RNA-RNA ANNEALING PROTEINS

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The goal of this research is to isolate and characterize proteins in HeLa cell nucleus which promote base-pairing between complementary RNA molecules. During the course of experiments in which antisense RNA was used as a probe for pre-mRNA splicing, proteins present in nuclear extracts were observed to greatly accelerate the rate of RNA-RNA annealing. Experiments are outlined here to determine the mechanism and specificity of annealing and the role that these proteins play in the cell. Nuclear extracts will be fractionated by chromatographic procedures to identify and purify proteins which promote RNA/RNA annealing. Assay methods will be established for different types of annealing reactions. Purified annealing proteins will be characterized with respect to their interactions with RNA and ability to promote base-pairing. Monoclonal antibodies will be prepared against annealing proteins and used to characterize intracellular localization and association with RNA as a first step towards understanding the biological role of these proteins. This work is important for understanding properties of proteins which mediate RNA-RNA base-pairing in vivo. This research will thus provide further insight into molecular interactions which regulate gene expression in higher organisms and their viruses and enhance our ability to modulate these interactions in vivo and in vitro.