The goal of this proposal is to examine the function of the Nova family of paraneoplastic neurologic disease antigens in neurons. We will use new methodology that we have developed to identify neuronal RNAs bound by Nova proteins. Specific hypotheses regarding the interaction of Nova with these RNAs will be examined in vitro and in vivo. Direct evidence for the in vivo function of Nova will be obtained with Nova null mice. PND's are autoimmune disorders that are believed to be triggered when tumor cells express proteins normally made only in neurons. This is associated with effective anti-tumor immunity, but neurologic degeneration as well. A number of groups have identified neuronal RNA binding proteins (n-RBPs) by expression vector cloning of brain libraries utilizing antiserum from PND patients. Two distinct groups of n-RBP's have emerged from this work. The Hu family of genes are human homologues of the Drosophila elav gene, and are also related to the Drosophila sxl gene, a regulator of splicing. The second, the Nova family of genes, are targeted in a disorder associated with breast cancer and brainstem/spinal cord motor dysfunction. The Nova antigen is related to hnRNP K, FMR-1 (fragile-X) and a number of proteins that regulate alternative splicing. In addition, FMR-1 and hnRNP K play cytoplasmic functions in ribosome binding and translational control. Aside from these sequence homologies, little is known regarding n-RBP function. We propose here a three-fold approach to the study of the Nova proteins. We have developed RNA selection methodology to identify target RNAs that are recognized by Nova-1. We propose to extend this methodology to identify neuronal RNAs bound by Nova proteins in vivo. We will utilize these RNAs to predict and examine the function of Nova as an RNA binding protein. These will include studies of Nova's role in the nucleus, as a possible regulator of alternative splicing, and in the cytoplasm, as a possible regulator of mRNA expression. Finally, we will examine the biologic role of Nova by generating and examining Nova null mice. These studies will contribute to our understanding of the role of Nova proteins in the development and function of the nervous system, and will clarify their role in the pathophysiology of the PND's including their association with autoimmunity and motor neuron dysfunction.
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