Figure 4.

LINC00173 stabilizes MGAT1 mRNA by recruiting HNRNPA2B1.(a) Through starBase, candidate RBPs that might interact with LINC00173 were selected under the condition of CLIP-Data >= 3, and potential RBPs that might interact with MGAT1 were screened out under the condition of CLIP-Data >= 6. Overlapped results were highlighted in box. (b-c) In RIP assay, the level of LINC00173 and MGAT1 precipitated by indicated antibodies was detected through RT-qPCR. (d-e) After RNA pull-down assay, western blot was performed to measure the protein level of HNRNPA2B1 pulled down by Bio-LINC00173, Bio-MGAT1-3’UTR or Bio-NC. (f) RNA pull-down assay and western blot detected the level of HNRNPA2B1 pulled down by Bio-NC or Bio-MGAT1-3’UTR before and after WT cells were transfected with sh-LINC00173-1. (g) By RT-qPCR, knockdown efficiency of sh-HNRNPA2B1-1/2/3 and overexpression efficiency of pcDNA3.1-HNRNPA2B1 were tested in HFWT and G-401 cells. (h) RT-qPCR and western blot were conducted to measure the mRNA and protein levels of MGAT1 in HFWT and G-401 cells transfected with sh-NC, sh-HNRNPA2B1-1 or sh-HNRNPA2B1-2. (i) Dual luciferase reporter assay assessed the putative affinity between HNRNPA2B1 and MGAT1 3’UTR in HFWT and G-401 cells. (j) MGAT1 mRNA stability was detected by RT-qPCR at different time points under the influence of HNRNPA2B1 knockdown, with β-actin as negative control. *P<0.05, **P<0.01.