Elucidating the synaptic interactome of the high risk autism gene ANK2

PROJECT SUMMARY Network gene analysis suggests that high confidence autism spectrum disorder (hcASD) genes operate through convergent mechanisms that predominantly support synaptic function, which, in turn, can determine anatomical and functional brain connectivity. Several variants in ANK2 have been identified in ASD patients and this gene has been consistently considered a top hcASD risk. ANK2 encodes ankyrin-B (AnkB), which roles in axonal biology may be an underlying factor in ASD pathophysiology. However, our recent findings indicate that ankyrin- B also plays important roles at the cortical excitatory postsynapse. Moreover, both predominant AnkB isoforms in neurons (220 kDa and 440 kDa AnkB) are abundantly expressed in dendrites, where they likely perform isoform-specific roles and interact with unique partners, including proteins encoded by other hsASD genes. Thus, it is important to define the isoform-specific roles and partners of AnkB at the postsynapse. To accomplish these goal we will 1) use in vivo proximity-dependent biotin identification (iBioID) combined with in tandem mass spectrometry (MS) to capture and identify AnkB interacting partners in the postsynapse by leveraging our novel biotin ligase BirA-tagged conditional AnkB (AnkB-BioID) knock-in mouse and isoform-specific AnkB knockout mice; 2) combine live and super-resolution microscopies and cellular assays to validate compelling AnkB interactors; and 3) define the significance of novel AnkB PSD complexes for synaptic development and function.