Fig 2: Evolutionary analysis of LZTR1 and the RAS proteins KRAS and RIT1.(a) Phylogenetic trees of KRAS, RIT1, and LZTR1 orthologs based on multiple protein sequence alignments performed with Clustal Omega (Figure 1—source data 1). Orthologs were searched in the following model organisms: chimpanzee (Pan troglodytes), pig (Sus scrofa), chicken (Gallus gallus), mouse (Mus musculus), African clawed frog (Xenopus laevis), zebrafish (Danio rerio), fruit fly (Drosophila melanogaster), roundworm (Caenorhabditis elegans), and budding yeast (Saccharomyces cerevisiae). (b) Pull-down assays using GST-tagged KRAS and RIT1 or their mouse, zebrafish, or fruit fly orthologs produced recombinantly in bacteria. Recombinant proteins were incubated with lysates from HEK-293T cells expressing their corresponding species’ HA-tagged LZTR1 ortholog. Representative results from three biological replicates. Figure 1—source data 1.Raw data for Figure 1.

Fig 3: Activation of ERK1/2 pathway after cerebral I/R in diabetic rats. After I/R injury in diabetic rats, the expression and activation of RAS, ERK1/2, and Drp-1 in rat brain tissue on the 1st, 3rd, and 7th day were detected by Western blot (A), and gray values were determined by ImageJ (B). In each group (n = 5), all the results are Mean ± SD, compared with DM group, **p < 0.01. DM, diabetes mellitus.

Fig 4: CircCD44 promotes tumorigenesis and progression of TNBCs in vivo. A Tumor volume of subcutaneous xenografts with the indicated modifications; ***, P < 0.001. B Subcutaneous xenograft tumors were collected, and KRAS and C-Myc were measured using IHC; scale bar: 200 μm. C Pulmonary tissue was harvested and subjected to HE staining, and a representative image of pulmonary metastatic tumors is shown; scale bar: 200 μm. D Blood was collected from mice, and the cycling tumor cells were identified using flow cytometry. The level of CTCs was normalized to that of peripheral blood monocytes; ***, P < 0.001. E Blood was collected, and human LINE1 DNA was detected and normalized to mouse MEFs; ***, P < 0.001. Data are representative of at least 2–3 experiments with similar results

Fig 5: CircCD44 inhibits miR-502–5p-mediated KRAS degradation. A The relative KRAS RNA level was detected in different cells with the indicated modifications; ***, P < 0.001. B Immunoblot of KRAS in cells with the indicated modifications. C Schematic image of the binding site between miR-502–5p and circCD44 and the mutant allele. The binding site was obtained from the online tool, TargetScan, and the mutant allele was established as illustrated. D HEK293T cells were transfected with different plasmids or microRNAs as indicated. Relative luciferase activity was detected and normalized to the control; ***, p < 0.001. E miR-502–5p inhibitor was transfected into circCD44 knockdown cells, and miR-502–5p mimic was transfected into circCD44-overexpressing cells (these two cell lines were named “Rescue”). Immunoblot for KRAS and downstream AKT signaling components in TNBCs with the indicated modifications was detected. Data are representative of at least 2–3 experiments with similar results