Characterization of the Role of Akt Kinase in Necroptosis.
Necroptosis in a novel form of programmed cell death that has been implicated in many
disease states including ischemia-reperfusion injuries, Huntington's disease,
pancreatitis, and Crohn's disease. It has also been shown to play an important role in
inflammatory diseases such as Systemic Inflammatory Response Syndrome (SIRS). Work done
in recent years has shown that necroptosis is ... read morea highly regulated process. Many signaling
proteins, most importantly RIP1 and RIP3 kinases, have been discovered to be involved in
the regulation of necroptosis. However, a complete understanding of molecular mechanisms
involved in necroptosis remains to be elucidated. The goal of this work was to
characterize the role of Akt kinase in necroptosis by gaining an understanding of how it
becomes activated in response to pro-necroptotic stimuli and elucidating the downstream
consequences of its activation in the process of cell death. We also sought to identify
novel Akt interacting proteins and to characterize the role that these proteins play in
necroptosis. In this work we show that Akt kinase activity is critical for necroptosis
in L929 cells and that it plays a key role in TNFα production. During necroptosis,
Akt is activated in a RIP1 dependent fashion through its phosphorylation specifically on
Thr308. In L929 cells, this activation requires independent signaling inputs from both
growth factors and RIP1. Akt controls necroptosis through downstream targeting of
mammalian Target of Rapamycin complex 1 (mTORC1). Akt activity, mediated in part through
mTORC1, links RIP1 to JNK activation and autocrine production of TNFα. In other
cell types, such as mouse lung fibroblasts and macrophages, Akt exhibited control over
necroptosis-associated TNFα production without contributing to cell death. We
discovered several putative Akt interacting proteins, including Arf1, Nur77, and GAPDH,
and began to describe the role that they play in cell death. Overall, our results
provide new insights into the mechanism of necroptosis and the role of Akt kinase in
both cell death and inflammatory regulation.
Thesis (Ph.D.)--Tufts University, 2013.
Submitted to the Dept. of Biochemistry.
Advisor: Alexei Degterev.
Committee: Brian Schaffausen, Peter Juo, and Philip Tsichlis.
Keywords: Biochemistry, and Cellular biology.read less