title: Human PrimPol mutation associated with high myopia has a DNA replication defect.
abstract: PrimPol is a primase-polymerase found in humans, and other eukaryotes, involved in bypassing lesions encountered during DNA replication. PrimPol employs both translesion synthesis and repriming mechanisms to facilitate lesion bypass by the replisome. PrimPol has been reported to be a potential susceptibility gene associated with the development of myopia. Mutation of tyrosine 89 to aspartic acid (PrimPolY89D) has been identified in a number of cases of high myopia, implicating it in the aetiology of this disorder. Here, we examined whether this mutation resulted in any changes in the molecular and cellular activities associated with human PrimPol. We show that PrimPolY89D has a striking decrease in primase and polymerase activities. The hydrophobic ring of tyrosine is important for retaining wild-type extension activity. We also demonstrate that the decreased activity of PrimPolY89D is associated with reduced affinities for DNA and nucleotides, resulting in diminished catalytic efficiency. Although the structure and stability of PrimPolY89D is altered, its fidelity remains unchanged. This mutation also reduces cell viability after DNA damage and significantly slows replication fork rates in vivo. Together, these findings establish that the major DNA replication defect associated with this PrimPol mutant is likely to contribute to the onset of high myopia.
authors.affiliation: Genome Damage and Stability Centre, School of Life Sciences, University of Sussex, Brighton BN1 9RQ, UK.
authors.forename: Benjamin A
authors.initials: BA
authors.lastname: Keen
authors.affiliation: Genome Damage and Stability Centre, School of Life Sciences, University of Sussex, Brighton BN1 9RQ, UK.
authors.forename: Laura J
authors.initials: LJ
authors.lastname: Bailey
authors.affiliation: Genome Damage and Stability Centre, School of Life Sciences, University of Sussex, Brighton BN1 9RQ, UK.
authors.forename: Stanislaw K
authors.initials: SK
authors.lastname: Jozwiakowski
authors.affiliation: Genome Damage and Stability Centre, School of Life Sciences, University of Sussex, Brighton BN1 9RQ, UK a.j.doherty@sussex.ac.uk.
authors.forename: Aidan J
authors.initials: AJ
authors.lastname: Doherty
mesh_terms: D004247:DNA / Q000378:metabolism; D019915:DNA Primase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D004261:DNA Replication; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D064251:Multifunctional Enzymes / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D009216:Myopia / Q000235:genetics*; D017354:Point Mutation
chemical_list: D064251:Multifunctional Enzymes; D004247:DNA; D019915:DNA Primase; C585901:PrimPol protein, human; D004259:DNA-Directed DNA Polymerase