Complex between α-bungarotoxin and an α7 nicotinic receptor ligand-binding domain chimaera.

To identify high-affinity interactions between long-chain α-neurotoxins and nicotinic receptors, we determined the crystal structure of the complex between α-btx (α-bungarotoxin) and a pentameric ligand-binding domain constructed from the human α7 AChR (acetylcholine receptor) and AChBP (acetylcholine-binding protein). The complex buries ~2000 Ų (1 Å=0.1 nm) of surface area, within which Arg³6 and Phe³² from finger II of α-btx form a π-cation stack that aligns edge-to-face with the conserved Tyr¹84 from loop-C of α7, while Asp³° of α-btx forms a hydrogen bond with the hydroxy group of Tyr¹84. These inter-residue interactions diverge from those in a 4.2 Å structure of α-ctx (α-cobratoxin) bound to AChBP, but are similar to those in a 1.94 Å structure of α-btx bound to the monomeric α1 extracellular domain, although compared with the monomer-bound complex, the α-btx backbone exhibits a large shift relative to the protein surface. Mutational analyses show that replacing Tyr¹84 with a threonine residue abolishes high-affinity α-btx binding, whereas replacing with a phenylalanine residue maintains high affinity. Comparison of the α-btx complex with that coupled to the agonist epibatidine reveals structural rearrangements within the binding pocket and throughout each subunit. The overall findings highlight structural principles by which α-neurotoxins interact with nicotinic receptors.

Tristability in cancer-associated microRNA-TF chimera toggle switch.

Cell fate decisions during embryonic development and tumorigenesis pose a major research challenge in modern developmental and cancer biology. Binary cell fate decisions are usually regulated by gene circuits incorporating either classical toggle switches with two mutually inhibiting transcription factor (TF) genes or chimera toggle switches with a mutually inhibiting pair of microRNA (miRNA) and TF gene. These circuits can explain binary cell fate decisions. Importantly, intermediate cell types can exist during the differentiation of both stem cells and cancer cells. It has been shown that TF-TF self-activating toggle switches (SATS) can have coexistence of three metastable states (tristability), yet the role of chimera toggle switches in opening these additional states remains elusive. Here we present a generalized framework for both the TF-TF SATS and miRNA-TF chimera SATS, starting from the TF-promoter and miRNA-mRNA binding/unbinding dynamics. We show that the chimera SATSs can also have tristability. We demonstrate that the dynamics of miRNA-TF SATS is qualitatively different from that of the TF-TF SATS because the nonlinear effects of translational silencing by miRNA are distinct from those of transcriptional repression. We discuss the possible relevance of these findings to fate decisions by cancer cells.