Dynamic remodeling of chromatin requires acetylation and methylation of histones, frequently affecting the same lysine residue. These alternative epigenetic modifications require the coordination of enzymes, writers and erasers, mediating them such as acetylases and deacetylases. In cells in G0/G1, DNA damage induced by doxorubicin causes an increase in histone H4K16ac, a marker of chromatin relaxation. In this context, we studied the role that VRK1, a chromatin kinase activated by DNA damage, plays in this early step. VRK1 depletion or MG149, a Tip60/KAT5 inhibitor, cause a loss of H4K16ac. DNA damage induces the phosphorylation of Tip60 mediated by VRK1 in the chromatin fraction. VRK1 directly interacts with and phosphorylates Tip60. Furthermore, the phosphorylation of Tip60 induced by doxorubicin is lost by depletion of VRK1 in both ATM +/+ and ATM−/− cells. Kinase-active VRK1, but not kinase-dead VRK1, rescues Tip60 phosphorylation induced by DNA damage independently of ATM. The Tip60 phosphorylation by VRK1 is necessary for the activating acetylation of ATM, and subsequent ATM autophosphorylation, and both are lost by VRK1 depletion. These results support that the VRK1 chromatin kinase is an upstream regulator of the initial acetylation of histones, and an early step in DNA damage responses (DDR).

Background Distal motor neuropathies with a genetic origin have a heterogeneous clinical presentation with overlapping features affecting distal nerves and including spinal muscular atrophies and amyotrophic lateral sclerosis. This indicates that their genetic background is heterogeneous. Patient and methods In this work, we have identified and characterized the genetic and molecular base of a patient with a distal sensorimotor neuropathy of unknown origin. For this study, we performed whole‐exome sequencing, molecular modelling, cloning and expression of mutant gene, and biochemical and cell biology analysis of the mutant protein. Results A novel homozygous recessive mutation in the human VRK1 gene, coding for a chromatin kinase, causing a substitution (c.637T > C; p.Tyr213His) in exon 8, was detected in a patient presenting since childhood a progressive distal sensorimotor neuropathy and spinal muscular atrophy syndrome, with normal intellectual development. Molecular modelling predicted this mutant VRK1 has altered the kinase activation loop by disrupting its interaction with the C‐terminal regulatory region. The p.Y213H mutant protein has a reduced kinase activity with different substrates, including histones H3 and H2AX, proteins involved in DNA damage responses, such as p53 and 53BP1, and coilin, the scaffold for Cajal bodies. The mutant VRK1(Y213H) protein is unable to rescue the formation of Cajal bodies assembled on coilin, in the absence of wild‐type VRK1. Conclusion The VRK1(Y213H) mutant protein alters the activation loop, impairs the kinase activity of VRK1 causing a functional insufficiency that impairs the formation of Cajal bodies assembled on coilin, a protein that regulates SMN1 and Cajal body formation.

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Background and ObjectivesTo conduct a genetic and molecular functional study of a family with members affected of hereditary spastic paraplegia (HSP) of unknown origin and carrying a novel pathogenic vaccinia-related kinase 1 (VRK1) variant.MethodsWhole-exome sequencing was performed in 2 patients, and their parents diagnosed with HSP. The novel VRK1 variant was detected by whole-exome sequencing, molecularly modeled and biochemically characterized in kinase assays. Functionally, we studied the role of this VRK1 variant in DNA damage response and its effect on the assembly of Cajal bodies (CBs).ResultsWe have identified a very rare homozygous variant VRK1-D263G with a neurologic phenotype associated with HSP and moderate intellectual disability. The molecular modeling of this VRK1 variant protein predicted an alteration in the folding of a loop that interferes with the access to the kinase catalytic site. The VRK1-D263G variant is kinase inactive and does not phosphorylate histones H2AX and H3, transcription factors activating transcription factor 2 and p53, coilin needed for assembly of CBs, and p53 binding protein 1, a DNA repair protein. Functionally, this VRK1 variant protein impairs CB formation and the DNA damage response.DiscussionThis report expands the neurologic spectrum of neuromotor syndromes associated with a new and rare VRK1 variant, representing a novel pathogenic participant in complicated HSP and demonstrates that CBs and the DNA damage response are impaired in these patients.