Hücre bölünmesi sırasında hücre yüzey düzenlenmesinin analizi

Plasma membrane is an essential compartment of the cells that contributes to a significant number of cellular events such as development, signal transduction and communication. Thus, understanding the changes in the plasma membrane is crucial to understand the regulation of these events. Cell division is one of these key processes in which the plasma membrane is physically re-arranged yet underlying biochemical regulations remains to be elucidated. Mass spectrometry based identification of the surface proteins is a powerful approach to quantify the changes in the surface proteome however runs into limitations related with the low abundance of the surface proteins. Thus, establishment of the new approaches that can improve the plasma membrane protein identification is essential. In the first part of thesis, I developed an alternative strategy to enrich the cell surface proteins. In this method, carboxylic acid groups on the proteins are targeted with a carboxyl reactive biotin reagent enabling the streptavidin based affinity capture. I used SILAC method the quantitatively compare this new approach with a well-established one, amine reactive labeling. My analysis revealed that carboxyl labeling can affectively enrich the surface proteins and can be used as an orthogonal approach to amine reactive labeling. In the second part of my thesis, I focused on two specific plasma membrane proteins; PCDH7 and PCDH1 which have mitotic surface localization. I tackled to understand the mechanism of cell cycle dependent translocation of PCDH7&1 and their roles in mitosis. I used proximity dependent biotinylation method and revealed a possible mechanism of the cell cycle dependent surface localization of PCDH7. Depleting the PCDH7 by CRISPR-Cas9 system resulted in abnormalities in the cell division implicating an important role of the PCDH7 during mitosis.