A CD4+T cells from wild-type (WT), CBLOST and CBLBOSTmice were left unstimulated (-) or stimulated for 1 min with anti-CD3 and anti-CD4 antibodies (+). Equal amounts of cell lysates were subjected to affinity purification on Strep-Tactin-Sepharose beads and the resulting protein eluates were analyzed by immunoblotting with anti-ubiquitin (UB), anti-CBLB or anti-CBL antibodies. Also shown are a loading control corresponding to equal amounts of proteins from whole cell lysates (WCL) probed with anti-CD5, and molecular masses (kDa). TCR stimulation resulted in some CBLB degradation and in the detection of a higher molecular weight CBLB form, likely due to its ubiquitylation (marked with an asterisk). In contrast, no degradation of CBL was detectable upon TCR stimulation and its molecular weight remained unchanged. Molecular masses are shown (kDa). Data are representative of at least 2 experiments. 