File Name : supplemental figure 1 - arterial clustering.tif

Caption : figure s1. jagged1 clusters in human arterial endothelial cells. representative fluorescence microscopy images of jagged1 (green) and nuclei (blue) in haec that have been cultured under 1 pa shear stress for 24 hours. jagged1 clustering was found in haec under flow, but not in static conditions. scale bar represents 10 μm.

File Name : supplemental figure 2 - dapt clustering.tif

Caption : figure s2. jagged1 localization in gamma-secretase inhibited ecs. representative fluorescence microscopy images of jagged1 (green) and nuclei (blue) in ecs that have been treated with dapt to inhibit notch signaling activity. jagged1 clustered both in dapt and control conditions. scale bar represents 10 μm.

File Name : supplemental figure 3 - permeabilization.tif

Caption : figure s3. confocal analysis of jagged1 clustering. a-d; representative confocal sections with side view of jagged1 (green) and integrin β1 (red) in ecs. confocal sections from basal (a) to apical (d) side at 500 nm increments. imaging direction is shown by arrows in (a). scale bar represents 2.5 μm 

File Name : supplemental figure 4 - dapt pcr.tif

Caption : figure s4. gene expression of ecs with and without dapt under flow and static conditions. inhibition of notch activity did not affect the shear stress response of endothelial cells. similar relative responses to shear stress are found in controls and notch activity inhibited cells. gene expression levels under static conditions (black bars) are normalized to 1. significance is indicated as * = p<0.05, ** = p<0.01, *** = p<0.001. error bars are defined as the standard deviation.

File Name : supplemental figure 5 - arterial pcr.tif

Caption : figure s5. notch gene expression profile in human arterial endothelial cells in response to shear stress. gene expression profile of haec under shear stress. notch1 showed no change. notch3, dll4 and hes1 were downregulated, jag1 and hey1 were upregulated. significance is indicated as * = p<0.05, ** = p<0.01,  *** = p<0.001. error bars are defined as the standard deviation.

File Name : supplemental figure 6 - mg132.tif

Caption : figure s6. no jagged1 clustering after proteasome inhibition. representative fluorescence microscopy images of jagged1 (green) and nuclei (blue) in huvec cultured statically that have been treated with the proteasome inhibitor mg132. no clustering could be observed in treated cells. scale bar represents 10 μm. 

File Name : supplemental figure 7 - kd-oe.tif

Caption : figure s7. quantification of jagged1 knockdown and overexpression. gene expression analysis of jagged1 knockdown and overexpression. knockdown resulted in a 42% reduction of jagged1 mrna. overexpression increased jagged1 mrna levels 3-fold. significance is indicated as * = p<0.05, ** = p<0.01, *** = p<0.001. error bars are defined as the standard deviation.

File Name : supplemental figure 8 - 3h flow.tif

Caption : figure s8. notch gene expression profile in huvec in response to 3 hour shear stress exposure. gene expression profile of huvec under shear stress after 3 hours. notch3, jag1, hes1 and hey1 were upregulated, dll4 was downregulated. significance is indicated as * = p<0.05, ** = p<0.01, *** = p<0.001. error bars are defined as the standard deviation.

File Name : supplemental figure 9 - 48h biotinylation.tif

Caption : figure s9. jagged1 membrane levels after flow recovery. a; immunoblot of jagged1 on streptavidin immunoprecipitates of biotin-labelled surface protein under shear stress, β-actin as loading control. b; quantification of jagged1 membrane levels. jagged1 membrane levels show a decreasing trend, not significant. error bars are defined as the standard deviation.