Molecular insights into the regulatory interactions of Dystrophia myotonica protein kinase

Abstract

Dystrophia Myotonica Protein Kinase (DMPK) is the defining member of a family of complex, multidomain kinases of major biomedical relevance. These kinases are characterized by a highly conserved catalytic domain and a coiled-coil motif (CC) involved in the regulation of their activity. DMPK has been related to a progressive neuromuscular disorder known as Myotonic Dystrophy (DM), the most prevalent muscular dystrophy in adults. DM is a multisystemic disease characterized by myotonia and progressive skeletal muscle weakness. Affected patients suffer from DMPK insufficiency due to retention of mRNA transcripts in the nucleus. However, it is not currently understood how low levels of this kinase affect cellular function and its involvement in disease remains elusive. The sequestration of the CUG-binding protein (CUGBP), a putative DMPK substrate by a trinucleotide repeat extension in the 3’ non-coding region of the DMPK gene has been suggested to play a role in the disease. DMPK is speculated to be involved in the modulation of the plasma membrane depolarization and reorganization of the actin cytoskeleton during tissue development, possibly, acting as a downstream effector of the actin cytoskeletonlinked GTPase Rac1. In order to gain a better understanding of the complex mechanism of regulation of this kinase, its oligomeric state and intrasteric regulation were investigated. Self-assembly is crucial for the regulation of DMPK and related kinases. Their CC domains are thought to form dimeric arrangements and, thus, to mediate dimerization in this family of kinases. In addition, a role as intrasteric regulators has been attributed to these moieties. In the current work, the role of the CC domain of DMPK in kinase assembly and possible Rac1 docking has been analyzed using structural, biophysical and biochemical approaches. Contrary to expectations, the self-assembly of DMPK is not dictated by the association properties of its CC domain, instead, it appears driven by sequence segments flanking both Nand C-termini of the catalytic kinase fraction, leading to the formation of head-tohead dimers. Our findings support a shared pattern of assembly across DMPK, ROCKs and MRCK members of this family.