Fig. 1.

Interactions of Stau1 with UPF1 and UPF2. a Schematic representation of the domain arrangements of human UPF1, UPF2, and Stau1. Globular domains are shown as rectangles and flexible linkers and low-complexity regions are denoted as lines. The helicase core of UPF1 consisting of the two RecA domains is colored yellow and the cis-inhibitory CH domain is in green. The three MIF4G domains of UPF2 are shown in shades of blue, while the C-terminal UPF1-binding region of UPF2 is shown as a hatched box. The four dsRNA-binding domains (dsRBDs) and the tubulin-binding domain (TBD) of Stau1 are in purple. The Staufen-swapping motif (SSM, hatched box) together with dsRBD5 comprises the Stau1 dimerization module (dd). The constructs of UPF1 and UPF2 used in this study are indicated. b RNA-dependent ATPase activity of UPF1 in the presence of UPF2 or Stau1, performed using an enzyme-coupled phosphate detection assay. The data points and their error bars represent the mean values and standard deviation (s.d.) from three independent experiments. c GST-pulldown assays of Stau1 and UPF2 with GST-UPF1CH as a bait. GST serves as a negative control. The top and bottom panels indicate input and precipitate in this and all other GST-pulldown experiments. Binding of Stau1 to UPF1CH is ~3-fold weaker than the binding of UPF2 to UPF1CH. d GST-pulldown assays of UPF1 and Stau1 in the absence and presence of UPF2. GST-UPF1 and UPF1CH are used as baits while GST serves as a negative control. The presence of UPF2 strengthens the interaction of Stau1 with UPF1 and UPF1CH by ~3-fold. e GST-pulldown assays to determine the Stau1-binding region of UPF2. Binding of GST-Stau1 to different UPF2 constructs spanning the entire protein identified the UPF2-MIF4G3 domain as the Stau1-binding site. The corresponding negative controls using GST as bait is shown in Supplementary Fig. 1c. The asterisk (*) indicates contaminants in d, e. The source data for this figure are provided as source data files