Intellectual merit. Epithelia are an essential tissue type in the body plan of all multicellular animals where they play a vital role in development, structure and physiology. Epithelia are also the most common tissue type in the body. To establish an epithelium successfully, cells must develop "polarity"--a strong asymmetry in the distribution of cellular proteins. Many epithelial functions, from nutrient uptake to organ formation, depend on such asymmetric distributions of proteins and this fundamental property of an epithelium. Polarity is generated by groups of proteins that reorganize the cell to transport different sets of proteins to each of the main epithelial surfaces: apical, lateral and basal. During this process, specific groups of proteins are tasked with the formation of the apical surface specifically. This project will analyze the role of a group of proteins involved in steps that lead to the stabilization of proteins at the apical surface using the fruit fly (Drosophila melanogaster) as a model system. These proteins include the membrane-associated proteins annexin B9 and beta H spectrin. Preliminary data has led to the hypothesis that this annexin and spectrin form a complex that determines the level of proteins at the apical membrane, and that they do this by modulating the movement of proteins inside the cell to and from the apical domain during a process called "protein recycling". To test this hypothesis the project will quantitatively measure the influence of these two proteins on protein recycling in normal and mutant cells and tissues. Annexin B9 and beta H spectrin also modulate a major polarity regulator called Rac during this process, and this relationship is also a focus of the research. Finally, the hypothesis suggests that many many proteins are likely to be modulated in this way. Therefore, the project will also develop a way to identify all such proteins using "proteomic" techniques. Although the action of the key proteins that initiate a polarized cell phenotype are beginning to be understood, the stabilization and elaboration of the incipient apical domain are still poorly understood. Thus, the project will produce new insights into the development and maintenance of epithelial organization. This project will also help redefine the role of spectrins in general. These proteins have been depicted in every basic cell biology textbook for over two decades as static structural proteins. This research is part of a growing body of evidence that demonstrates that these proteins actually have a much wider and more dynamic role in the cell that challenges this long-held view.
Broader impacts. The broader impacts of this project will be in the area of science education. Not only will the project provide training for graduate students and postdoctoral fellows, but it will also integrate undergraduate-driven experiments with the research. Undergraduate participants will be drawn from those doing honors theses projects and also from underrepresented groups through the WISER (Women In Science and Engineering Research) and MURE (Minority Undergraduate Research Experience) programs at Penn State. In addition, lab experiments will be used for a "Reality Science" approach in a large (>500 student) introductory cell biology class, which serves several majors. For this approach a tractable research thread that is performed by an undergraduate researcher is then integrated into the curriculum (including examinations). The undergraduate's experiments are followed in class throughout the semester with regular updates on that student's progress. This will demonstrate real-life applications of techniques that are mentioned in class, but which cannot be covered in the lab sections of the course. At the same time this will impart a feeling for the nature, pace, and thought processes that underly basic research amongst students who may choose not to become professional biologists, but nevertheless will become science-literate citizens.
