Dynamics of GFP-Fusion p110 and p110 Isoforms of PI3K Signaling Pathway in Normal and Cancer Cells

Cancer therapeutics is a hot subject and PI3K class 1A isoforms (p110 and p110) are pursued as major targets. Genetic analysis, biochemical approaches, and structural studies have demonstrated crucial roles for these isoforms in several physiological processes. p110 is critical for insulin signaling, whereas p110 is essential for the growth and differs from p110 in many ways. Here, we have generated GFP-fusion clones of wildtype and mutant version of p110 and p110 and expressed them in HEK293 and cancer cells to examine their subcellular localization and their impact on downstream signaling. In HEK293 cells, p110 GFP-fusion protein is translocated into the nucleus, whereas p110-GFP stays exclusively in the cytoplasm. This study demonstrates that p110 and p110 oncogenecity, kinase activity, and interaction with p85 regulatory subunit does not have any impact on their subcellular localization. PI3K pathway specific inhibitor, LY294002, abrogated PI3K signaling by reducing pAkt levels, however, the subcellular localization of p110 and p110 remained unchanged. Furthermore, we analyzed the expression of recombinant p110 and p110 in a panel of human cancer cells and observed remarkable differences in their expression levels. The differential expression of recombinant p110 and p110 was observed to be mainly regulated by the endogenous levels of pAkt. Unlike in HEK293, p110 showed nuclear localization in cancer cells in a similar fashion to p110. Moreover, we observed the PI3K signaling activities in low pAkt expressing cells are mediated by PDK1 and S6K proteins. Finally, p110 and p110 were seen to play an essential role in promoting the cell cycle progression in MCF-7 and HCT-116 cells. J. Cell. Biochem. 117: 2864-2874, 2016. (c) 2016 Wiley Periodicals, Inc.