Nab2, Molecular Recognition of Polyadenosine RNA by a Zinc Finger Protein

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This project, which is at the interface of biology and physics, examines a very common macromolecular interaction domain, the zinc finger motif. Zinc finger proteins, which are among the most abundant proteins in eukaryotes, play critical functions in many biological processes. The researchers use a comprehensive approach to understand how an evolutionarily conserved family of zinc finger proteins including, the Saccharomyces cerevisiae Nab2 protein, which contains tandem (CCCH) zinc fingers, interacts with RNA. Nab2 is an essential yeast protein that plays critical roles in both mRNA processing and mRNA export from the nucleus to the cytoplasm. Previous studies have demonstrated that the Nab2 zinc finger domain is required for mRNA binding and the preliminary data suggests preferential binding to polyadenosine RNA. Three areas of research will be pursued: 1) determining whether the zinc finger motifs present in Nab2 and ZC3H14 do indeed confer sequence specific binding to poly(A) RNA; 2) defining how multiple zinc fingers contribute to binding specificity and/or high affinity nucleic acid binding thus providing insight into how tandem zinc fingers confer sequence-specific binding to poly(A) RNA; and 3) exploiting a zinc finger mutant of Nab2 (C437S) with documented decreased RNA binding to understand the requirement for RNA binding in vivo and to identify factors that regulate the interaction of Nab2 with mRNA transcripts. The Intellectual Merit of the research is two-fold: 1) the insights that will be gained into how a novel family of zinc finger proteins recognizes RNA; and 2) the development of biophysical methods not typically employed to study protein nucleic acid interactions. The researchers' approach employs Fluorescence Correlation Spectroscopy (FCS), biochemical approaches, and genetic studies in yeast. The studies described are the collaborative effort of the Corbett (Biochemistry Department, Emory School of Medicine) and the Berland (Physics Department, Emory College) laboratories. Thus, these studies lie at the interface of biology and physics. The Broader Impacts resulting from this research are significant contributions to training undergraduate students, graduate researchers, and postdoctoral fellows, including cross-disciplinary training through the focused interactions of trainees in physics and biology. Importantly, these studies also set the stage for the development of interdisciplinary learning in the classroom and afford enhanced opportunities for interface with the community beyond Emory. Both PI's laboratories have a long-standing history of interaction with students at all levels including those in high schools in the Atlanta area through hosting both students and teachers in the lab. The proposed studies would enhance interactions with students interested in Physics/Biophysics in and provide a strong illustration of the strength of interdisciplinary research.The goal of this research is to understand how information within the genetic material of cells is actually read and used as a blueprint to create the building blocks needed to make and maintain cells. The researchers will study the messenger molecule, RNA, that moves the information from the cell nucleus out to the cell cytoplasm where the machinery is present to actually translate the genetic information. This process, called messenger RNA export is a critical step in gene expression or reading the genetic code. The work combines biochemistry and physics to approach this important question from a new direction and also to develop methods not previously used to study this question. Much of the work includes undergraduate students who work jointly between a Biological laboratory and a Physics laboratory. This interface between Biology and Physics also allows the researchers to develop new training methods including interdisciplinary courses and laboratories.