PRMT1-mediated methylation of the EGF receptor regulates signaling and cetuximab response
Hsin-Wei Liao, … , Scott Kopetz, Mien-Chie Hung
Hsin-Wei Liao, … , Scott Kopetz, Mien-Chie Hung
Published November 16, 2015
Citation Information: J Clin Invest. 2015;125(12):4529-4543. https://doi.org/10.1172/JCI82826.
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Research Article Cell biology Oncology

PRMT1-mediated methylation of the EGF receptor regulates signaling and cetuximab response

Abstract

Posttranslational modifications to the intracellular domain of the EGFR are known to regulate EGFR functions; however, modifications to the extracellular domain and their effects remain relatively unexplored. Here, we determined that methylation at R198 and R200 of the EGFR extracellular domain by protein arginine methyltransferase 1 (PRMT1) enhances binding to EGF and subsequent receptor dimerization and signaling activation. In a mouse orthotopic colorectal cancer xenograft model, expression of a methylation-defective EGFR reduced tumor growth. Moreover, increased EGFR methylation sustained signaling activation and cell proliferation in the presence of the therapeutic EGFR monoclonal antibody cetuximab. In colorectal cancer patients, EGFR methylation level also correlated with a higher recurrence rate after cetuximab treatment and reduced overall survival. Together, these data indicate that R198/R200 methylation of the EGFR plays an important role in regulating EGFR functionality and resistance to cetuximab treatment.

Authors

Hsin-Wei Liao, Jung-Mao Hsu, Weiya Xia, Hung-Ling Wang, Ying-Nai Wang, Wei-Chao Chang, Stefan T. Arold, Chao-Kai Chou, Pei-Hsiang Tsou, Hirohito Yamaguchi, Yueh-Fu Fang, Hong-Jen Lee, Heng-Huan Lee, Shyh-Kuan Tai, Mhu-Hwa Yang, Maria P. Morelli, Malabika Sen, John E. Ladbury, Chung-Hsuan Chen, Jennifer R. Grandis, Scott Kopetz, Mien-Chie Hung

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Figure 4

PRMT1 upregulates EGFR dimerization and activation, and EGFR-dependent cell proliferation through R198/200 methylation.

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PRMT1 upregulates EGFR dimerization and activation, and EGFR-dependent c...
(A) Immunoblot comparing EGFR and downstream ERK activation level of GEO cells expressing control vector, WT, and methylation-site mutant EGFR upon EGF stimulation. (B) Immunoblot comparing EGFR and downstream ERK activation level of HT29 cells expressing control vector, WT, and R198/200K mutant EGFR with or without PRMT1 knocking down upon EGF stimulation. Blots shown are representative of 5 independent experiments. (C) Cell proliferation assay of HT29 cells expressing control vector, WT, and R198/200K mutant EGFR with or without PRMT1 knocking down. (n = 6). Data are expressed as mean ± SD. (D) Left: Inactive tethered conformation of EGFR. The structure was prepared based on the crystal structure of the inactive human EGFR (PDB accession 1IVO) (http://www.rcsb.org/pdb/explore.do?structureId=1IVO). D1–D4 are color-coded; the weakly bound EGF is in cyan. R198 and R200 are highlighted. Right: Active dimerized form of EGFR (based on the crystal structure of human EGFR, PDB accession 1NQL). The second EGFR and EGF molecules are shown in gray and black, respectively. (E and F) Zoom onto R198 (E) and R200 (F) shown as stick figures. The molecular surface of EGFR (with the exception of the region 198-200) is colored in blue, positively charged atoms; red, negatively charged atoms; green, hydrophobic atoms; salmon, polar oxygens; marine, polar nitrogens; yellow, sulfur). EGFR domains are indicated on the surfaces, as is the location of D206. (G) Dimerization assay of SKCO1 cells exogenously expressing vector control and PRMT1. Anti-EGFR antibody was used to detect EGFR monomer and dimer. (H) Dimerization assay of SKCO1 cells expressing vector control and PRMT1 shRNA. (I) Dimerization assay of GEO cells expressing WT and R198/200K mutant EGFR with or without knocking down of PRMT1. Blots shown are representative of 4 independent experiments.