Fig 2: Knockdown of SRCIN1 expression partially inhibited the effects of miR-208a downregulation on the proliferation and invasion of H460 and A549 cells. H460 and A549 cells were co-transfected with an miR-208a inhibitor and SRCIN1 siRNA or NC siRNA. (A) Western blot analysis was performed to examine SRCIN1 protein expression levels in the differently treated cells 72 h following transfection. *P<0.05 vs. the NC siRNA group. #P<0.05 vs. the miR-208a inhibitor+SRCIN1 siRNA group. Cell Counting Kit-8 and Matrigel invasion assays were performed to determine (B) cell proliferation and (C) invasion, respectively, in the differently treated cells. *P<0.05 vs. the NC siRNA group. #P<0.05 vs. the miR-208a inhibitor+SRCIN1 siRNA group. miR, microRNA; SRCIN1, Src kinase signaling inhibitor 1; NC, negative control; siRNA, small interfering RNA; OD, optical density.

Fig 3: Validation of spinophilin interactions. WT or spinophilin KO striatal (STR) or olfactory tubercle (OT) lysates were immunoprecipitated with a spinophilin antibody. Lysates or immunoprecipitates were immunoblotted for spinophilin and three interacting proteins that were detected in the HA immunoprecipitates that had a decreased (SAP102) or increased (Clathrin heavy chain and SRCIN1) interaction with spinophilin in amphetamine-treated animals. Spinophilin and all associated proteins were detected in the spinophilin immunoprecipitates from WT animals, but were absent in immunoprecipitates isolated from KO animals.

Fig 4: SRC kinase signaling inhibitor 1 (SRCIN1) is a direct target of miR-510 in NSCLC. (A) The predicted miR-510 binding sites in the 3'-UTR of SRC kinase signaling inhibitor 1 (SRCIN1) by bioinformatic analysis. (B) 3'-UTR luciferase reporter assay was performed in HEK293T cells cotransfected with pGL3-SRCIN1-3'-UTR Wt or pGL3-SRCIN1-3'-UTR Mut, and miR-510 inhibitor or NC inhibitor. Luciferase activity was examined at 48 h posttransfection. (C) The mRNA expression levels of SRCIN1 were measured in A549 and H460 cells transfected with miR-510 inhibitor or NC inhibitor. (D) Western blotting analysis of SRCIN1 protein in A549 and H460 cells transfected with miR-510 inhibitor or NC inhibitor. *p < 0.05.

Fig 5: miR-208a directly targets SRCIN1 by directly binding to its 3'-UTR. (A) Putative Wt and Mut miR-208 binding sites in the 3'-UTR of SRCIN1. A luciferase reporter assay was performed to detect luciferase activity in (B) H460 and (C) A549 cells that were co-transfected with miR-208a inhibitor or NC inhibitor and pMIR-SRCIN1-Wt-3'-UTR or pMIR-SRCIN1-Mut-3'-UTR. *P<0.05 vs. the NC inhibitor group. H460 and A549 cells were transfected with a miR-208a inhibitor or NC inhibitor. RT-qPCR and western blot analysis were performed to measure (D) SRCIN1 mRNA and (E) protein expression levels in the indicated cell lines. *P<0.05 vs. the NC inhibitor group. (F) Expression levels of SRCIN1 mRNA in 52 primary NSCLC tissues and adjacent non-cancerous tissues were examined using RT-qPCR. *P<0.05 with comparisons shown by lines. (G) Spearman's correlation analysis was performed to assess the association between miR-208a and SRCIN1 mRNA in NSCLC tissues. miR, microRNA; SRCIN1, Src kinase signaling inhibitor 1; 3'-UTR, 3'untranslated region; Wt, wild type; Mut, mutant type; NC, negative control; NSCLC, non-small cell lung cancer; RT-qPCR, reverse transcription-quantitative polymerase chain reaction.