The tyrosine-phosphorylated GPRC5A expression is increased in NSCLC histological tissue specimens. A, HEK293T cells were co-transfected with plasmids of EGFR, GPRC5A WT, and mutants, as indicated. Two days later, the cells were treated with or without EGF. Cell lysates were harvest for Immunoblot using anti-tyrosine-phosphorylated GPRC5A (Y317-P/Y320-P) and anti-GPRC5A. B, H292G and Calu-1 cells were pretreated with DMEM containing 0% overnight, then treated with or without EGF (50 ng/ml) for 3 hour as indicated. Cell lysates were harvested for Immunoblot using anti-EGFR-P (Y1068), anti-EGFR, anti-tyrosine-phosphorylated GPRC5A or anti-GPRC5A as indicated. The representative photomicrographs of IHC staining were shown for normal lung and NSCLC tissues with ant-GPRC5A (n = 129) (C) and anti-tyrosine-phosphorylated GPRC5A (n = 150) (E). Negative (-), positive (+), (++), (+++) staining samples, and the IHC score (IS), were as indicated. Dot-plots depicting GPRC5A (D) and the tyrosine-phosphorylated (F) expression in IHC staining were shown for adjacent normal lung (AN) and NSCLC samples. The differences between normal lung and NSCLCs were statistically significant as indicated as *(P <0.01; two-sided z test).