EBV Conference 2008 Guangzhou - Baylor College of Medicine

86 (RegID: 1921; 1922)
Ryan Takeshita
Institution: University of Colorado at Boulder
e-mail: ryantake@colorado.edu
CHARACTERIZATION OF LMP-1 BEHAVIOR IN LIVE CELLS
Ryan Takeshita, Chris Stockburger, and Jennifer Martin (Univ. of Colorado at Boulder)
Posterabstract:
Latent Membrane Protein-1, the EBV oncoprotein essential for B cell immortalization, constitutively
mimics the signaling activity of the ligand-dependent CD40 receptor. Upon binding to its ligand, CD40
homo-trimerizes and migrates to lipid rafts (membrane microdomains rich in signaling molecules) in order
to generate a functional signaling complex. Because LMP-1 and CD40 trigger similar signaling
pathways (NFκB, JAK/STAT, JNK, etc.), and based on evidence from biochemical experiments (ie:
co-immunoprecipitation and ultracentrifugation of cellular lystates in sucrose gradients), most of the
assumptions regarding LMP-1 complex formation are rooted in analogy to that of CD40. However, there
is little known about the behavior of LMP-1 in live, intact cells.
In an effort to characterize the behaviors of LMP-1 necessary for LMP-1 signaling and, in turn,
EBV-driven B cell immortalization, we utilize a variety of high-resolution confocal fluorescence
microscopy techniques in live cells (epithelial and lymphoid). We show that LMP-1 homo-oligomerizes
by Förster Resonance Energy Transfer and describe the stoichiometry and subcellular localization of these
LMP-1 clusters. We also investigate the relationship between oligomerization, lipid raft-association, and
LMP-1 signaling by measuring the effect of cholesterol depletion upon LMP-1 homo-oligomerization and
LMP-1’s ability to induce NFκB. By using live-cell imaging techniques, we can move away from
studying LMP-1 based on its mimicry of activated CD40 and towards the aspects of LMP-1 that make it
unique from CD40. This transition will assist in the creation of therapies for EBV-related malignancies
without side-effects on endogenous CD40 activity.
EBV Conference 2008 Guangzhou
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