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Trafficking defects and loss of ligand binding are the underlying causes of all reported DDR2 missense mutations found in SMED-SL patients.

Ali BR, Xu H, Akawi NA, John A, Karuvantevida NS, Langer R, Al-Gazali L, Leitinger B - Hum. Mol. Genet. (2010)

Bottom Line: We found that all SMED-SL missense mutants were defective in collagen-induced receptor activation and that the three previously reported mutants (p.T713I, p.I726R and p.R752C) were retained in the endoplasmic reticulum.The novel mutant (p.E113K), in contrast, trafficked normally, like wild-type DDR2, but failed to bind collagen.Our data thus demonstrate that SMED-SL can result from at least two different loss-of-function mechanisms: namely defects in DDR2 targeting to the plasma membrane or the loss of its ligand-binding activity.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Medicine and Health Sciences, United Arab Emirates University, Al-Ain, United Arab Emirates.

ABSTRACT
Spondylo-meta-epiphyseal dysplasia (SMED) with short limbs and abnormal calcifications (SMED-SL) is a rare, autosomal recessive human growth disorder, characterized by disproportionate short stature, short limbs, short broad fingers, abnormal metaphyses and epiphyses, platyspondyly and premature calcifications. Recently, three missense mutations and one splice-site mutation in the DDR2 gene were identified as causative genetic defects for SMED-SL, but the underlying cellular and biochemical mechanisms were not explored. Here we report a novel DDR2 missense mutation, c.337G>A (p.E113K), that causes SMED-SL in two siblings in the United Arab Emirates. Another DDR2 missense mutation, c.2254C>T (p.R752C), matching one of the previously reported SMED-SL mutations, was found in a second affected family. DDR2 is a plasma membrane receptor tyrosine kinase that functions as a collagen receptor. We expressed DDR2 constructs with the identified point mutations in human cell lines and evaluated their localization and functional properties. We found that all SMED-SL missense mutants were defective in collagen-induced receptor activation and that the three previously reported mutants (p.T713I, p.I726R and p.R752C) were retained in the endoplasmic reticulum. The novel mutant (p.E113K), in contrast, trafficked normally, like wild-type DDR2, but failed to bind collagen. This finding is in agreement with our recent structural data identifying Glu113 as an important amino acid in the DDR2 ligand-binding site. Our data thus demonstrate that SMED-SL can result from at least two different loss-of-function mechanisms: namely defects in DDR2 targeting to the plasma membrane or the loss of its ligand-binding activity.

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Related in: MedlinePlus

Cartoon drawing of the DDR2 discoidin domain (selected side chains: W52, cyan; R105, blue; E113, red) bound to a collagen peptide (selected side chains: M21 and O24 of leading chain, F23 of middle chain). Hydrogen bonds involving E113 are shown as dashed black lines. The Figure was prepared using the coordinates of PDB entry 2WUH (6).
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DDQ103F8: Cartoon drawing of the DDR2 discoidin domain (selected side chains: W52, cyan; R105, blue; E113, red) bound to a collagen peptide (selected side chains: M21 and O24 of leading chain, F23 of middle chain). Hydrogen bonds involving E113 are shown as dashed black lines. The Figure was prepared using the coordinates of PDB entry 2WUH (6).

Mentions: Unlike the ER-retained mutant DDR2 receptors, E113K-DDR2 was found at the cell surface, similar to wild-type DDR2 (Fig. 3A–F), indicating a different disease-causing mechanism. Glu113 is located in the DDR2 extracellular discoidin domain. The DDR2 discoidin domain adopts a β-barrel structure consisting of eight β-strands (23). At the top of the barrel, five protruding loops create a trench that forms the collagen-binding site (6,23). Glu113, which forms a salt bridge with Arg105, is one of several key residues that contact collagen (6). Figure 8, which shows a cartoon drawing of the DDR2 discoidin domain bound to a collagen peptide, illustrates how Glu113 is involved in the DDR2-collagen interaction. Data presented in Figure 7 demonstrate that E113K-DDR2 had lost most of the collagen-binding activity. This result agrees well with the previous observation that mutation of E113Q reduced collagen-binding to ∼10% of wild-type DDR2 (23). Thus, Glu113 is one of the main DDR2 residues contributing to collagen-binding. It is also noted that Glu113 and Arg105 are conserved in DDR1, further underscoring the importance of the Glu113-Arg105 salt bridge in the DDR-collagen interaction. This is the first report on the clinical relevance of substitutions at this site.


