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. 2013 Apr 11;50(1):149–156. doi: 10.1016/j.molcel.2013.02.024

Figure 4.

Figure 4

Coimmunoprecipitation Experiments Do Not Confirm STAT1 as a β-Arrestin1-Interacting Molecule

(A) Immunoprecipitations using anti-FLAG-tag antibody and extracts from untreated and IFNγ-treated HEK293T cells coexpressing FLAG-tagged STAT1 and β-arrestin1 (arrb1-GFP) or STAT1 (STAT1-GFP). Shown are inputs (In.) and immunoprecipitates (IP). Immunoblotting was done consecutively with anti-GFP, anti-FLAG-tag, and anti-β-arrestin1 antibodies. Molecular weight marker positions are indicated on the left, and arrowheads point to overexpressed proteins. Asterisks indicate immunoglobulin heavy chains. The intensity (arbitrary light units) of arrb1-GFP and STAT1-GFP bands, corrected for background, is reported below the middle and bottom panels where feasible.

(B) Shown are immunoprecipitations as in (A) using extracts from HEK293T cells coexpressing FLAG-tagged STAT1 and β-arrestin1 (arrb1-GFP) in the combinations indicated. Prior to antibody incubation, the membrane was stained with Ponceau S to reveal immunoglobulin heavy-chain bands, marked with (∗). In lanes 5–8, their upper and lower boundaries were marked with needle pinches, which remain visible in the immunoblots.