Functional Characterization of Colon Cancer-Associated Mutations in ADAM17: Modifications in the Pro-Domain Interfere with Trafficking and Maturation

Abstract

Colorectal cancer is one of the most commonly diagnosed malignancies in the Western world and is associated with elevated expression and activity of epidermal growth factor receptors (EGF-R). The metalloproteinase ADAM17 is involved in EGF-R activation by processing EGF-R ligands from membrane-bound pro-ligands. Underlining the link between colon cancer and ADAM17, genetic intestinal cancer models in ADAM17-deficient mice show a reduced tumor burden. In this study, we characterize point mutations within the ADAM17 gene found in the tissue of colon cancer patients. In order to shed light on the role of ADAM17 in cancer development, as well as into the mechanisms that regulate maturation and cellular trafficking of ADAM17, we here perform overexpression studies of four ADAM17 variants located in the pro-, membrane-proximal- and cytoplasmic-domain of the ADAM17 protein in ADAM10/17-deficient HEK cells. Interestingly, we found a cancer-associated point mutation within the pro-domain of ADAM17 (R177C) to be most impaired in its proteolytic activity and trafficking to the cell membrane. By comparing this variant to an ADAM17 construct lacking the entire pro-domain, we discovered similar functional limitations and propose a crucial role of the pro-domain for ADAM17 maturation, cellular trafficking and thus proteolytic activity.

Differential effects on human cytochromes P450 by CRISPR/Cas9-induced genetic knockout of cytochrome P450 reductase and cytochrome b5 in HepaRG cells

“…Although there is evidence for CYB5 showing preferences for certain cytochrome P450 isozymes as well as particular reactions, we are not aware of any studies that included CYP2C8 57 . Interestingly, we found that HepaRG cells are homozygous for CYP2C8*3 , a relatively common variant reported to show altered interaction properties with POR and CYB5 in vitro 58 , 59 .…”

Strategies to Target ADAM17 in Disease: From Its Discovery to the iRhom Revolution

“…This prevents the enzyme from cleaving proteins in an unspecific manner before reaching the Golgi, where its removal occurs by action of a pro-protein furin convertase [ 48 ]. The importance of the pro-domain in the regulation of ADAM17 is demonstrated in vivo by a cancer-associated point mutation that falls within its sequence (R177C) [ 49 ]. A further analysis in vitro shows that this mutation impaired ADAM17 proteolytic activity and its tracking to the cell membrane in a similar manner as a lack of the entire pro-domain affected its maturation [ 49 ].…”

Comparison of ADAM17 gene expression level in acute lymphoblastic leukemia patients

“…ADAM17 (ADAM metallopeptidase domain 17) is a membrane protein that contains 824 amino acids and its gene is located on chromosome 2. ADAM17 is widely expressed in various tissues including brain, heart, kidney, and skeletal muscle and its expression changes during embryonic development and life in adulthood (13). ADAM17 protein can regulate cellular signaling and affect cellular behavior, but the outcome always depends on the cell substrate.…”