Transcriptional activity of megakaryoblastic leukemia 1 (MKL1) is repressed by SUMO modification

Megakaryoblastic leukemia 1 (MKL1) was originally identified as a gene translocated in megakaryoblastic leukemia. It has been shown that MKL1 functions as a RhoA‐regulated transcriptional coactivator of serum response factor (SRF). In order to identify a protein that regulates the function of MKL1, we performed yeast two‐hybrid screening and isolated cDNA that encodes UBC9, an E2 enzyme of small ubiquitin‐related modifier‐1 (SUMO‐1), as an MKL1‐binding protein. UBC9 was found to physically interact with MKL1 by GST pull‐down assay, and MKL1 was covalently modified with SUMO‐1 in 293T cells and in vitro reconstitution system. MKL1 sumoylation is enhanced by either serum stimulation or co‐expression of constitutively active form of RhoA. Mutational analysis showed that lysine residues at 499, 576, and 624 are the major acceptor sites for SUMO‐1. In addition, reporter gene analysis revealed that mutation of the three sumoylation sites strongly enhances the transcriptional activity of MKL1. The covalent attachment of SUMO‐1 to MKL1 by gene fusion represses MKL1‐dependent transcription in a complementary manner. Finally, mutation of the sumoylation sites of MKL1 also enhances SRF‐dependent transcription without affecting MKL1‐SRF interaction. The combined results demonstrated that MKL1 is sumoylated and this modification represses transcriptional activity of MKL1.

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