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. 2024 Oct 29;221(11):e20231832. doi: 10.1084/jem.20231832

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© 2024 Hsu et al.

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Figure 1. — IMPA1/inositol axis is enriched in ALDH-positive PCSCs and maintains prostate cancer stemness. (A and B) Dot plots show the upregulation of genes and metabolites in ALDH^high compared with ALDH^low/− of PC3 cells. PC3 cells were isolated using FACS based on ALDH expression. (C) Metabolic profiling of PC3 or DU145 cells expressing shLuc or shIMPA1 was revealed by targeted mass spectrometry analysis of phosphoinositides metabolism including inositol (Ins), PI, inositol 1,3,4-trisphosphate (IP₃), phosphatidylinositol 3-phosphate (PIP), and inositol 4-phosphate (IP). Each metabolite in the shIMPA1 group was normalized by shLuc showing relative peak intensity. The mean ± SEM showed three independent experiments for each group. *, P < 0.05; **, P < 0.01 by two-tailed unpaired t test. (D) The level of inositol of PC3 cells stably expressing shLuc or IMPA1-two specific shRNA lentivirus (#1 and #2) was determined using a K-INOSL assay kit according to the manufacturer’s instructions. The inositol levels were normalized by protein concentration in each experimental group. The mean ± SEM showed three independent experiments for each group. *, P < 0.05; **, P < 0.01 by two-tailed unpaired t test. (E) Spheres from LNCaP, PC3, DU145, and TRAMP-C2 stably expressing shLuc or IMPA1-two specific shRNA lentivirus (#1 and #2) were shown. Scale bar, 200 μm. (F–I) Quantification of the number of spheres from E was shown for LNCaP (F), PC3 (G), DU145 (H), and TRAMP-C2 (I). The mean ± SEM showed three independent experiments for each group. **, P < 0.01; ***, P < 0.001 by two-tailed unpaired t test. (J–M) Immunoblotting of LNCaP (J), PC3 (K), DU145 (L), and TRAMP-C2 (M) cells stably expressing shLuc or shIMPA1 (#1 and #2) with indicated antibodies. Immunoblotting data were verified in at least two independent experiments. Source data are available for this figure: SourceData F1.