TY - JOUR
T1 - Possible function of the second RecJ-like protein in stalled replication fork repair by interacting with Hef
AU - Nagata, Mariko
AU - Ishino, Sonoko
AU - Yamagami, Takeshi
AU - Simons, Jan Robert
AU - Kanai, Tamotsu
AU - Atomi, Haruyuki
AU - Ishino, Yoshizumi
N1 - Funding Information:
This work was supported by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan [grant numbers JP21113005, JP23310152, and JP26242075 to Y. I.], and a Grant-in-Aid for Japan Society for the Promotion of Science (JSPS) Fellows [JP16J02633 to M.N.].
Publisher Copyright:
© 2017 The Author(s).
PY - 2017/12/1
Y1 - 2017/12/1
N2 - RecJ was originally identified in Escherichia coli and plays an important role in the DNA repair and recombination pathways. Thermococcus kodakarensis, a hyperthermophilic archaeon, has two RecJ-like nucleases. These proteins are designated as GAN (GINS-associated nuclease) and HAN (Hef-associated nuclease), based on the protein they interact with. GAN is probably a counterpart of Cdc45 in the eukaryotic CMG replicative helicase complex. HAN is considered mainly to function with Hef for restoration of the stalled replication fork. In this study, we characterized HAN to clarify its functions in Thermococcus cells. HAN showed single-strand specific 3′ to 5′ exonuclease activity, which was stimulated in the presence of Hef. A gene disruption analysis revealed that HAN was non-essential for viability, but the ΔganΔhan double mutant did not grow under optimal conditions at 85 °C. This deficiency was not fully recovered by introducing the mutant han gene, encoding the nuclease-deficient HAN protein, back into the genome. These results suggest that the unstable replicative helicase complex without GAN performs ineffective fork progression, and thus the stalled fork repair system including HAN becomes more important. The nuclease activity of HAN is required for the function of this protein in T. kodakarensis.
AB - RecJ was originally identified in Escherichia coli and plays an important role in the DNA repair and recombination pathways. Thermococcus kodakarensis, a hyperthermophilic archaeon, has two RecJ-like nucleases. These proteins are designated as GAN (GINS-associated nuclease) and HAN (Hef-associated nuclease), based on the protein they interact with. GAN is probably a counterpart of Cdc45 in the eukaryotic CMG replicative helicase complex. HAN is considered mainly to function with Hef for restoration of the stalled replication fork. In this study, we characterized HAN to clarify its functions in Thermococcus cells. HAN showed single-strand specific 3′ to 5′ exonuclease activity, which was stimulated in the presence of Hef. A gene disruption analysis revealed that HAN was non-essential for viability, but the ΔganΔhan double mutant did not grow under optimal conditions at 85 °C. This deficiency was not fully recovered by introducing the mutant han gene, encoding the nuclease-deficient HAN protein, back into the genome. These results suggest that the unstable replicative helicase complex without GAN performs ineffective fork progression, and thus the stalled fork repair system including HAN becomes more important. The nuclease activity of HAN is required for the function of this protein in T. kodakarensis.
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U2 - 10.1038/s41598-017-17306-0
DO - 10.1038/s41598-017-17306-0
M3 - Article
C2 - 29209094
AN - SCOPUS:85037100662
SN - 2045-2322
VL - 7
JO - Scientific reports
JF - Scientific reports
IS - 1
M1 - 16949
ER -