N-terminal and central domains of APC function to regulate branch number, length and angle in developing optic axonal arbors in vivo

Abstract

During formation of neuronal circuits, axons navigate long distances to reach their target locations in the brain. When axons arrive at their target tissues, in many cases, they extend collateral branches and/or terminal arbors that serve to increase the number of synaptic connections they make with target neurons. Here, we investigated how Adenomatous Polyposis Coli (APC) regulates terminal arborization of optic axons in living Xenopus laevis tadpoles. The N-terminal and central domains of APC that regulate the microtubule cytoskeleton and stability of β-catenin in the Wnt pathway, were co-expressed with GFP in individual optic axons, and their terminal arbors were then imaged in tectal midbrains of intact tadpoles. Our data show that the APCNTERM and APCβ-cat domains both decreased the mean number, and increased the mean length, of branches in optic axonal arbors relative to control arbors in vivo. Additional analysis demonstrated that expression of the APCNTERM domain increased the average bifurcation angle of branching in optic axonal arbors. However, the APCβ-cat domain did not significantly affect the mean branch angle of arbors in tecta of living tadpoles. These data suggest that APC N-terminal and central domains both modulate number and mean length of branches optic axonal arbors in a compensatory manner, but also define a specific function for the N-terminal domain of APC in regulating branch angle in optic axonal arbors in vivo. Our findings establish novel mechanisms for the multifunctional protein APC in shaping terminal arbors in the visual circuit of the developing vertebrate brain.

Cannabinoid Receptor Type 1 regulates growth cone filopodia and axon dispersion in the optic tract of Xenopus laevis tadpoles

“…We expressed the plasmid pCS2‐GFP in small numbers of RGCs in X. laevis embryos using a lipofection procedure (Dao et al, 2019; Elul et al, 2003; Holt et al, 1990; Jin et al, 2018; Ohnuma et al, 2002; Wiley et al, 2008). pCS2‐GFP (at a concentration >1 μg/μl) was combined with a 1,2‐dioleoyloxy‐3‐trimethylammonium propane chloride (DOTAP) lipofection reagent at a 1:3 weight to volume ratio.…”

“…In other experiments, we mixed a pCS2‐β‐catenin N‐terminal domain (β‐catNTERM) plasmid with pCS2‐GFP at a 1:1 weight ratio and then combined this mixture with DOTAP at a 1:3 ratio (Elul et al, 2003; Wiley et al, 2008). We injected 50–200 nl of these DNA‐DOTAP solutions into eyebud primordia of 1‐day‐old Xenopus embryos, as described previously (Dao et al, 2019; Jin et al, 2018; Wiley et al, 2008; Elul et al, 2003; Ohnuma et al, 2002; Holt et al, 1990; Figure 1a).…”

With the Permission of Microtubules: An Updated Overview on Microtubule Function During Axon Pathfinding

“…In vivo , despite initial contradictory results obtained in Drosophila , APC has been shown to play an important role in neural circuits formation. APC mutant mice exhibit gross misrouting defects in the internal capsule, posterior commissure or thalamocortical axons, and APC-deficient neurons displayed an abnormal axonal arborization and curling at the tips ( Rusan et al, 2008 ; Yokota et al, 2009 ; Jin et al, 2018 ). Besides its MT-stabilizing role at the plus-end, APC participates in the MT-based transport of mRNAs, such as those encoding β-actin, Tubb2b or the dynein complex subunit Lis1, toward the axon.…”