(A) The indicated ZFP568 wild type (lines 2, 5 and 9) and mutant proteins in reporter luciferase assays against the mouse Igf2-P0. The mutation N-terminal to ZF1 in the linker region in human alleles compared with chimp is marked with a *. Data in panels A and F are shown as mean±SD, t-test, *p<0.05, **p<0.01, n=3.
(B) Y484 and L490 of ZF4 in mZFP568 are packed against each other via a hydrophobic force.
(C) The ZF domain of human–C1 shows much reduced DNA binding.
(D) Comparison of binding affinities by mZFP568, cZFP568 and human-C1 against their respective Igf2-P0 sequences.
(E) Rhesus macaque and human placenta RNA-seq tracks at the Igf2 locus. The most abundant junctions are shown below the signal tracks.
(F) Relative luciferase activity of Igf2-P0 reporters cloned from indicated species transfected into 293T cells.