The Drosophila Gene CheB42a Is a Novel Modifier of Deg/ENaC Channel Function
Figure 5
CHEB42A and LLZ can form a protein complex.
A. Schematic of transfected proteins. B. Co-expression of tagged CHEB42A-HA and LLZ-GFP proteins. Immunoprecipitation with anti-HA antibody co-precipitated LLZ. “mix” indicates an experiment in which equal amounts of protein from the singly-transfected cells were mixed prior to immunoprecipitation as a control for non-specific interactions. Expression of llz produced two protein bands, the expected lower molecular mass band, plus a band of higher molecular mass. We do not know the identity of the more slowly migrating band; it might represent an LLZ-containing multiunit complex that is resistant to SDS denaturation or might represent post-translational modifications of LLZ [45]. We also attempted to detect HA-CHEB42A after immunoprecipitating LLZ-GFP, but were not successful. C. CHEB42A did not affect LLZ surface expression. COS7 cells were transfected with llz and either GFP or CheB42a. Surface expression level was estimated with biotinylation of surface proteins followed by neutravidin precipitation. To estimate total protein, protein input was directly blotted with anti-GFP antibody (right panel). We did not observe the larger LLZ' band in the surface expression study. We speculate that differences in sample processing protocols between the IP and surface expression studies could affect solubility and detection of larger protein complexes.