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Ted Cybulski 11/19/09

Ted Cybulski 11/19/09. Non-genetic origins of cell-to-cell variability in TRAIL-induced apoptosis - Spencer et al., Nature 2009. TRAIL and Cell Death. Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) activates a broad signal cascade resulting in apoptosis

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Ted Cybulski 11/19/09

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  1. Ted Cybulski 11/19/09 Non-genetic origins of cell-to-cell variability in TRAIL-induced apoptosis- Spencer et al., Nature 2009

  2. TRAIL and Cell Death • Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) activates a broad signal cascade resulting in apoptosis • Cells exhibit heterogeneous TRAIL response phenotype • Time response between TRAIL treatment varies greatly both in sensitive and resistant cells • “Fractional Killing” is a key problem for cancer theraputics Spencer et al., 2009

  3. Where does this variability come from? • Genetic/epigenetic differences, or stochastic “noise” • If caused by genetic differences, sister cells should behave identically • If caused by stochastic fluctuations, sister cells should not be correlated

  4. Correlation between sister cells decreases with time

  5. Is variance in protein levels to blame? • Did flow cytometry for expression of 5 proteins in TRAIL pathway • Ran ordinary differential simulation of TRAIL pathway • Measured differences in protein levels account for variability in Td

  6. What steps in the TRAIL pathway cause the time variance?

  7. How does pre-MOMP and apoptosis time vary? • Casp8 and Casp10 cleave BID to form a truncated BID (tBID) • tBID activates pore-forming proteins BAX and BAK • Timing of MOMP relies on the rate of tBID accumulation to a threshold set by BCL2 proteins • Can model this using the rate of IC-RP cleavage and fraction of IC-RP cleaved at MOMP

  8. Almost all variation comes from differences in tBIT accumulation rate • Td is more correlated with rate than threshold (R^2=.82 vs. R^2=.22) • Almost all variation in pre-MOMP time comes from the rate at which BID is converted to tBID

  9. Are levels of upstream proteins important to apoptosis time? • Several proteins play a role in the rate of tBID accumulation. • Modelling predicts that any single upstream protein will have minimal predictive value • Other variations in the system are too great

  10. Levels of BID have a predictive power on apoptotic death time • According to simulation, as BID levels increase, Td decreases • Similarly, Var(Td) decreases with BID levels • This is all confirmed in assays with GFP-tagged BID

  11. Conclusions • Cell-to-cell variation in TRAIL-response is transiently heritable • Variability in phenotype comes from cell-to-cell differences in protein level that exist pre-TRAIL exposure • The rate of apoptosis is correlated with the conversion of BID to tBID, but cannot be predicted by concentrations of other proteins in the pathway • Reducing cell-to-cell variability in protein concentrations could reduce problem of “fractional killing”

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