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Date of Award
Biochemistry and Molecular Biology
Phosphorylation and de-phosphorylation of many proteins is a key regulator of cellular life. It maintains cellular activity through an array of signaling pathways like cell division, proliferation and growth. However, Overexpression or mutations by the constitutive activation of phosphorylation machinery disrupt the balance in the cell, driving the inappropriate activation or deactivation of the cellular processes it controls. FAM129B is a protein whose activity is partly maintained by phosphorylation and dephosphorylation at the six serine residues at the C-terminal. FAM129B is expressed highly in many forms of cancer and its main function is to suppress apoptosis and enhances cancer cell invasion. In this project, FAM129B phosphorylation studies are done to see how the mutation at the serine residues affects its localization to membrane in confluent HeLa cells. We demonstrated that endogenous FAM129B colocalizes with N-cadherin at the adherens junction in confluent HeLa cells. However, deletion of the six serine residues phosphorylated by MEK, prevented its localization to membrane in confluent cells. It is seen that out of the six serine residues, the two residues Ser679 and Ser683 plays an important role in FAM129B localization. The alanine mutant of Ser679Ala and Ser683Ala prevented FAM129B localization to membrane in confluent HeLa cells and the activity is restored by the phosphomimic glutamic acid mutant Ser679Asp and Ser683Asp shows membrane localization at the cell-cell adherens junction. Thus, blocking phosphorylation of Ser679 and Ser683 prevented FAM129B localization to membrane and its phosphorylation takes the FAM129B to the membrane in confluent cells. It is also studied that the phosphorylation and dephosphorylation of the other serine phosphorylation sites, Ser628, Ser633 and Ser652 has no effect on FAM129B membrane localization in confluent HeLa cell.
Thompil Somasekharan, Lakshmi, "Fam129b Phosphorylation And Its Effect On Membrane Localization In Confluent Hela Cells" (2017). Wayne State University Theses. 590.