Fig. 3.

ERG28 interacts with itself and NSDHL and SC4MOL but not with EBP. A: An overview of the NanoBiT® luciferase system for protein-protein interactions. 1) Fusion constructs of ERG28 with the LgBiT™ construct and proteins of interest (ERG28, NSDHL, SC4MOL, EBP, or a HaloTag) with the SmBiT™ construct are transfected into HEK293T cells. 2) If the proteins of interest interact with ERG28, the functional NanoLuciferase is formed. 3) When the NanoGlo substrate is supplied to the cells, luciferase activity can be measured and used to infer interaction. B: HEK293T cells were transfected for 24 h with ERG28 LgBiT, and the indicated SmBiT plasmid, split into 96-well plates, and luminosity was measured according to the manufacturer’s protocol. The relative luminosity is normalized to the response of the negative control pair (ERG28 LgBiT and HaloTag™-SmBiT), which was set to 1. Data are presented as mean + SEM of n = 3–4 independent experiments, each conducted in duplicate. Statistical significance from the ERG28 LgBiT:HaloTag SmBiT pairing was determined using a paired one-tailed t-test (∗P < 0.05).