Trafficking defects and loss of ligand binding are the underlying causes of all reported DDR2 missense mutations found in SMED-SL patients.

Ali BR, Xu H, Akawi NA, John A, Karuvantevida NS, Langer R, Al-Gazali L, Leitinger B - Hum. Mol. Genet. (2010)

Cartoon drawing of the DDR2 discoidin domain (selected side chains: W52, cyan; R105, blue; E113, red) bound to a collagen peptide (selected side chains: M21 and O24 of leading chain, F23 of middle chain). Hydrogen bonds involving E113 are shown as dashed black lines. The Figure was prepared using the coordinates of PDB entry 2WUH (6).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2865377&req=5

DDQ103F8: Cartoon drawing of the DDR2 discoidin domain (selected side chains: W52, cyan; R105, blue; E113, red) bound to a collagen peptide (selected side chains: M21 and O24 of leading chain, F23 of middle chain). Hydrogen bonds involving E113 are shown as dashed black lines. The Figure was prepared using the coordinates of PDB entry 2WUH (6).
Mentions: Unlike the ER-retained mutant DDR2 receptors, E113K-DDR2 was found at the cell surface, similar to wild-type DDR2 (Fig. 3A–F), indicating a different disease-causing mechanism. Glu113 is located in the DDR2 extracellular discoidin domain. The DDR2 discoidin domain adopts a β-barrel structure consisting of eight β-strands (23). At the top of the barrel, five protruding loops create a trench that forms the collagen-binding site (6,23). Glu113, which forms a salt bridge with Arg105, is one of several key residues that contact collagen (6). Figure 8, which shows a cartoon drawing of the DDR2 discoidin domain bound to a collagen peptide, illustrates how Glu113 is involved in the DDR2-collagen interaction. Data presented in Figure 7 demonstrate that E113K-DDR2 had lost most of the collagen-binding activity. This result agrees well with the previous observation that mutation of E113Q reduced collagen-binding to ∼10% of wild-type DDR2 (23). Thus, Glu113 is one of the main DDR2 residues contributing to collagen-binding. It is also noted that Glu113 and Arg105 are conserved in DDR1, further underscoring the importance of the Glu113-Arg105 salt bridge in the DDR-collagen interaction. This is the first report on the clinical relevance of substitutions at this site.

Bottom Line: We found that all SMED-SL missense mutants were defective in collagen-induced receptor activation and that the three previously reported mutants (p.T713I, p.I726R and p.R752C) were retained in the endoplasmic reticulum.The novel mutant (p.E113K), in contrast, trafficked normally, like wild-type DDR2, but failed to bind collagen.Our data thus demonstrate that SMED-SL can result from at least two different loss-of-function mechanisms: namely defects in DDR2 targeting to the plasma membrane or the loss of its ligand-binding activity.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Medicine and Health Sciences, United Arab Emirates University, Al-Ain, United Arab Emirates.

ABSTRACT
Spondylo-meta-epiphyseal dysplasia (SMED) with short limbs and abnormal calcifications (SMED-SL) is a rare, autosomal recessive human growth disorder, characterized by disproportionate short stature, short limbs, short broad fingers, abnormal metaphyses and epiphyses, platyspondyly and premature calcifications. Recently, three missense mutations and one splice-site mutation in the DDR2 gene were identified as causative genetic defects for SMED-SL, but the underlying cellular and biochemical mechanisms were not explored. Here we report a novel DDR2 missense mutation, c.337G>A (p.E113K), that causes SMED-SL in two siblings in the United Arab Emirates. Another DDR2 missense mutation, c.2254C>T (p.R752C), matching one of the previously reported SMED-SL mutations, was found in a second affected family. DDR2 is a plasma membrane receptor tyrosine kinase that functions as a collagen receptor. We expressed DDR2 constructs with the identified point mutations in human cell lines and evaluated their localization and functional properties. We found that all SMED-SL missense mutants were defective in collagen-induced receptor activation and that the three previously reported mutants (p.T713I, p.I726R and p.R752C) were retained in the endoplasmic reticulum. The novel mutant (p.E113K), in contrast, trafficked normally, like wild-type DDR2, but failed to bind collagen. This finding is in agreement with our recent structural data identifying Glu113 as an important amino acid in the DDR2 ligand-binding site. Our data thus demonstrate that SMED-SL can result from at least two different loss-of-function mechanisms: namely defects in DDR2 targeting to the plasma membrane or the loss of its ligand-binding activity.

Show MeSH
Related in: MedlinePlus