The RhoGAP-myosin Myo9b regulates ocular lens pit morphogenesis.

BACKGROUND: During eye development the lens placode invaginates to form the lens pit. Further bending of lens epithelium and separation from ectoderm leads eventually to a spherical lens vesicle with enclosed extracellular fluid. Changes in epithelial morphology involve the actin cytoskeleton and its regulators. The myosin Myo9b is simultaneously an actin-based motor and Rho GTPase-activating protein that regulates actin cytoskeleton organization. Myo9b-deficient adult mice and embryos were analyzed for eye malformations and alterations in lens development. RESULTS: Myo9b-deficient mice showed a high incidence of microphthalmia and cataracts with occasional blepharitis. Formation of the lens vesicle during embryonic lens development was disordered in virtually all embryos. Lens placode invagination was less deep and gave rise to a conical structure instead of a spherical pit. At later stages either no lens vesicle was formed or a significantly smaller one that was not enclosed by the optic cup. Expression of the cell fate marker Pax6 was not altered. Staining of adherens junctions and F-actin was most intense at the tip of conical invaginations, suggesting that mechanical forces are not properly coordinated between epithelial cells that form the pit. CONCLUSIONS: Myo9b is a critical regulator of ocular lens vesicle morphogenesis during eye development.

Local Myo9b RhoGAP activity regulates cell motility.

To migrate, cells assume a polarized morphology, extending forward with a leading edge with their trailing edge retracting back toward the cell body. Both cell extension and retraction critically depend on the organization and dynamics of the actin cytoskeleton, and the small, monomeric GTPases Rac and Rho are important regulators of actin. Activation of Rac induces actin polymerization and cell extension, whereas activation of Rho enhances acto-myosin II contractility and cell retraction. To coordinate migration, these processes must be carefully regulated. The myosin Myo9b, a Rho GTPase-activating protein (GAP), negatively regulates Rho activity and deletion of Myo9b in leukocytes impairs cell migration through increased Rho activity. However, it is not known whether cell motility is regulated by global or local inhibition of Rho activity by Myo9b. Here, we addressed this question by using Myo9b-deficient macrophage-like cells that expressed different recombinant Myo9b constructs. We found that Myo9b accumulates in lamellipodial extensions generated by Rac-induced actin polymerization as a function of its motor activity. Deletion of Myo9b in HL-60-derived macrophages altered cell morphology and impaired cell migration. Reintroduction of Myo9b or Myo9b motor and GAP mutants revealed that local GAP activity rescues cell morphology and migration. In summary, Rac activation leads to actin polymerization and recruitment of Myo9b, which locally inhibits Rho activity to enhance directional cell migration.