Hierarchical Microtubule Organization Controls Axon Caliber and Transport and Determines Synaptic Structure and Stability

Abstract

SUMMARY The dimensions of axons and synaptic terminals determine cell-intrinsic properties of neurons; however, the cellular mechanisms selectively controlling establishment and maintenance of neuronal compartments remain poorly understood. Here, we show that two giant Drosophila Ankyrin2 isoforms, Ank2-L and Ank2-XL, and the MAP1B homolog Futsch form a membrane-associated microtubule-organizing complex that determines axonal diameter, supports axonal transport, and provides independent control of synaptic dimensions and stability. Ank2-L controls microtubule and synaptic stability upstream of Ank2-XL that selectively controls microtubule organization. Synergistically with Futsch, Ank2-XL provides three-dimensional microtubule organization and is required to establish appropriate synaptic dimensions and release properties. In axons, the Ank2-XL/Futsch complex establishes evenly spaced, grid-like microtubule organization and determines axonal diameter in the absence of neurofilaments. Reduced microtubule spacing limits anterograde transport velocities of mitochondria and synaptic vesicles. Our data identify control of microtubule architecture as a central mechanism to selectively control neuronal dimensions, functional properties, and connectivity.

A Dynamic Protein Interaction Landscape of the Human Centrosome-Cilium Interface

“…Defects in MT post-translational modifications were previously observed when CEP120 or SPICE1 were overexpressed (Comartin et al, 2013) or depleted (Figure 5B). These observations are consistent with recent reports implicating ANK2 in MT organization (Stephan et al, 2015), and reveal ANK2 as a key component of a PxI network that controls MT organization and centriole biogenesis.…”

Bilaterian Giant Ankyrins Have a Common Evolutionary Origin and Play a Conserved Role in Patterning the Axon Initial Segment

“…The distinct distribution pattern of Ank2L4 in the proximal axon is clear when compared to the general membrane marker mCD8-RFP, which was used to outline the cells (Fig 6C). This ability of part of the polypeptide encoded by the L exon to concentrate in the proximal axon suggested that Drosophila giant ankyrins may well localize to the proximal axon in addition to distal axonal regions as has been reported for motor neurons [15, 21]. …”

“…The bulk of the Drosophila XL exon has a core array of 93 highly-conserved repeats encoding 76 amino acids which has been postulated to serve as a molecular spacer, although only a fraction of the array is required for Ank2 function in maintaining axon diameter [21]. Arrays of homologous repeats are not present in vertebrate AnkGs [14], and we found repeat arrays in only a subset of the long exons (S4 Table).…”

“…In this study we focused on disruption of isoforms that include the L exon to show that these giant isoforms are required to set up an axonal diffusion barrier and to localize a K + channel marker (GFP-Shal) to the proximal axon. Because loss of L exon-containing isoforms is known to also disrupt localization and expression of XL exon-containing isoforms [16, 21], it is not currently possible for us to determine whether L, XL or both L and XL isoforms are required for the diffusion barrier and GFP-Shal localization in the proximal axon.…”

“…elegans unc-44 is required for normal axonal growth [19], and an unidentified isoform of unc-44 is also required to maintain axon identity and concentrate the microtubule organizer CRMP to the proximal axon [20]. Drosophila Ank2 contains two long exons, XL and L, and the giant isoforms they encode work together to regulate axon diameter and synapse stability through microtubules [15, 16, 21]. Interestingly, genetic deletion of Ank2 leads to ectopic branching in the proximal axon of Drosophila sensory neurons [22].…”