Fig. 1

miR-130a & miR-145, and TβRII mRNA in Gr-1⁺CD11b⁺ cells. a Left panels: qRT-PCR for fold changes of TβRII transcript in Gr-1⁺CD11b⁺ cells isolated from the spleen of 4T1 tumor-bearing mice (Tumor) compared to that from healthy control mice (Healthy) (n = 3). Right panel: Immunofluorescence images of TβRII (green) and DAPI (blue) in Gr-1⁺CD11b⁺ cells. scale bar=10 μm. b Stability of TβRII and GAPDH transcripts in Gr-1⁺CD11b⁺ cells derived from the spleen of 4T1 tumor-bearing mice (T) compared to those from healthy control mice (N). Total RNA was isolated at the indicated times after Actinomycin D treatment. 18sRNA was used to normalize TβRII and GAPDH expression. The experiment was performed three times with three biological replicates in each group. c Strategies to identify miRNAs targeting TβRII. d Heat map of differentially expressed miRNAs in Gr-1⁺CD11b⁺ cells from 4T1 tumor-bearing mice (Tumor) compared to the healthy control (Healthy) (n = 3). Yellow and blue colors indicate increased and decreased expression, respectively. Below: Schematic diagram showing the location of miR-130a and miR-145 consensus binding sites in 3′-UTR of TβRII transcript. e Expression of the indicated miRNAs by qRT-PCR. qRT-PCR of miR-130a and miR-145 in Gr-1⁺CD11b⁺ myeloid cells sorted from the spleen of healthy control or tumor-bearing mice, with miR-19a and miR-93 as controls. f Relative luciferase activity of HeLa cells transfected with pGL3-TβRII-3′-UTR vector and miR-16, miR-130a, or miR-145 mimics by electroporation. The miR-16 was used as a negative control. g Time course experiments for the expression of miR-130a (left), miR-145 (middle) and TβRII (right) in sorted Gr-1⁺CD11b⁺ myeloid cells as measured by qRT-PCR. Y-axis: fold changes; X-axis: days after tumor injection in the MFP (n = 3/each time point). Right panel: Spearman analysis of the correlations of miR-130a or miR-145 level with TβRII level. r: Pearson’s correlation coefficient. h qRT-PCR for expression of miR-130a (left), miR-145 (middle) and TβRII (right) in myeloid subsets CD11b⁺Ly6C⁺ and CD11b⁺Ly6G⁺ sorted from spleens of 4T1 tumor-bearing (Tumor) or healthy control mice (Healthy). The data are represented as mean±SEM, and Student’s t test was performed. *p < 0.05, **p < 0.01, ***p < 0.001