Oligomerization of DDR1 ECD affects receptor–ligand binding

Abstract

Discoidin Domain Receptor 1 (DDR1) is a widely expressed receptor tyrosine kinase (RTK) which regulates cell differentiation, proliferation and migration and remodeling of the extracellular matrix. Collagen(s) are the only known ligand for DDR1. We have previously reported that collagen stimulation leads to oligomerization of the full length receptor. In this study we investigated the effect of oligomerization of the DDR1 extracellular domain (ECD) pre and post ligand binding. Solid phase binding assays showed that oligomers of recombinant DDR1-Fc bound more strongly to collagen compared to dimeric DDR1-Fc alone. In addition, DDR1-Fc itself could oligomerize upon in-vitro binding to collagen when examined using atomic force microscopy. Inhibition of dynamin mediated receptor endocytosis could prevent ligand induced endocytosis of DDR1-YFP in live cells. However inhibition of receptor endocytosis did not affect DDR1 oligomerization. In summary our results demonstrate that DDR1 ECD plays a crucial role in receptor oligomerization which mediates high-affinity interactions with its ligand.

Mechanical signaling through the discoidin domain receptor 1 plays a central role in tissue fibrosis

“…Recent work using cells expressing DDR1d, a kinase-deficient isoform, supports the involvement of the kinase domain in regulating receptor oligomerization, auto-phosphorylation and clustering at the cell surface [118]. We found that upon fibrillar collagen stimulation, activated DDR1 (phosphorylation at the kinase domain (Y792)) was mainly present as oligomers, which supports the notion that receptor oligomerization precedes receptor activation [13,117]. Cells expressing the kinase-depleted DDR1d and cells expressing a kinase-dead DDR1 isoform (DDR1e) formed clusters but cells that express kinase-active DDR1 (DDR1b) exhibited large increases in cluster size.…”

“…One proposed mechanism for activation involves collagen-induced DDR1 clustering [114,116] since DDR1 oligomerization is required for high affinity binding to collagen [117]. Recent work using cells expressing DDR1d, a kinase-deficient isoform, supports the involvement of the kinase domain in regulating receptor oligomerization, auto-phosphorylation and clustering at the cell surface [118].…”

Discoidin Domain Receptor 1 Mediates Myosin-Dependent Collagen Contraction

“…Work using cells expressing DDR1d, a kinase-deficient isoform, supports the involvement of the kinase domain in regulating receptor oligomerization, auto-phosphorylation, and clustering at the cell surface (Fu et al, 2014). We found that in cells binding to fibrillar collagen, activated DDR1 was mainly oligomeric, supporting the notion that receptor oligomerization precedes receptor activation (Yeung et al, 2013). Furthermore, we found that cells expressing the kinase-depleted DDR1 (DDR1d), cells expressing kinase-dead DDR1 (DDR1e), or DDR1 OE cells cultured on non-activating fibronectin substrates all formed clusters.…”

“…WT OE W W W WT WT WT WT W WT WT WT W WT W WT W WT W W WT WT WT WT WT WT T WT T T WT WT WT W WT W W WT W WT WT W W W W W WT WT W WT WT WT T T T WT WT WT WT WT W WT W W W WT WT W W W WT T T T W WT WT WT T WT T T W W W WT W W WT T WT T T T W W WT T high-affinity binding to collagen (Yeung et al, 2013). Work using cells expressing DDR1d, a kinase-deficient isoform, supports the involvement of the kinase domain in regulating receptor oligomerization, auto-phosphorylation, and clustering at the cell surface (Fu et al, 2014).…”

Discoidin domain receptors (DDRs): Potential implications in atherosclerosis

“…Unlike most RTKs that undergo dimerization upon ligand binding, DDRs appear to exist in dimers prior to interaction with the ligand (Abdulhussein et al, 2008;Mihai et al, 2009;Noordeen et al, 2006). Studies from Agarwal et al (2007) and Leitinger (2003) showed that DDR dimerization was a prerequisite for high affinity binding to collagen, and DDR oligomerization greatly enhanced their affinity to collagen (Agarwal et al, 2007;Mihai et al, 2006;Yeung et al, 2013). Collagen interacts with pre-formed DDR dimers and upon binding induces dimer oligomerization, conformational changes and autophosphorylation on multiple tyrosine residues within cytosolic JM and kinase domain of DDRs, and thus results in receptor activation.…”