Fig. 1.

Yeast two-hybrid analysis of NR1–yotiao interaction. A, C1 exon cassette is necessary for an interaction between yotiao and the NR1 C-terminal tail. NR1 C-terminal tail deletion constructs were subcloned into the bait vector pBHA and assayed by yeast two-hybrid for binding to GAL4 activation domain constructs of yotiao, α-actinin-2, or CaM. Vector only, pGAD10 with no insert. Interaction was determined by activation of the two reporter genes, HIS3 and β-gal.Inset, Membrane topology of NR1. The intracellular tail consists of the membrane proximal segment C0, and alternatively spliced exons C1 and C2. B, A 272 amino acid region of yotiao is sufficient for interaction with the NR1 C-terminal tail in a C1-dependent manner. Clone A1.7 and further deletion constructs of yotiao were tested (as GAL4-activation-domain fusions) by yeast two-hybrid for binding to NR1 C-terminal deletion constructs as inA. Numbers refer to the amino acid residues at the boundaries of each construct. The location of the polypeptide fragment encoded by clone A1.7 within the full-length yotiao protein is shown.