| Research Abstract |
Neurons have two types of highly polarized cell processes, axons and dendrites, both of which differentiate from common immature neurites. Initial neuronal polarization begins when one immature neurite becomes an axon and the remaining immature neurites then become dendrites. However, the intracellular mechanisms, which direct the processes to become an axon or dendrites, are largely unclear. Rho family GTPases regulate cytoskeletons, cell adhesions, neuronal polarity and cell migration by extra cellular signals. We have previously identified Rho-kinase, as an effector of Rho. Rho-kinase phosphorylates collapsin response mediator protein-2 (CRMP-2). CRMP-2 plays crucial roles in axonal growth and determination of axon-dendrite fate, thereby establishing and maintaining neuronal polarity. However, the molecular mechanisms by which the CRMP-2 or Rho family GTPases regulate neuronal polarity have been unknown. In this project, we have found the following results. We identified tublin, Num
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b and LARG as CRMP- 2-interacting molecules. (1) Functional analysis of CRMP-2 ; CRMP-2 binds to tubulin-heterodimer and regulates microtubule assembly. CRMP-2 is involved in polarized Numb-mediated endocytosis. (2) Effect of Rho family GTPases on neuronal polarity ; Phosphorylated CRMP-2 by Rho-kinase do not bind to tubulin. CRMP-2 negatively regulates the activity of LARG/Rho signal. Moreover, IQGAP1, an effector of Racl and Cdc42, interacts with CLIP-170. CLIP-170 binds to the growing ends of microtubules and plays pivotal roles in orientation. Rac1/Cdc42 marks special cortical spots where the IQGAP1 and CLIP-170 complex is targeted, leading to a polarized microtubule array and cell polarization. (3) Whole body functional analysis ; In C.elegans, UNC-33 is CRMP2 homologue and is expressed in neurons. We identified UNC-43 (calcuim-calmodulin dependent kinase II)-UNC-33-interacting molecule. The results obtained here lead us to better understanding how the Rho family GTPases regulate neuronal polarity. Less
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