Abstract
Thymocytes convert graded T cell receptor (TCR) signals into positive selection or deletion, and activation of extracellular signal-related kinase (ERK), p38, and Jun N-terminal protein kinase (JNK) mitogen-activated protein kinases (MAPKs) has been postulated to play a discriminatory role. Two families of Ras guanine nucleotide exchange factors (RasGEFs), SOS and RasGRP, activate Ras and the downstream RAF-MEK-ERK pathway. The pathways leading to lymphocyte p38 and JNK activation are less well defined. We previously described how RasGRP alone induces analog Ras-ERK activation while SOS and RasGRP cooperate to establish bimodal ERK activation. Here we employed computational modeling and biochemical experiments with model cell lines and thymocytes to show that TCR-induced ERK activation grows exponentially in thymocytes and that a W729E allosteric pocket mutant, SOS1, can only reconstitute analog ERK signaling. In agreement with RasGRP allosterically priming SOS, exponential ERK activation is severely decreased by pharmacological or genetic perturbation of the phospholipase Cγ (PLCγ)-diacylglycerol-RasGRP1 pathway. In contrast, p38 activation is not sharply thresholded and requires high-level TCR signal input. Rac and p38 activation depends on SOS1 expression but not allosteric activation. Based on computational predictions and experiments exploring whether SOS functions as a RacGEF or adaptor in Rac-p38 activation, we established that the presence of SOS1, but not its enzymatic activity, is critical for p38 activation.
SUPPLEMENTAL MATERIAL
Supplemental material for this article may be found at http://dx.doi.org/10.1128/MCB.01593-12.
ACKNOWLEDGMENTS
We thank T. Kurosaki for DT40 lines and J. Stone for RasGRP1-deficient mice. We are grateful to the members of the Chakraborty and Roose groups for helpful suggestions. Particularly, we thank Grace Maher and Mary Ho for their technical assistance.
This research was supported by a NIH Director's Pioneer award (to A.K.C.), Physical Science Oncology Center grant U54CA143874 (to A.K.C. and J.P.R.), and NIH grant 1P01AI091580-01 (to J.E.J., M.Y., H.C., A.K.C., and J.P.R.).