Figure 4.

POLDIP3 can be sumoylated both in vitro and in vivo. (A) The SUMO consensus sequences in human POLDIP3 protein. (B) In vitro sumoylation assay. Purified recombinant proteins were used in the in vitro sumoylation assay in the presence (+) or absence (−) of ATP. The black arrow indicates the mono-sumoylated POLDIP3. The dashed red line marks the poly-sumoylated POLDIP3. (C) In vivo sumoylation assay. HEK293T cells were transfected with either empty vector (EV) or plasmids expression His-SUMO-1, or His-SUMO-2, or His-SUMO-3. Ni-NTA beads were used to pull-down the sumoylated proteins, which were then immunoblotted (IB) with antibodies as indicated on the right.