Hurst SE, Paulusma C, Dunlap J, Biggerstaff J, Dhar M
Atp10c, located on mouse chromosome 7, is a putative phospholipid translocase or flippase which encodes for a type IV P-type ATPase. Based on numerous studies in yeast and in mice, we hypothesize that ATP10C due to its flippase nature plays a role in diet-induced obesity and diabetes; mice heterozygous for Atp10c display this phenotype. For our investigation, we decided to look at Atp10c/ATP10C in order to gleam information about its biological role and/or function. In order to do this, an ATP10C-GFP plasmid construct was commercially obtained and transfected into HEK293T cells. Immunoblots with these cells revealed band at the expected molecular weight, about 165 kD. When the fusion protein was transfected into UPS-1 cells, we discovered that unlike other type IV P-type ATPases, ATP10C does not require chaperone proteins, CDC50A or 50B, for function, nor does it act on phosphatidylserine in the lipid bilayer. Next, immunolocalization with known protein markers by fluorescence microscopy was employed and we were able to show that the ATP10C-GFP fusion protein localized to the plasma membrane and appeared as a punctate pattern in the nucleus of HEK293T cells, thus strengthening its role in protein trafficking and cell signaling pathways. Funding for this research was provided by the American Diabetes Association.
Hurst SE, Paulusma C, Dunlap J, Biggerstaff J, Dhar M. 2012. Flipping out! Overexpression and localization of a novel flippase, ATP10C. FASEB Journal 26.