We are proud to announce that Dr. Patricia F. Maness and colleagues have published a groundbreaking article in The Journal of Biological Chemistry titled, “Structural interactions of ankyrin B with NrCAM and β2 spectrin.” This research uses state-of-the-art AI-based molecular modeling to reveal how the 220 kDa isoform of Ankyrin B (AnkB)—a scaffold protein encoded by the autism risk gene ANK2—forms critical connections with the neuronal cell adhesion molecule NrCAM and β2-spectrin. By validating these predicted interactions with targeted mutagenesis and co-immunoprecipitation experiments, the study pinpoints how specific missense mutations linked to ASD can disrupt AnkB’s interactions and neuronal function. These findings not only deepen our understanding of neuronal connectivity and synapse development but also highlight potential molecular mechanisms contributing to ASD.
A note from the first author
In this work, we focused on Ankyrin B (AnkB) — a scaffold protein encoded by the high-confidence autism spectrum disorder (ASD) risk gene ANK2 — and investigated its molecular interactions with the cytoplasmic domain of the L1-family cell adhesion molecule NrCAM and with β2‑spectrin. These findings advance our mechanistic understanding of how AnkB links neuronal cell-adhesion molecules and the spectrin-actin cytoskeleton to regulate dendritic spine dynamics and excitatory synapse maturation.
Abstract
Ank2 is a high confidence autism spectrum disorder (ASD) gene encoding the spectrin-actin scaffold protein Ankyrin B (AnkB). The 220 kDa isoform of AnkB has multiple functions including developmental spine pruning through L1 family cell adhesion molecules (L1-CAMs) and class 3 Semaphorins on dendrites of pyramidal neurons to achieve an appropriate excitatory balance in the neocortex. Molecular modeling employing AlphaFold was used to predict the structure and interactions of AnkB with the cytoplasmic domain of Neuron-glial Related L1-CAM (NrCAM), and with β2-Spectrin. The validity of the models was assessed by analyzing protein-protein interactions by co-immunoprecipitation from HEK293 cell lysates after mutating key residues in AnkB predicted to impair these associations. Results revealed a pocket with critical residues in the AnkB membrane-binding domain that engages NrCAM at the conserved cytoplasmic motif – FIGQY. AlphaFold modeling of the AnkB/β2-Spectrin complex identified key interactions between the AnkB spectrin-binding domain and β2-Spectrin repeats 14-15. Selected ASD-linked mutations in AnkB predicted to impact binding to NrCAM or β2-Spectrin were then assayed for protein interactions. Maternally inherited ASD missense mutations AnkB A368G located in the NrCAM binding pocket and AnkB R977Q in the Zu5A subdomain disrupted associations with NrCAM and β2-Spectrin, respectively. Moreover, AnkB A368G impaired the neuronal function of 220 kDal AnkB for Semaphorin 3F-induced spine pruning in mouse cortical neuron cultures. These new findings provide structural insights into the L1-CAM/AnkB complex and the molecular basis of ASD etiology associated with AnkB missense mutations.
Citation
Chirasani, V. R., Haberman, V. A., Oldre, E. N., Webb, B. D., Pereira, E. B., Yang, W., & Maness, P. F. (2025). Structural interactions of ankyrin B with NrCAM and β2 spectrin. Journal of Biological Chemistry, 301(1), 110872.
