Justin L Sparks

Publications (2)22.63 Total impact

• Article: RNase H2 roles in genome integrity revealed by unlinking its activities.

  Hyongi Chon, Justin L Sparks, Monika Rychlik, Marcin Nowotny, Peter M Burgers, Robert J Crouch, Susana M Cerritelli
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  ABSTRACT: Ribonuclease H2 (RNase H2) protects genome integrity by its dual roles of resolving transcription-related R-loops and ribonucleotides incorporated in DNA during replication. To unlink these two functions, we generated a Saccharomyces cerevisiae RNase H2 mutant that can resolve R-loops but cannot cleave single ribonucleotides in DNA. This mutant definitively correlates the 2-5 bp deletions observed in rnh201Δ strains with single rNMPs in DNA. It also establishes a connection between R-loops and Sgs1-mediated replication reinitiation at stalled forks and identifies R-loops uniquely processed by RNase H2. In mouse, deletion of any of the genes coding for RNase H2 results in embryonic lethality, and in humans, RNase H2 hypomorphic mutations cause Aicardi-Goutières syndrome (AGS), a neuroinflammatory disorder. To determine the contribution of R-loops and rNMP in DNA to the defects observed in AGS, we characterized in yeast an AGS-related mutation, which is impaired in processing both substrates, but has sufficient R-loop degradation activity to complement the defects of rnh201Δ sgs1Δ strains. However, this AGS-related mutation accumulates 2-5 bp deletions at a very similar rate as the deletion strain.
  Nucleic Acids Research 01/2013; · 8.03 Impact Factor
• Article: RNase H2-Initiated Ribonucleotide Excision Repair.

  Justin L Sparks, Hyongi Chon, Susana M Cerritelli, Thomas A Kunkel, Erik Johansson, Robert J Crouch, Peter M Burgers
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  ABSTRACT: Ribonucleotides are incorporated into DNA by the replicative DNA polymerases at frequencies of about 2 per kb, which makes them by far the most abundant form of potential DNA damage in the cell. Their removal is essential for restoring a stable intact chromosome. Here, we present a complete biochemical reconstitution of the ribonucleotide excision repair (RER) pathway with enzymes purified from Saccharomyces cerevisiae. RER is most efficient when the ribonucleotide is incised by RNase H2, and further excised by the flap endonuclease FEN1 with strand displacement synthesis carried out by DNA polymerase δ, the PCNA clamp, its loader RFC, and completed by DNA ligase I. We observed partial redundancy for several of the enzymes in this pathway. Exo1 substitutes for FEN1 and Pol ε for Pol δ with reasonable efficiency. However, RNase H1 fails to substitute for RNase H2 in the incision step of RER.
  Molecular cell 08/2012; 47(6):980-6. · 14.61 Impact Factor