Appropriate T cell signaling is essential for the function of the human immune system. T cells regulate and direct the body’s response to foreign pathogens such as virus and bacteria, or threats from within in the form of cancerous cells. Aberrant T cell activation may lead to either immune deficiency or autoimmune diseases such as diabetes or rheumatoid arthritis.
The Src-family tyrosine kinase Lck is of vital importance in the intracellular T cell signaling machinery. We have targeted Lck using siRNA-mediated RNA interference and investigated signaling properties in T cells with Lck knockdown (Lck-kd). As expected, proximal signaling was reduced in Lck-kd cells, as determined by overall tyrosine phosphorylation, CD3-zeta phosphorylation and Ca2+ mobilization. Despite this, NFAT-AP-1 activation and IL-2 secretion was increased. This paradoxical result required further investigation. We found that Grb2-SOS1 was recruited to hypophosphorylated CD3-zeta in Lck-kd cells, leading to sustained Ras-Raf-1-ERK1/2 activation. Furthermore, endocytosed TCR/CD3-containing vesicles were not targeted to lysosomes, and as a result, CD3-zeta levels remained elevated for several hours after stimulation. Prolonged survival of internalized TCR/CD3-complexes may lead to sustained signaling through the Grb2-SOS1-Ras-ERK pathway, and if allowed to operate in the absence of negative feedback normally imposed by Lck, this mechanism my result in hyperresponsive signaling. Our data demonstrate a crucial role for Lck as both a positive and negative regulator of T cell activation. Alternative or aberrant mechanisms of T cell activation may be of importance in the development of immune system pathologies, such as autoimmune diseases
