In this study, we investigate the role of the receptor-like protein tyrosine phosphatase CD148 in T-cell activation. Overexpression of CD148 in the Jurkat T-cell line inhibited activation of the transcription factor nuclear factor of activated T cells following T-cell receptor (TCR) stimulation but not following stimulation through a heterologously expressed G protein-coupled receptor, the human muscarinic receptor subtype 1. Using a tetracycline-inducible expression system, we show that the TCR-mediated activation of both the Ras and calcium pathways was inhibited by expression of CD148 at levels that approximate those found in activated primary T cells. These effects were dependent on the phosphatase activity of CD148. Analysis of TCR-induced protein tyrosine phosphorylation demonstrated that most phosphoproteins were unaffected by CD148 expression. However, phospholipase Cgamma1 (PLCgamma1) and LAT were strikingly hypophosphorylated in CD148-expressing cells following TCR stimulation, whereas the phosphorylation levels of Slp-76 and Itk were modestly reduced. Based on these results, we propose that CD148 negatively regulates TCR signaling by interfering with the phosphorylation and function of PLCgamma1 and LAT.