Quantitative proteomics analysis of signalosome dynamics in primary T cells identifies the surface receptor CD6 as a Lat adaptor-independent TCR signaling hub.

T cell antigen receptor (TCR)-mediated activation of T cells requires the interaction of dozens of proteins. Here we used quantitative mass spectrometry and activated primary CD4(+) T cells from mice in which a tag for affinity purification was knocked into several genes to determine the composition and dynamics of multiprotein complexes that formed around the kinase Zap70 and the adaptors Lat and SLP-76. Most of the 112 high-confidence time-resolved protein interactions we observed were previously unknown. The surface receptor CD6 was able to initiate its own signaling pathway by recruiting SLP-76 and the guanine nucleotide-exchange factor Vav1 regardless of the presence of Lat. Our findings provide a more complete model of TCR signaling in which CD6 constitutes a signaling hub that contributes to the diversification of TCR signaling.

The lymphoid lineage-specific actin-uncapping protein Rltpr is essential for costimulation via CD28 and the development of regulatory T cells.

Although T cell activation can result from signaling via T cell antigen receptor (TCR) alone, physiological T cell responses require costimulation via the coreceptor CD28. Through the use of an N-ethyl-N-nitrosourea-mutagenesis screen, we identified a mutation in Rltpr. We found that Rltpr was a lymphoid cell-specific, actin-uncapping protein essential for costimulation via CD28 and the development of regulatory T cells. Engagement of TCR-CD28 at the immunological synapse resulted in the colocalization of CD28 with both wild-type and mutant Rltpr proteins. However, the connection between CD28 and protein kinase C-θ and Carma1, two key effectors of CD28 costimulation, was abrogated in T cells expressing mutant Rltpr, and CD28 costimulation did not occur in those cells. Our findings provide a more complete model of CD28 costimulation in which Rltpr has a key role.

A hypomorphic mutation in the Gfi1 transcriptional repressor results in a novel form of neutropenia.

Using N-ethyl-N-nitrosourea-induced mutagenesis, we established a mouse model with a novel form of neutropenia resulting from a point mutation in the transcriptional repressor Growth Factor Independence 1 (Gfi1). These mice, called Genista, had normal viability and no weight loss, in contrast to mice expressing null alleles of the Gfi1 gene. Furthermore, the Genista mutation had a very limited impact on lymphopoiesis or on T- and B-cell function. Within the bone marrow (BM), the Genista mutation resulted in a slight increase of monopoiesis and in a block of terminal granulopoiesis. This block occurred just after the metamyelocytic stage and resulted in the generation of small numbers of atypical CD11b(+) Ly-6G(int) neutrophils, the nuclear morphology of which resembled that of mature WT neutrophils. Unexpectedly, once released from the BM, these atypical neutrophils contributed to induce mild forms of autoantibody-induced arthritis and of immune complex-mediated lung alveolitis. They additionally failed to provide resistance to acute bacterial infection. Our study demonstrates that a hypomorphic mutation in the Gfi1 transcriptional repressor results in a novel form of neutropenia characterized by a split pattern of functional responses, reflecting the distinct thresholds required for eliciting neutrophil-mediated inflammatory and anti-infectious responses.