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An X-ray crystallographic study has suggested that vertebrate discoidin domain receptors (DDRs) have a conserved Ca2+ binding site. DDR1 and DDR2 transfected in HEK293 cells were expressed mainly as 120 and 130 kDa forms, respectively, as they are sufficiently N-glycosylated. However, both of them showed the molecular weight of 110 kDa predominantly in the cells cultured with Ca2+-depleted media. DDR2-carrying D234A mutation at the conserved Ca2+-binding site expressed the 110 kDa form dominantly even in normal culture condition. DDR2 becomes 100 kDa form in glucose-depleted culture condition and its molecular weight increases up to 130 kDa with re-feeding glucose. However, in the mutant DDR2, the increase came to a halt at 110 kDa. The 110 kDa form had premature N-glycosyl carbohydrates and located predominantly within the endoplasmic reticulum. These results suggest that DDRs require Ca2+-binding to complete their N-glycan processing and generate the form targeted to cell membrane.

Cell-surface expression of the discoidin domain receptor (DDR) tyrosine kinase family in high molecular mass form was controlled sensitively by the glucose concentration through a post-translational N-glycosylation process. Cycloheximide time-course experiments revealed that the high-molecular-mass forms of DDR1 and DDR2 were significantly less stable than control receptor tyrosine kinases. Site-directed mutational analysis of the consensus N-glycosylation sites of the DDRs revealed that mutations of asparagine 213 of DDR2 and asparagine 211 of DDR1, a conserved N-glycosylation site among vertebrate DDRs, inhibited the generation of the high-molecular-mass isoform. Taken together, these results suggest a mechanism to control the activity of the DDR family by regulating its cell-surface expression. Due to low stability, the steady-state population of functional DDR proteins in the cell surface depends sensitively on its maturation process via post-translational N-glycosylation, which is controlled by the glucose supply and the presence of a conserved N-glycosylation site.

Burn scar contracture that follows the healing of deep dermal burns causes severe deformation and functional impairment. However, its current therapeutic interventions are limited with unsatisfactory outcomes. When we treated deep second-degree burns in rat skin with activin-like kinase 5 (ALK5) inhibitor A-83-01, it reduced wound contraction and enhanced the area of re-epithelialization so that the overall time for wound closing was not altered. In addition, it reduced myofibroblast population in the dermis of burn scar with a diminished deposition of its biomarker proteins such as α-SMA and collagen. Treatment of rat dermal fibroblast with A-83-01 inhibited transforming growth factor-β1 (TGF-β1)-dependent induction of α-SMA and collagen type I. Taken together, these results suggest that topical application of ALK5 inhibitor A-83-01 could be effective in preventing the contraction of burn wound without delaying the wound closure by virtue of its inhibitory activity against the TGF-β-induced increase of myofibroblast population.