Regulation of the Tumor‐Suppressor Ikaros Transcription Factor by Vasoactive Intestinal Peptide Receptor‐1 in HuT 78 Cells

Abstract

Mutations to the germline encoding Ikaros (IK), a master regulator of lymphopoesis, result in a hyperproliferative phenotype leading to T cell leukemia. There are three identified ser/thr phosphorylation sites termed p1, p2 and p3. Hyperphosphorylation of IK at the p1 site is regulated via casein kinase II (CKII), which reduces its DNA binding affinity and mediates G1/S transition. However, other kinase consensus sites have been identified including protein kinase A (PKA). The G protein coupled receptor (GPCR) called vasoactive intestinal peptide receptor 1 (VPACR‐1) negatively regulates TCR signal transduction by invoking cAMP/PKA activity and suppresses G1/S transition. We hypothesize that VPACR‐1 induced PKA activity phosphorylates IK and impedes CKII phosphorylation, maintaining a quiescent naïve phenotype. To test our hypothesis, naïve and activated HuT 78 cells will be utilized in the presence and absence of VPACR‐1 ligand, vasoactive intestinal peptide, to determine whether different isoelectric (pI) patterns of IK are observed. Presently, 2D gel electrophoresis followed by anti‐IK immunoblot detects three isoforms exhibiting multiple pI's. VPACR‐1 induced PKA‐dependent phosphorylation of IK, which impedes cell cycle entry, will lead to a better understanding of a poorly understood anti‐proliferative GPCR regulate T cell activation. Funding: COBRE‐2P20RR015566 and 1KO1 DK064828.