Figure 4

Mutations in a potential dimerization contact in ZF4 have no effect. A. Schematic of the potential ZF4-ZF4 dimerization contact identified in MBNL1 crystal structures. The RNA binding face of one ZF4 unit interacts with the opposite face of the second ZF4 unit. Mutation of Y224 and Q244 is predicted to interfere with the potential protein-protein interaction, but not with RNA binding by ZF4. B. The Vldlr minigene was transfected alone (lane 1), or with MBNL1 FL (lanes 2–7). The MBNL1 was WT (lane 7) or carried the indicated ZF4 mutations. C. The Tpm1 ΔBP DMS2 minigene was transfected alone (lane 1), or co-transfected with MS2 (lane 7), or MS2 fused to the N-terminal (aa 2–253) of MBNL1 (lanes 2–6). The MBNL-MS2 fusion proteins were WT (lane 6) or had the indicated ZF4 mutations (lanes 2–5). D. The Vldlr MS2 minigene was transfected alone (lane 1), or co-transfected with MS2 (lane 6), or MS2 fused to the N-terminal (aa 2–253) of MBNL1 (lanes 2–5). The MBNL-MS2 fusion proteins had the indicated ZF mutations (lanes 2–5). E. Anti-MS2 western blot of MBNL1-MS2 fusion proteins corresponding to lanes 1–6 of panel C.