Alpha 4 interacts with MID1 and MID2. (A) Yeast two-hybrid analysis of the interaction between MID1 and Alpha 4 as well as MID2 and Alpha 4. Yeast agar plate (leu- trp- his-, 75 mM 3-AT) showing growth for MID1/Alpha 4 and MID2/Alpha 4 interactions as well as positive control two-hybrid combinations and no growth for negative controls. (B) Detection of full-length myc tagged-Alpha 4 when co-expressed with GFP-MID1 and GFP-MID2 fusion proteins in transiently transfected Cos1 cells. (a) GFP-MID1 fluorescence (green), (b) anti-myc antibody detecting myc-Alpha 4 localisation (red), (c) overlay of (a), (b) showing co-localisation of the myc-Alpha 4 fusion protein and GFP-MID1, with DAPI stain for DNA (blue). (d) GFP-MID2 fluorescence (green), (e) myc-Alpha 4 localisation (using same detection as b) (red), (f) overlay of (d), (e) with DAPI (blue) showing co-localisation of the myc-Alpha 4 fusion protein and GFP-MID2. (g) Detection of transiently expressed myc-Alpha 4 fusion protein in Cos1 cells, (h) overlay of (g) and DAPI stain showing cytoplasmic distribution of myc-Alpha 4 fusion protein.