Ashley Bate

Date Approved


Degree Type

Open Access Senior Honors Thesis

Department or School



XEphA4 is a cellular receptor that functions to regulate cell and tissue interactions in amphibian embryos via a repulsive mechanism that involves actin cytoskeleton reorganization. Ectopic EphA4 signaling in Xenopus embryos results in a loss of cell-adhesion and rounded cell morphology, and this phenotype is consistent with EphA4 signaling in cultured A6 cells. How EphA4 achieves its effects on the actin cytoskeleton at the molecular level is largely unknown. One known step in the pathway is that EphA4 causes inhibition of the small GTPase RhoA. RhoA has many downstream effectors that cause cytoskeletal reorganization; the most recognized of these are the ROCK proteins (Rho-associated kinases). ROCK exists in two isoforms, ROCKI and ROCKII. We hypothesize that ROCK inhibition is one step in EphA4 signaling. To test our hypothesis we used ROCK inhibitors and mutants. ROCK inhibitors resulted in loss of cell-adhesion, but not change in cell shape in Xenopus embryo, and rounded morphology in A6 culture. Dominant-negative ROCKII did not result in a phenotype similar to EphA4 and constitutively-active ROCKII did not rescue Xenopus embryos from the EphA4 phenotype. Expression of dominant-negative ROCKI in A6 culture produces a phenotype similar to EphA4 and more importantly expression of constitutively-active ROCKI in Xenopus embryos resulted in partial rescue of the embryos from the effects of EphA4 signaling. These data support a role for ROCKI but not ROCKII in EphA4 signaling. However, the lack of a change in cell shape in Xenopus embryos suggests RhoA has multiple downstream effectors involved in EphA4 signaling.