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VEGFR-2 conformational switch in response to ligand binding.

Sarabipour S, Ballmer-Hofer K, Hristova K - Elife (2016)

Bottom Line: Ligand binding leads to a change in the TM domain conformation, resulting in increased kinase domain phosphorylation.Inter-receptor contacts within the extracellular and TM domains are critical for the establishment of the unliganded dimer structure, and for the transition to the ligand-bound active conformation.We further show that the pathogenic C482R VEGFR-2 mutant, linked to infantile hemangioma, promotes ligand-independent signaling by mimicking the structure of the ligand-bound wild-type VEGFR-2 dimer.

View Article: PubMed Central - PubMed

Affiliation: Department of Materials Science and Engineering, Johns Hopkins University, Baltimore, United States.

ABSTRACT
VEGFR-2 is the primary regulator of angiogenesis, the development of new blood vessels from pre-existing ones. VEGFR-2 has been hypothesized to be monomeric in the absence of bound ligand, and to undergo dimerization and activation only upon ligand binding. Using quantitative FRET and biochemical analysis, we show that VEGFR-2 forms dimers also in the absence of ligand when expressed at physiological levels, and that these dimers are phosphorylated. Ligand binding leads to a change in the TM domain conformation, resulting in increased kinase domain phosphorylation. Inter-receptor contacts within the extracellular and TM domains are critical for the establishment of the unliganded dimer structure, and for the transition to the ligand-bound active conformation. We further show that the pathogenic C482R VEGFR-2 mutant, linked to infantile hemangioma, promotes ligand-independent signaling by mimicking the structure of the ligand-bound wild-type VEGFR-2 dimer.

No MeSH data available.


Related in: MedlinePlus

Dimerization curves for the wild-type VEGFR-2 in the absence of ligand, the D4→D5 mutant in the absence of ligand, and the D4→D5 mutant in the presence of VEGF-A121.DOI:http://dx.doi.org/10.7554/eLife.13876.014
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fig5s1: Dimerization curves for the wild-type VEGFR-2 in the absence of ligand, the D4→D5 mutant in the absence of ligand, and the D4→D5 mutant in the presence of VEGF-A121.DOI:http://dx.doi.org/10.7554/eLife.13876.014

Mentions: A monomer-dimer equilibrium model was successfully fitted to the FRET data for the EC+TM VEGFR-2 D4(VEGFR-2)→D5(VEGFR-1) construct (Figure 5—figure supplement 1). Dimerization of the D4 mutant was stronger than for the wild type, and the mutant dimer exhibited much higher Intrinsic FRET. Thus, the substitution of the D4 domain caused a structural change in the unliganded VEGFR-2 EC domain favoring non-productive dimer formation in the absence of ligand. On the other hand, a monomer-dimer model could not be fitted to the D7 mutant data. This is indicative of the formation of higher order oligomers (King et al., 2016). Thus, the substitution of both D4 and D7 significantly altered the association of the VEGFR-2 molecules in the absence of ligand.


VEGFR-2 conformational switch in response to ligand binding.

Sarabipour S, Ballmer-Hofer K, Hristova K - Elife (2016)

Dimerization curves for the wild-type VEGFR-2 in the absence of ligand, the D4→D5 mutant in the absence of ligand, and the D4→D5 mutant in the presence of VEGF-A121.DOI:http://dx.doi.org/10.7554/eLife.13876.014
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4829425&req=5

fig5s1: Dimerization curves for the wild-type VEGFR-2 in the absence of ligand, the D4→D5 mutant in the absence of ligand, and the D4→D5 mutant in the presence of VEGF-A121.DOI:http://dx.doi.org/10.7554/eLife.13876.014
Mentions: A monomer-dimer equilibrium model was successfully fitted to the FRET data for the EC+TM VEGFR-2 D4(VEGFR-2)→D5(VEGFR-1) construct (Figure 5—figure supplement 1). Dimerization of the D4 mutant was stronger than for the wild type, and the mutant dimer exhibited much higher Intrinsic FRET. Thus, the substitution of the D4 domain caused a structural change in the unliganded VEGFR-2 EC domain favoring non-productive dimer formation in the absence of ligand. On the other hand, a monomer-dimer model could not be fitted to the D7 mutant data. This is indicative of the formation of higher order oligomers (King et al., 2016). Thus, the substitution of both D4 and D7 significantly altered the association of the VEGFR-2 molecules in the absence of ligand.

Bottom Line: Ligand binding leads to a change in the TM domain conformation, resulting in increased kinase domain phosphorylation.Inter-receptor contacts within the extracellular and TM domains are critical for the establishment of the unliganded dimer structure, and for the transition to the ligand-bound active conformation.We further show that the pathogenic C482R VEGFR-2 mutant, linked to infantile hemangioma, promotes ligand-independent signaling by mimicking the structure of the ligand-bound wild-type VEGFR-2 dimer.

View Article: PubMed Central - PubMed

Affiliation: Department of Materials Science and Engineering, Johns Hopkins University, Baltimore, United States.

ABSTRACT
VEGFR-2 is the primary regulator of angiogenesis, the development of new blood vessels from pre-existing ones. VEGFR-2 has been hypothesized to be monomeric in the absence of bound ligand, and to undergo dimerization and activation only upon ligand binding. Using quantitative FRET and biochemical analysis, we show that VEGFR-2 forms dimers also in the absence of ligand when expressed at physiological levels, and that these dimers are phosphorylated. Ligand binding leads to a change in the TM domain conformation, resulting in increased kinase domain phosphorylation. Inter-receptor contacts within the extracellular and TM domains are critical for the establishment of the unliganded dimer structure, and for the transition to the ligand-bound active conformation. We further show that the pathogenic C482R VEGFR-2 mutant, linked to infantile hemangioma, promotes ligand-independent signaling by mimicking the structure of the ligand-bound wild-type VEGFR-2 dimer.

No MeSH data available.


Related in: MedlinePlus