Fig. 3: NTF forms complexes with NOTCH3 detectable in CADASIL vessels.
From: Oligomerization, trans-reduction, and instability of mutant NOTCH3 in inherited vascular dementia
Fc-NTF co-immunoprecipitated with NOTCH3-HA (a). A version of Fc-NTF with all cysteines mutated to serine (Fc-NTF C > S) and the Fc tag alone were unable to pull down NOTCH3-HA. Experiments were performed at least three times using distinct samples with similar results. Unprocessed western blots can be found in Fig. S9. PLA was performed on paraffin-embedded postmortem CADASIL and control brains using an antibody specific for NTF (UMI-F11) and a second antibody recognizing a different epitope of NOTCH3 (1E4; Sigma Aldrich30). Positive PLA signal was identified in 4/4 CADASIL brains and 0/3 age-matched control brains. A representative leptomeningeal vessel from a CADASIL brain is displayed in (b) at ×400 and a representative leptomeningeal vessel from a control is displayed in (c) at ×400. A representative penetrating white matter vessel from a CADASIL brain is displayed in (d) at ×400 magnification and in (f) at ×1000 magnification, and a representative penetrating white matter vessel from a control vessel is displayed in (e) at ×400 magnification and in (g) at ×1000 magnification. Representative vessels from a CADASIL brain stained with UMI-F and 1E4 by immunohistochemistry are shown at ×400 in (h) and (i), respectively.
