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. 2021 Jul 13;12(7):696. doi: 10.1038/s41419-021-03991-3

Fig. 3. miR-301a-3p mediated trastuzumab resistance by downregulating LRIG1 expression via directly targeting its 3′-UTR in ER stressed GC cells.

Fig. 3

a The bioinformatics analyses of TargrtScan7.1 and miRDB databases indicated that the 3′-UTR sequence of LRIG1 mRNA contained two binding sites of miR-301a-3p at nucleotide positions of 469-475 and 1066-1072. b The miR-301a-3p mimics, or NC and the luciferase reporter plasmids containing wide type or mutant 3′-UTR region of LRIG1 were co-transfected into NCI-N87 and MKN45 cells. Relative luciferase activity was then detected. (n = 3). c qRT-PCR analysis for the LRIG1 mRNA in NCI-N87 and MKN45 cells treated with or without 1 μM TG for 12 h followed by 40 or 400 μg/ml trastuzumab treatment for 72 h (n = 3). d Representative western blot and quantification for LRIG1 in NCI-N87 and MKN45 cells treated with or without 1 μM TG for 12 h following by 40 or 400 μg/ml trastuzumab treatment for 72 h. e The indicated proteins of NCI-N87 and MKN45 cells, which were stably transfected with miR-301a-3p or control and treated with or without 1 μM TG for 12 h were detected by western blot. f The indicated proteins of NCI-N87 and MKN45 cells, which were stably transfected with miR-301a-3p or control and treated with 1 μM TG for 12 h followed by 400 μg/ml trastuzumab treatment for 72 h were detected by western blot. Statistical analysis was performed by Student’s t-test, error bars indicate SD (*p < 0.05, **p < 0.01, ***p < 0.001).