The archaeal ATPase PINA interacts with the helicase Hjm via its carboxyl terminal KH domain remodeling and processing replication fork and Holliday junction

Zhai, Binyuan; DuPrez, Kevin; Han, Xing; Yuan, Zenglin; Ahmad, Sohail; Xu, Chengpei; Gu, Lichuan; Ni, Jinfeng; Фан, Ли; Shen, Yulong

doi:10.1093/nar/gky451

Cited by 10 publications

(15 citation statements)

References 73 publications

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“…We also characterized Tbar-aLhr2 as a monomeric DNA/RNA helicase able to process DNA:RNA and RNA:RNA duplexes (Figure S4 & Figure 7, left panels). Interestingly, we highlighted that the in vitro unwinding and annealing activities of Tbar-aLhr2 differ drastically from those of Hel308, described as a DNA helicase involved in DNA repair [35,44]. Indeed, we showed that while Tbar-aLhr2 is more prone to anneal nucleic strands than to unwind them, Paby-Hel308 unwinds 3 DNA:DNA homoduplexes but does not form them from ssDNA molecules (Figure 7).…”

Section: Discussionmentioning

confidence: 93%

Phylogenetic Diversity of Lhr Proteins and Biochemical Activities of the Thermococcales aLhr2 DNA/RNA Helicase

et al. 2021

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Helicase proteins are known to use the energy of ATP to unwind nucleic acids and to remodel protein-nucleic acid complexes. They are involved in almost every aspect of DNA and RNA metabolisms and participate in numerous repair mechanisms that maintain cellular integrity. The archaeal Lhr-type proteins are SF2 helicases that are mostly uncharacterized. They have been proposed to be DNA helicases that act in DNA recombination and repair processes in Sulfolobales and Methanothermobacter. In Thermococcales, a protein annotated as an Lhr2 protein was found in the network of proteins involved in RNA metabolism. To investigate this, we performed in-depth phylogenomic analyses to report the classification and taxonomic distribution of Lhr-type proteins in Archaea, and to better understand their relationship with bacterial Lhr. Furthermore, with the goal of envisioning the role(s) of aLhr2 in Thermococcales cells, we deciphered the enzymatic activities of aLhr2 from Thermococcus barophilus (Tbar). We showed that Tbar-aLhr2 is a DNA/RNA helicase with a significant annealing activity that is involved in processes dependent on DNA and RNA transactions.

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Section: Discussionmentioning

confidence: 93%

Phylogenetic Diversity of Lhr Proteins and Biochemical Activities of the Thermococcales aLhr2 DNA/RNA Helicase

et al. 2021

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“…The damages at the stalled replication forks may be repaired by translesion DNA synthesis, BER, MMR, or HR in different phases of cell cycle (Jones and Petermann, 2012). We have recently proposed a role of Hjc in repair of stalled replication fork in concert with Hjm and PINA (Zhai et al, 2018). The inhibition of Hjc activity by phosphorylation at S34 might avoid HJ cleavage and stalled replication fork collapse.…”

Section: Discussionmentioning

confidence: 99%

“…DNA replication fork is usually blocked by a lesion. The stalled replication fork would be reversed to a HJ DNA by proteins such as Hjm and/or PINA (PIN domain-containing ATPase), followed by the recruitment of Hjc (Zhai et al, 2018). Phosphorylation states of Hjc might control different repair pathways: Non-phosphorylated Hjc would cleave the HJ forming a DSB which would be repaired by HR; phosphorylated Hjc at S34 would not cleave the HJ and the lesion might be repaired by mismatch repair (MMR) or base excision repair (BER).…”

