The long term goal of this proposal is to increase our understanding of nervous system development. Such an understanding will be important for the design of treatments for nerve injury and neurological disorders. The experiments will (1) test the hypothesis that beta1 integrins, a major class of receptors for extracellular matrix (ECM), have distinct functions in growth cone motility on the ECM molecules laminin (LN) and (FN) and (2) test for a functional relationship between beta1 integrins and the beta1-binding protein, talin. The experiments will use chromophore assisted laser inactivating (CALl), a unique method that will allow inactivation of beta1 integrins in the growth cone without disrupting beta1 function elsewhere on the cell. Experimental strategy: (1) We will demonstrate that (CALI) can be used to specifically inactivate beta1 integrins. (2) We will use CALI to inactivate beta1 in the filopodia of growth cones of DRG neurons on LN and FN. Changes in filopodial motility will be recorded by time lapse microscopy and analyzed statistically for significance. (3) We will use CALI to inactivate beta1 integrins and talin simultaneously and compare the resulting phenotype to inactivation of beta1 and talin individually. This experiment will be a direct test of the functional relationship between beta1 integrin and talin, a relationship long suspected on the basis of beta1-talin binding and colocalization.
