Table 1 Cell lines used for analysis.

In some experiments, additional proteins were transiently expressed as indicated by a “+” and the name of the transiently expressed protein. The HA-tagged form of N1 (N1-HA) had the HA tag after the epidermal growth factor (EGF) repeat 22, which was detectable without permeabilization, but did not interfere with activation. The fusion proteins between DTX4 and GFP or Cherry had the fluorescent protein fused to the C terminus of DTX4. N1 fused to GFP had the GFP fused to the amino acid 2097 of N1. Dll1 fused to the VSV tag had the tag on the amino acid 45, which was exposed extracellularly. See fig. S1 for diagrams of the proteins.

Cell line nameParent cellsStable expressionTransient expressionGene deletion
OP9-Dll1OP9Dll1-VSVNoneNone
U2OS-N1U2OSN1-HA+DTX4-VSV
+DTX4-GFP
+DTX4-Cherry
+DynK44A
+siDTX4
+siDTX1
+siDTX3
+siAP2
+siDyn2
+siADAM10
None
N1/DTX4-KOU2OSN1-HANoneDTX4
N1/DTX4-GFPU2OSN1-HA
DTX4-GFP
+DTX4-Cherry
+Cherry
None
DTX4-ChU2OSDTX4-CherryNone
N1-GFP/DTX4-ChU2OSN1-GFP
DTX4-Cherry
None
GFP/DTX4-ChU2OSGFP
DTX4-Cherry
None
N1/DTX4-ChU2OSN1-HA
DTX4-Cherry
None
N1/CherryU2OSN1-HA
Cherry
None
N1ΔE-GFP/DTX4-ChU2OSN1ΔE-GFP
DTX4-Cherry
None