Inhibition of DHHC20-Mediated EGFR Palmitoylation Creates a Dependence on EGFR Signaling

Abstract

SUMMARY Inappropriate activation of the receptor tyrosine kinase EGFR contributes to a variety of human malignancies. Here we show a mechanism to induce vulnerability to an existing first line treatment for EGFR driven cancers. We find that inhibiting the palmitoyltransferase DHHC20 creates a dependence on EGFR signaling for cancer cell survival. The loss of palmitoylation increases sustained EGFR signal activation and sensitizes cells to EGFR tyrosine kinase inhibition. Our work shows that the reversible modification of EGFR with palmitate “pins” the unstructured C-terminal tail to the plasma membrane; impeding EGFR activation. We identify by mass spectrometry palmitoylated cysteine residues within the C-terminal tail where mutation of the cysteine residues to alanine is sufficient to activate EGFR signaling promoting cell migration and transformation. Our results reveal that the targeting of a peripheral modulator of EGFR signaling, DHHC20, causes a loss of signal regulation and susceptibility to EGFR inhibitor induced-cell death.

The depalmitoylase APT1 directs the asymmetric partitioning of Notch and Wnt signaling during cell division

“…DHHC20 is also known to be expressed in MDA-MB-231 cells, where it palmitoylates EGFR and attenuates EGFR signaling (28). We asked whether loss of DHHC20 would affect protein localization.…”

Induced sensitivity to EGFR inhibitors is mediated by palmitoylated cysteine 1025 of EGFR and requires oncogenic Kras

“…1A) (16). Although inhibition of DHHC20 by shRNA leads to an increase in sensitivity to gefitinib-induced cell death, it is still unclear if this effect was directly mediated by inhibiting EGFR palmitoylation or if it is the effect of another unknown target of DHHC20.…”

“…Although inhibition of DHHC20 by shRNA leads to an increase in sensitivity to gefitinib-induced cell death, it is still unclear if this effect was directly mediated by inhibiting EGFR palmitoylation or if it is the effect of another unknown target of DHHC20. We previously showed that EGFR is palmitoylated on the C-terminal tail and that mutating one of the palmitoylated cysteine residues 1025 to alanine is sufficient to reduce receptor palmitoylation, induce receptor autophosphorylation, adaptor binding and downstream signaling to AKT and ERK when transiently expressed in NIH3T3 cells (16). To test if blocking EGFR palmitoylation directly increases gefitinib sensitivity of human cancer cells, we developed a conditional system for expressing wild type EGFR (EGFR WT ) or palmitoylation defective EGFR with cysteine 1025 mutated to alanine (EGFR C1025A ) in MDA-MB-231 cells with a tetracycline-inducible promoter.…”

“…DHHC20 palmitoylates the epidermal growth factor receptor (EGFR) on specific cysteine residues on the C-terminal tail, suppressing its activation. Unexpectedly, inhibition of DHHC20 in both breast and lung cancer cells increases induction of cell death in response to the EGFR inhibitor gefitinib, despite the fact the cells harbor activated Kras mutations and wild type EGFR (16). Mutations in Kras, predominantly an amino acid substitution at codon 12 or 13, lead to upregulation of the Ras/MAPK signaling and ERK activation, ultimately driving cell division.…”

EZH2 Palmitoylation Mediated by ZDHHC5 in p53-Mutant Glioma Drives Malignant Development and Progression

“…DHHC proteins and their substrates play important roles in tumorigenesis (13,14). Palmitoylation of H-and N-Ras facilitates Ras localization to the plasma membrane and is required for protein activity (15,16), and inhibiting ZDHHC20 palmitoyltransferase leads to the dependence of cancer cells on EGF receptor signaling for survival (17).…”