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. 2007 Mar 19;104(13):5413–5418. doi: 10.1073/pnas.0700809104

Fig. 2.

Fig. 2.

c-Cbl E3 ligase activity is not associated with proteolysis of PLCγ1. (A) PAE cells either expressing wild-type CKR alone or coexpressing c-Cbl were treated with CHX (20 μg/ml) and CSF-1 for indicated times and whole cell lysates were immunoblotted with an anti-PLCγ1 antibody. (B) PAE and CKR/PAE cells expressing either a control siRNA (CKR/Scrambled siRNA) or a siRNA targeting c-Cbl (CKR/c-Cbl siRNA) were unstimulated (−) or stimulated (+) with CSF-1 for indicated periods of time, and whole cell lysates were immunoblotted with an anti-PLCγ1 antibody. (C) A parallel immunoblot of whole cell lysate aliquots was probed with an anti-c-Cbl antibody. (D) PAE cells expressing constitutively active PLCγ1 (HA-Palm-PLCγ1) were preincubated for 2 h with either DMSO or MG-132 (50 μM) followed by a 30-min preincubation with CHX (20 μg/ml). Cells were then either lysed (0 min) or incubated in the continued presence of CHX for the indicated periods with or without MG-132. At each time point, whole cell lysates were immunoblotted with an anti-phospho-PLCγ1 antibody (pTyr783). (E) A parallel immunoblot of whole cell lysates was probed with an anti-HA antibody.