title: Depletion of the protein kinase VRK1 disrupts nuclear envelope morphology and leads to BAF retention on mitotic chromosomes.
abstract: Barrier to autointegration factor (BAF), which is encoded by the BANF1 gene, binds with high-affinity to double-stranded DNA and LEM domain-containing proteins at the nuclear periphery. A BANF1 mutation has recently been associated with a novel human progeria syndrome, and cells from these patients have aberrant nuclear envelopes. The interactions of BAF with its DNA- and protein-binding partners are known to be regulated by phosphorylation, and previously we validated BAF as a highly efficient substrate for the VRK1 protein kinase. Here we show that depletion of VRK1 in MCF10a and MDA-MB-231 cells results in aberrant nuclear architecture. The immobile fraction of green fluorescent protein (GFP)-BAF at the nuclear envelope (NE) is elevated, suggesting that prolonged interactions of BAF with its binding partners is likely responsible for the aberrant NE architecture. Because detachment of BAF from its binding partners is associated with NE disassembly, we performed live-imaging analysis of control and VRK1-depleted cells to visualize GFP-BAF dynamics during mitosis. In the absence of VRK1, BAF does not disperse but instead remains chromosome bound from the onset of mitosis. VRK1 depletion also increases the number of anaphase bridges and multipolar spindles. Thus phosphorylation of BAF by VRK1 is essential both for normal NE architecture and proper dynamics of BAF-chromosome interactions during mitosis. These results are consistent with previous studies of the VRK/BAF signaling axis in Caenorhabditis elegans and Drosophila melanogaster and validate VRK1 as a key regulator of NE architecture and mitotic chromosome dynamics in mammalian cells.
authors.affiliation: Department of Microbiology and Molecular Genetics, Medical College of Wisconsin, Milwaukee, WI 53226.
authors.forename: Tyler P
authors.initials: TP
authors.lastname: Molitor
authors.forename: Paula
authors.initials: P
authors.lastname: Traktman
mesh_terms: D045744:Cell Line, Tumor; D002875:Chromosomes / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000172:deficiency* / Q000235:genetics; D008938:Mitosis; D009685:Nuclear Envelope / Q000378:metabolism / Q000648:ultrastructure*; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000172:deficiency* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D008941:Spindle Apparatus / Q000378:metabolism / Q000648:ultrastructure*
chemical_list: C501959:BANF1 protein, human; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D017346:Protein Serine-Threonine Kinases; C109890:VRK1 protein, human