Figure 5. Peripheral T cells expressing a mutant form of SLP-76 fail to transduce TCR-mediated signals despite normal development and TCR levels.

WT and SLP-76^flox→Y3F mice were treated with tam for five days. (A) No differences in the expression of CD4 and CD8 (top panels) and TCRβ (bottom panels) on splenocytes analyzed by FACS was detected. Representative of four experiments (B) Cell lysates prepared from splenocytes stimulated with anti-CD3 for the indicated times were used for western blotting to detect phospho-ZAP-70 (p-ZAP-70), phospho-LAT (p-LAT), phospho-PLCγ1 (p-PLCγ1), total PLCγ, phospho-ERK (p-ERK), and total ERK. Representative of two experiments (C) Splenocytes from SLP-76^WT (solid line), SLP-76^flox→Y3F (dotted line), SLP-76^CKO (dashed line) with anti-CD3 overnight and CD69 levels were assessed by flow cytometry on gated CD4+ T cells (top panel) and CD8+ T cells (bottom panel) Shaded areas represent staining of unstimulated cells. Representative of two experiments. (D) Thymidine incorporation was measured after overnight anti-CD3 stimulation of splenocytes from SLP-76^WT (solid line), SLP-76^CKO (dashed line), and SLP-76^flox→Y3F (dotted line) mice. Representative of two experiments.