Figure 9. Syntaxin 3B in the mouse retina is phosphorylated at T14.
A. Left panel (1-3): Western blot analysis of mouse retina extract using a peptide antibody directed against a phosphopeptide derived from the N-terminus of syntaxin 3B detected a band of the predicted size of syntaxin 3B (1). Addition of the immunizing phospopeptide to the western blot blocked the signal (lane 3), whereas the corresponding unphosphorylated peptide had only a weak effect on the phospo-syntaxin 3B signal (lane 2). Right panel (4-6): Retina extract was incubated for 1 hr at 370C in the presence (6) or absence (5) of alkaline phosphatase and analyzed by western blotting using the phospho-syntaxin 3 antibody. Unincubated extract was used as a control (4). Stability of syntaxin 3B protein during the incubation was confirmed by blotting with the syntaxin 3 antibody.
B. Retina sections were labeled with the phospho-syntaxin 3 antibody with or without blocking antibodies as in A. Strong labeling of the ribbon synapse containing layers (IPL and OPL) with the phospho-specific antibodies indicates that syntaxin 3B in ribbon synapses is phosphorylated at T14. Scale bar = 20μm.