FIG. 2.

Lysine 445, the primary myocardin SUMO-1 target site in myocardin. (A) Schematic diagram of the conserved canonical sumoylation sequence in myocardin. (B) The K445R mutant blocked sumoylation. Lysates taken from HeLa cells transfected with wild-type (wt) myocardin or its K445R mutant in the presence or absence of SUMO-1 as indicated were stained with V5 antibody. Data are representative of three independent assays. (C and D) The K445R mutant displayed impaired activity. Ca-Actin-Luc activity was assayed for CV1 cells transfected with dosed myocardin or its mutant K445R (C) or with increasing amounts of SUMO-1 alone or in the presence of either myocardin or the K445R mutant (D). Data obtained from three separate experiments carried out in duplicate are shown as activation levels (n-fold) relative to that for the empty vector group, which was taken as 1. Cont, control.