Project description:ATAC-Seq of wild-type and nm1-knockout mouse embryonic fibroblasts to investigate the impact of nm1 loss on chromatin accessibility
Project description:Hi-C sequencing of wild-type and nm1-knockout mouse embryonic fibroblasts to investigate the impact of nm1 loss on genome architecture
Project description:Mouse embryonic fibroblasts were grown in DMED supplemented with 10%FBS. The same number of cells were plated in eight 6cm dishes. When the cells were ~70% confluent, four dishes received ethanol (carrier) and the other four received tamoxifen (1uM final) to induce knockout. The cells and media were harvested 4 days later. The media harvested from control and knockout dishes will be compared with the starting medium. The cells were washed with cold PBS twice while they are attached to the dishes. The whole dish with cells attached were immediately frozen in dry ice and wrapped with aluminum foil and stored at -80C.
Project description:Plasma membrane tension is an important feature that determines the cell shape and influences processes such as cell motility, spreading, endocytosis and exocytosis. Unconventional class 1 myosins are potent regulators of plasma membrane tension because they physically link the plasma membrane with the adjacent cytoskeleton. We identified nuclear myosin 1 (NM1) - a putative nuclear isoform of myosin 1c (Myo1c) - as a new player in the field. Although having specific nuclear function, we show that NM1 localizes preferentially to the plasma membrane. Deletion of NM1 causes more than 50% increase in the elasticity of the plasma membrane around the actin cytoskeleton as measured by atomic force microscopy. This higher elasticity of NM1 knock-out cells leads to 25% higher resistance to short-term hypotonic environment and rapid cell swelling. In contrary, overexpression of NM1 in WT cells leads to additional 30% reduction of their survival. We have brought evidence that NM1 has a direct functional role in the cytoplasm as a dynamic linker between the cell membrane and the underlying cytoskeleton, regulating the degree of effective plasma membrane tension. We used tissues from NM1 WT and KO mice with highest expression of NM1, lungs and heart (3 replicates each) and skin fibroblast derived from each mice (2 replicates each)