CAREER: Genome Regulation and Nuclear Rheology

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0954421DahlThe sequencing of the human genome has provided a wealth of scientific information, but this information is limited since mechanisms which control gene expression are poorly understood. The role of force in regulating gene expression is an emerging and uniquely multi-disciplinary area of study. Mechanical force is important during organism development and for proper maintenance of cells and tissues. Specifically, shear stress on endothelial cells alters gene expression and changes cell behavior. There are many known mechanosensors in cells, but short-lived signaling events are not sufficient to explain both the long-time responses and permanent changes of cells. The DNA inside the nucleus, which is organized into the genome, has unique mechanical properties capable of responding proportionally to both the magnitude and duration of applied stress. Nuclear deformation may be important since placement of genes within the nucleus is correlated with their expression. This grant will test the hypothesis that the reorganization of the nucleus helps regulate gene expression in cells exposed to force. Subnuclear reorganization in endothelial cells will be monitored under dose dependent extracellular shear stress using real time particle tracking of fluorescent proteins. Nuclear structural proteins will be up- and down-regulated to determine the role of nuclear mechanics on controlling gene expression and gene movement stimulated by extracellular shear stress. The effects of chemically-stimulated angiogenesis, blood vessel formation, on subnuclear reorganization, gene expression and cellular phenotype will specifically be examined. By correlating changes in subnuclear rearrangement, gene accessibility and gene expression with changes in nuclear mechanics, the amount of force the DNA experiences in a cell under stress can be determined. The changes associated with modifications of individual nuclear structural proteins bridges the gap between nanoscale structure and microscale mechanics of the nucleus. This project aims to prove a new form of cellular mechanotransduction using a combination of particle tracking, biological manipulation, and gene analysis in cells under shear stress. While this project will answer important biological questions using novel engineering techniques and approaches, it will also show the power of combining biology with engineering.Changes in gene regulation associated with force, independent of or to enhance chemical factors, will help scientists and engineers to better control cells for a variety of technological applications including areas of tissue engineering and stem cell therapies. Working at the interface of science, engineering and medicine includes many unique ethical situations. Seminars and web-based tutorials promoting ethical conduct in the classroom and good research practices are being developed in collaboration with the Center for Applied Ethics and will be stressed throughout graduate research, undergraduate laboratory and summer research curricula.