Figure 1.

Expression of adhesion and degranulation promoting adapter protein (ADAP) in hippocampal neurons. (A) ADAP expression can be detected in the somata of primary hippocampal neurons and along their MAP2-positive dendritc and MAP2-negative/Tau-positive axonal structures. Scale bar, 100 μm. (B) Immunoblot analysis with two different ADAP antibodies confirms the expression in juvenile (postnatal day 9) and adult (postnatal day 90) hippocampus, as well as in neurally differentiated PC12 cells (after 4 days of NGF treatment). ADAP expression in murine CD3⁺ T-cells is shown for comparison. (C) The specificity of immunocytochemical ADAP labeling is demonstrated in HEK-293T cells, which do not express endogenous ADAP. Positive signals are strictly limited to those cells that have been transfected with His-tagged ADAP, whereas non-transfected cells marked by DAPI staining alone are negative for ADAP immunoreactivity. Scale bar, 100 μm. (D) In 7 days in vitro (DIV7) hippocampal neurons, SKAP-HOM is distributed in soma and proximal dendrites along with ADAP, but not in a MAP2-negative neurite (arrow). Scale bar, 100 μm. (E) Immunoprecipitation (IP) of hippocampal lysate identifies SKAP-HOM, RAPL and MST1, but not RIAM as binding partners of ADAP in neuronal tissue. WL, whole lysate.