Fig. 1: SAT1 promoted anchorage-independent cell survival and peritoneal metastasis.

a SKOV3 were transduced with individual shRNAs that targeted each of the 111 rate-limiting metabolic enzymes. The transduced cells were detached 24 h before cell viability measurement. Data are presented as a volcano plot of three biologically independent experiments (two-tailed Student’s t test). shRNAs with P < 0.05 and a relative cell viability level (normalized to the control pLKO) of <0.8 or >1.2 were biologically meaningful. The top hit genes are labeled. b Apoptosis of detached SAT1-knockdown cells rescued with the control vector or the SAT1 recombinant vector rSAT1. c Viability of attached or detached cells (detachment 48 h) with or without SAT1 knockdown. d SAT1 expression detected by western blot in attached or detached cells. e Gene set enrichment analysis (GSEA) of RNA-seq data showing enrichment of KEGG hallmark hypoxia in detached cells relative to attached cells; GEO accession: GSE267614. f Representative microscopy images of detached ovarian cancer cells forming clusters and Image-iT green hypoxia reagent staining. Scale bars represent 20 μm. g Binding of HIF-1α to the SAT1 promoter in detached cells determined by a ChIP assay followed by qPCR. h–l Effect of SAT1 knockdown on the ID8 peritoneal dissemination model. h Experimental design (upper panel) and confirmation of SAT1 knockdown by western blot (lower panel). i–j, Average photon flux (i) and representative bioluminescence image (j) of tumors at the indicated timepoints. k, l Representative images (k) and quantification (l) of hemorrhagic ascites and peritoneal tumor nodules in mice at the endpoint. Data shown are the means ± SDs from 3 biological replicates for (b), (c), and (g) and are representative of 3 independent biological experiments for (d) and (f). The error bars represent the SDs of the tumor luminescence signals (i). Statistical analysis was performed via 1-way ANOVA for (b), (c) and (g); 2-way ANOVA for (i); and two-tailed Student’s t test for (l). The samples derive from the same experiment but different gels for SAT1 and ACTB were processed in parallel for (d) and (h). See also supplementary Figs. S1–S3.