Figure 6. Zn²⁺ promotes condensation and the DNA‐induced LLPS of cGAS.

• A
  Bright‐field microscope images of indicated groups (left) and a quantitative analysis of total area of droplets (right). n = 3 biological replicates. 10 μM cGAS, 5 μM G3BP1 and 50 μM KCl, 50 μM MgCl₂, 50 μM ZnCl₂ were used.
• B
  cGAMP production assays. Recombinant cGAS (10 μM) and indicated ions (100 μM) were incubated with or without dsDNA (100 nM). The production of cGAMP was analyzed by LC‐MS/MRM. n = 3 technical replicates.
• C
  Fluorescent images and analysis of DNA‐induced cGAS LLPS in the presence of the indicated reagent. n = 3 biological replicates. 10 μM cGAS, 50 μM of each ion and 100 nM dsDNA were used in these assays.
• D
  Fluorescent images and analysis of DNA‐induced LLPS of cGAS in the presence of different amounts of ZnCl₂. n = 3 biological replicates. 10 μM cGAS and 100 nM dsDNA were used in the assay.
• E
  Fluorescent images and analysis of DNA‐induced LLPS of cGAS in the presence of G3BP1 (2 μM) and/or ZnCl₂ (20 μM) as indicated. n = 3 biological replicates.
• F
  cGAMP production assays. Recombinant cGAS (10 μM), G3BP1(10 μM) and ZnCl₂ (100 μM) were incubated with or without dsDNA (100 nM) as indicated. The produced cGAMP was quantitatively analyzed by LC‐MS/MRM. n = 3 technical replicates.
• G
  Immunofluorescence staining of cGAS in HeLa cells stimulated with ZnCl₂ (100 μM). The images were acquired by ZEISS LSM 880 Confocal Microscopy.

Data information: Representative images are shown (A, C–E and G). Scale bars, 10 μm. The partition coefficient was calculated as the total fluorescence intensity of droplets/bulk fluorescence intensity of background (C–E). Hoechst (blue), nuclear staining. Error bars, mean with s.d. (A‐F), *P < 0.05, **P < 0.01, ***P < 0.001, two‐tailed t‐test. NS, non‐significant; NT, non‐treated.