Mislocalized RGCs retain cytoskeletal markers and show altered morphology in LKB1 mutants. RGC migration-related microtubule proteins were identified with antibodies to β-tubulin and doublecortin (magenta). (A–D) Representative images from E16 (A and B) and P0 (C and D) of migrating neurons positive for β-tubulin (A and C) and doublecortin (B and D). At E16, migrating RGCs in both control and LKB1^RET animals are positive for microtubule-associated proteins (arrows). At P0, β-tubulin and doublecortin are largely absent from the outer retina in controls, but both proteins remain localized to mistargeted RGC nuclei in LKB1^RET animals (arrows). Scale bars = 50 µm. (E) Representative high magnification images of migrating RGCs in control and LKB1^RET mutants show that control RGCs retain long basal processes (solid arrow), while basal processes are shorter or absent in mutant RGCs (dotted arrow). (F and G) Quantification of the percent of RGCs without a basal neurite (F), and measurements of the length of the basal neurite when it was extended (G) at E16 in controls and LKB1^RET mutants. Significantly more RGCs lacked a basal neurite in LKB1^RET mutants, and among those RGCs that possessed a basal neurite, its length was significantly shorter. (H and I) Representative single reconstructions of migrating RGCs in control and LKB1^RET mutants at E16 (H) and P0 (I). At E16, control RGCs showed apical and basal process, while in LKB1 mutants, RGCs showed variable morphologies with some retaining process (solid arrows) and some lacking processes (dotted arrows), suggesting some mutant RGCs may attempt bipolar migration. At P0 and P3, LKB1^RET RGC that remain in the outer retina take on different morphologies, consistent with alternative forms of migration. These include tear drop shapes (star) and multipolar processes extending in many directions (arrowheads). n = 3 control and n = 3 LKB1^RET animals. Scale bars = 25 µm. ***P < 0.001 and ***P < 0.01, unpaired two-tailed Student’s t test. n.s., nonsignificant.