nephrocyte indicating that the Lpp driver is not expressed in this tissue. The arrows show the presence of RFP in the pericardial nephrocytes of the larvae. The brightness of the image is modified to improve visualization of the signal.
  • F
    Representative images of L3 larvae overexpressing NimB4‐RFP, SPvkg‐RFP, or CD8‐GFP in the hemocyte. NimB4‐RFP and SPvkg‐RFP accumulate in the pericardial nephrocytes indicating that both proteins are secreted into the hemolymph. CD8‐GFP is absent from the pericardial nephrocyte indicating that the Hml driver is not expressed in nephrocytes. The arrows show the presence of RFP in the pericardial nephrocytes of the larvae. The brightness of the image is modified to improve visualization of the signal.