Exploring Effects of Altered Nup153 Expression Through CRISPR/dCas9 Systems
Medicine and Health Sciences
Cardiomyopathy is a disease of the heart with unclear etiology. There are as many as 1 in 500 cases of cardiomyopathy in adults, making the disease incredibly prevalent. This disease remodels cardiac muscle tissue, ultimately leading to impaired heart function. Recent work has investigated potential causes of cardiomyopathy mediated by nuclear envelope proteins called nucleoporins (nups). In particular, studies have implicated a role for Nup153 in the development and/or progression of cardiopathology. In this study we use a CRISPR/dCas9 activation and inhibition approach to alter mRNA and protein levels of Nup153 in HeLa cells. Using lentiviral-based techniques, we successfully transfected HeLa cells with dCas9 plasmids designed to increase or decrease Nup153 expression. From this transfection step, we will purify the population of HeLa cells with selectable markers to target cells that have successfully taken up the dCas9 constructs. We will then add targeting guide RNAs (gRNAs) to alter Nup153 expression and assess those changes. After validating this approach in stably transfected HeLa cells, we will apply this strategy to human induced pluripotent stem cells (hiPSCs) that can be differentiated into hiPSC-derived cardiomyocytes. Using this pluripotent stem cell methodology, we will be able to determine the regulatory effects of Nup153 expression changes, if any, as applied to a cardiogenic paradigm.
Burdine, Ryan, "Exploring Effects of Altered Nup153 Expression Through CRISPR/dCas9 Systems" (2020). IdeaFest. 176.