INTRACELLULAR ACTIONS OF SPHINGOSINE-1-PHOSPHATE TO REGULATE UBIQUITINATION OF RIP1, NEMO RECRUITMENT, AND NF-kB ACTIVATION

ALVAREZ SE; MILSTIEN S; SPIEGEL S

Richmond, VA USA

Conferencia; Massey Cancer Center 2008 Research Retreat; 2008

Sphingosine 1-phosphate (S1P) is a lipid mediator that elicits multiple cellular processes, including cell migration, survival, and angiogenesis. S1P is produced inside the cell by the activation of sphingosine kinases (SphK1 and SphK2). Here, we demonstrate a critical role for SphK1 in tumor necrosis factor alpha (TNFa)-induced activation of nuclear factor (NF)-kB in melanoma cells. TNFa-induced NF-kB DNA binding activity and activation of IkB kinases (IKK) were markedly impaired by downregulation of SphK1 expression. Sphk1 is important for nuclear translocation and phosphorylation of the p65 subunit of NF-kB induced by TNFa and is recruited to lipid rafts following TNF stimulation. Interestingly, NF-kB activation was independent of S1P cell surface receptors, since neither S1P nor antagonists of these receptors affected NF-kB activation. Moreover, ubiquitination of receptor interacting protein 1 (RIP1) and NF-kB essential modulator (NEMO) by TNFa receptor associated 2 (TRAF2) E3 ubiquitin ligase, which are required for initiation of NF-kB signaling by TNFa, was greatly attenuated by downregulation of SphK1. Taken together, our data point to a unique intracellular role for SphK1 and production of S1P for regulation of ubiquitination of RIP1, NEMO recruitment, and NF-kB activation.