Retrotransposons are discrete segments of DNA (of viral origin) that contain all of the genetic information necessary to generate a copy of themselves and insert that copy back into the host cell DNA. The ability of retrotransposons to increase their copy number has ensured their success as invaders of host cell DNA in organisms from yeast to primates. The potentially mutagenic activity of retrotransposons is tightly suppressed in most cells, except those that divide asymmetrically, including stem cells and cells of the single-cell organism, brewer's yeast. Asymmetric cell division produces two cells with different potential, one with the capacity for self-renewal and the other with the capacity to perform specialized functions. A key requirement for asymmetric cell division is that each of these cells inherit a specific centrosome. The centrosomes comprise two intracellular bodies onto which chromosomes are anchored. Migration of the centrosomes, each with their complement of chromosomes, to opposite ends of a dividing cell ensures that chromosomes are equally distributed between the two cell products of cell division. The research outlined in this project stems from the investigagtor's discovery that stress-triggered retrotransposon activity in brewer's yeast interferes with the typical pattern of centrosome inheritance. The project will determine how the activity of retrotransposons influences the inheritance of a specific centrosome by the self-renewing cell and differentiated cell, and explores the consequences of disrupting the normal pattern of centrosome inheritance on chromosome segregation, cell division and the production of two cells with different potential. This research aims to explain how events occurring in single cells can influence the reproductive capacity and lifespan of the entire populations. The Broader Impact activities will include extensive laboratory-based training for a postdoctoral fellow, a graduate student, a high school student and also an undergraduate student to participate in the project each summer, with priority given to underrepresented minorities from institutions with limited research opportunities. The undergraduate research opportunity will focus on building self-identification as a research scientist, regardless of gender or racial representation in science.
Many types of asymmetrically dividing cells, including embryonic and germline stem cells, neural progenitors and Saccharomyces cerevisiae cells have a canonical pattern of centrosome or spindle pole body (SPB) inheritance that intrinsically regulates the orientation of the mitotic spindle, chromosome segregation and the capacity of cells to self-renew. Remarkably, these same asymmetrically dividing cell types are also permissive for the activity of retrotransposons, although host defense mechanisms usually hold retrotransposons in check to maintain the integrity of asymmetric cell division. It is not known how unleashing retrotransposon activity compromises asymmetric cell division. While examining the inherent asymmetry of nucleocapsid assembly sites of the Ty1 retrotransposon in Saccharomyces cerevisiae, the principal investigator's laboratory discovered that derepressing Ty1 activity substantially perturbs the canonical pattern of SPB inheritance. The goal of this project is to understand how Ty1 retrotransposon activity influences SPB inheritance and how the resulting alterations in SPB inheritance affect spindle orientation, chromosome segregation and the self-renewing potential of yeast cells. Protein-protein interaction assays will be used to investigate the mechanism of cross-talk between Ty1 nucleocapsid components and SPB components, particularly the coiled-coil SPB assembly protein, Spc42. Moreover, genetic and environmental manipulation of Ty1 retrotransposon activity and quantitative fluorescence microscopy-based assays of single cells will probe potential correlations between post-transcriptional steps in retrotransposition and (a) the SPB inheritance pattern, (b) the orientation of the mitotic spindle, (c) chromosome transmission fidelity, and (d) the replicative potential of mother and daughter cells. This research addresses a critical question that has not been addressed in the scientific literature previously: can sites of retrotransposon nucleocapsid assembly sequester host proteins with essential roles in SPB assembly to alter spindle orientation and the maintenance of differential cell fates by asymmetric cell division? Research to address this question will provide new insights into the relationship between retrotransposon activity and cell division.
This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.
