Fig 2: Cytosolic SUCLA2-mediated stress granules induce anoikis resistance in cancer cells. (A–D) Effect of stress granule disruption on anoikis. Stress granule formation was inhibited by NUFIP2 (A and B) or G3BP1 (C and D) knockdown and its effect on anoikis was assessed by annexin V staining. Disruption of stress granules was confirmed by staining detached cells with stress granule markers G3BP1 or TIA-1. (E and F) Anoikis rates and stress granule formation in detached cells with SUCLA2 knockdown and rescue of stress granules by G3BP1 overexpression. (G and H) Effect of cytosolic localization of SUCLA2 on stress granules. Mitochondria and cytosolic localization of SUCLA2 WT and SUCLA2 mutant lacking mitochondria targeting sequence (MTS) was assessed using fractionated cell lysates (G). 293T cells with SUCLA2 knockdown were transduced with SUCLA2 WT or SUCLA2 del-MTS. Stress granules were induced by 24 h of detachment, 0.5 mM sodium arsenite or 1 mM hydrogen peroxide treatment for 1 h, or GFP-G3BP1 overexpression for 24 h. Stress granules were assessed by G3BP1 staining or GFP. The number of stress granules per cell was quantified using image J software and shown on the top of (B, D, F, and H). Scale bars represent 5 µm for (B, D, F, and H). Error bars represent ± SD from three replicates for (A–F) and (H). P values were obtained by two-tailed Student’s t test for (A–D) and (H), and one-way ANOVA for (E) and (F) (*: 0.01 < P < 0.05; **P < 0.01).