Section: Discussionmentioning

confidence: 99%

Phosphorylation of the Archaeal Holliday Junction Resolvase Hjc Inhibits Its Catalytic Activity and Facilitates DNA Repair in Sulfolobus islandicus REY15A

et al. 2019

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Protein phosphorylation is one of the main protein post-translational modifications and regulates DNA repair in eukaryotes. Archaeal genomes encode eukaryotic-like DNA repair proteins and protein kinases (ePKs), and several proteins involved in homologous recombination repair (HRR) including Hjc, a conserved Holliday junction (HJ) resolvase in Archaea, undergo phosphorylation, indicating that phosphorylation plays important roles in HRR. Herein, we performed phosphorylation analysis of Hjc by various ePKs from Sulfolobus islandicus . It was shown that SiRe_0171, SiRe_2030, and SiRe_2056, were able to phosphorylate Hjc in vitro . These ePKs phosphorylated Hjc at different Ser/Thr residues: SiRe_0171 on S34, SiRe_2030 on both S9 and T138, and SiRe_2056 on T138. The HJ cleavage activity of the phosphorylation-mimic mutants was analyzed and the results showed that the cleavage activity of S34E was completely lost and that of S9E had greatly reduced. S. islandicus strain expressing S34E in replacement of the wild type Hjc was resistant to higher doses of DNA damaging agents. Furthermore, SiRe_0171 deletion mutant exhibited higher sensitivity to DNA damaging agents, suggesting that Hjc phosphorylation by SiRe_0171 enhanced the DNA repair capability. Our results revealed that HJ resolvase is regulated by protein phosphorylation, reminiscent of the regulation of eukaryotic HJ resolvases GEN1 and Yen1.

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“…On the other hand, different KH domains in the same protein may have different sequence preferences and affinity, such as the four KH domains in KSRP ( 17 ) and the 14 domains in vigilin ( 18 ). In addition to nucleic acid binding, the KH domain in PINA mediates the interactions of PINA with Hjm and Hjc and regulates the hexameric assembly of PINA ( 19 ).…”

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confidence: 99%

The KH domain facilitates the substrate specificity and unwinding processivity of DDX43 helicase

Yadav

Singh

Hogan

et al. 2021

Journal of Biological Chemistry

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The K-homology (KH) domain is a nucleic acid binding domain present in many proteins. Recently we found that the DEAD-box helicase DDX43 contains a KH domain in its N-terminus; however, its function remains unknown. Here, we purified recombinant DDX43 KH domain protein and found that it prefers binding single-stranded (ss)DNA and ssRNA. Electrophoretic mobility shift assay (EMSA) and nuclear magnetic resonance (NMR) revealed that the KH domain favors pyrimidines over purines. Mutational analysis showed that the GXXG-loop in the KH domain is involved in pyrimidine binding. Moreover, we found that an alanine residue adjacent to the GXXG loop is critical for binding. SELEX (systematic evolution of ligands by exponential enrichment), chromatin immunoprecipitation (ChIP)-seq, and crosslinking immunoprecipitation (CLIP)-seq showed that the KH domain binds C/T rich DNA and U rich RNA. Bioinformatics analysis suggested that the KH domain prefers to bind promoters. Using 15N-HSQC NMR, the optimal binding sequence was identified as TTGT. Finally, we found that the full-length DDX43 helicase prefers DNA or RNA substrates with TTGT or UUGU single strand tails, and that the KH domain is critically important for sequence specificity and unwinding processivity. Collectively, our results demonstrated that the KH domain facilitates the substrate specificity and processivity of the DDX43 helicase.

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The archaeal ATPase PINA interacts with the helicase Hjm via its carboxyl terminal KH domain remodeling and processing replication fork and Holliday junction

Cited by 10 publications

References 73 publications

Phylogenetic Diversity of Lhr Proteins and Biochemical Activities of the Thermococcales aLhr2 DNA/RNA Helicase

Phylogenetic Diversity of Lhr Proteins and Biochemical Activities of the Thermococcales aLhr2 DNA/RNA Helicase

Phosphorylation of the Archaeal Holliday Junction Resolvase Hjc Inhibits Its Catalytic Activity and Facilitates DNA Repair in Sulfolobus islandicus REY15A

The KH domain facilitates the substrate specificity and unwinding processivity of DDX43 helicase

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