Divergent structures of Mammalian and gammaherpesvirus uracil DNA glycosylases confer distinct DNA binding and substrate activity

Mu, Yunxiang; Zelazowska, Monika A.; Chen, Zaowen; Plummer, Joshua B.; Dong, Qiwen; Krug, Laurie T.; McBride, Kevin M.

doi:10.1016/j.dnarep.2023.103515

Cited by 3 publications

(19 citation statements)

References 67 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Notably, the frequencies of transversion mutations of the N-terminal mutants, mUNGΔN (1.7X10 -4 tv/bp) and MHV68/mUNG (1.9X10 -4 tv/bp) were also significantly decreased compared to mUNG (6.6 X10 -4 tv/bp) (p<0.01) (Figure 3F). Importantly, this difference in mutational frequency was not due to differences in enzymatic activity, since our previous analysis of catalytic activity demonstrated that mUNG and mUNGΔN had comparable catalytic activity on both ssDNA and dsDNA uracil containing substrates and MHV68/mUNG had slightly lower activity 23 . This result suggests that the N-terminal domain of UNG plays a major role in dictating error-prone repair outcome.…”

Section: Resultsmentioning

confidence: 89%

“…To determine whether the N-terminal or leucine loop extension structures contribute to differential mutation outcomes, we constructed chimeric UNG mutants with swapped domains 23 . Utilizing the NTZ reporter assay we compared the frequency of mutation in cells expressing mUNG, MHV68UNG or mutants with their leucine loops swapped (Ls) (Figure 3A).…”

Section: Resultsmentioning

confidence: 99%

“…Gammaherpesviruses (GHVs) which include the oncogenic human herpesviruses (HHV), Epstein–Barr virus (EBV/HHV-4), Kaposi sarcoma associated herpesvirus (KSHV/ HHV-8) and the model murine Gammaherpesvirus 68 (MHV68/MHV-4) infect and maintain life-long latency in host B lymphocytes 16 . All GHVs studied to date encode a functional UNG 17–23 . In the murine model pathogen system, the viral UNG of MHV68 (MHV68UNG) promotes virus replication in the lungs 21,22 .…”

Section: Introductionmentioning

confidence: 99%

“…MHV68 then migrates to secondary lymphoid organs where it latently expands in the GC and establishes life-long latency in the B cell compartment 22 . MHV68UNG has uracil glycosylase activity on ssDNA and dsDNA 23 and compensates for CSR deficiency in UNG knockout (KO) B cells 21 . While GHVUNGs share structure and sequence identity with mammalian UNGs in their core catalytic domain, GHVUNGs diverge from mammalian counterparts in two domains: the N-terminal region, and an extension of the leucin loop structure in their DNA binding domain 20,23 .…”

Section: Introductionmentioning

confidence: 99%

“…In GHVs, the unique leucine loop extension of UNG mediates a difference in DNA substrate choice. The GHVUNG leucine loop also imparts affinity to AP sites, a quality not found in mammalian UNGs 23 . During its life cycle, MHV68 inhabits GC cells.…”

Section: Introductionmentioning

confidence: 99%

See 4 more Smart Citations

UNG-RPA interaction governs the choice between high-fidelity and mutagenic uracil repair

Mu,

Chen,

Plummer

et al. 2024

Preprint

Self Cite

View full text Add to dashboard Cite

Mammalian Uracil DNA glycosylase (UNG) removes uracils and initiates high-fidelity base excision repair to maintain genomic stability. During B cell development, activation-induced cytidine deaminase (AID) creates uracils that UNG processes in an error-prone fashion to accomplish immunoglobulin (Ig) somatic hypermutation (SHM) or class switch recombination (CSR). The mechanism that governs high-fidelity versus mutagenic uracil repair is not understood. The B cell tropic gammaherpesvirus (GHV) encodes a functional homolog of UNG that can process AID induced genomic uracils. GHVUNG does not support hypermutation, suggesting intrinsic properties of UNG influence repair outcome. Noting the structural divergence between the UNGs, we define the RPA interacting motif as the determinant of mutation outcome. UNG or RPA mutants unable to interact with each other, only support high-fidelity repair. In B cells, transversions at the Ig variable region are abated while CSR is supported. Thus UNG-RPA governs the generation of mutations and has implications for locus specific mutagenesis in B cells and deamination associated mutational signatures in cancer.

show abstract

Section: Resultsmentioning

confidence: 89%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

UNG-RPA interaction governs the choice between high-fidelity and mutagenic uracil repair

Mu,

Chen,

Plummer

et al. 2024

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

Uracil-DNA glycosylase of murine gammaherpesvirus 68 binds cognate viral replication factors independently of its catalytic residues

Smith,

Paul,

Dong

et al. 2023

mSphere

Self Cite

View full text Add to dashboard Cite

Herpesviruses are large double-stranded DNA viruses that encode core replication proteins and accessory factors involved in nucleotide metabolism and DNA repair. Mammalian uracil-DNA glycosylases (UNG) excise deleterious uracil residues from their genomic DNA. Each herpesvirus UNG studied to date has demonstrated conservation of the enzymatic function to excise uracil residues from DNA. We previously reported that a murine gammaherpesvirus (MHV68) with a stop codon in ORF46 (ORF46.stop) that encodes for vUNG was defective in lytic replication and latency in vivo . However, a mutant virus that expressed a catalytically inactive vUNG (ORF46.CM) had no replication defect unless coupled with additional mutations in the catalytic motif of the viral dUTPase (ORF54.CM). The disparate phenotypes observed in the vUNG mutants led us to explore the non-enzymatic properties of vUNG. Immunoprecipitation of vUNG followed by mass spectrometry in MHV68-infected fibroblasts identified a complex comprising the cognate viral DNA polymerase, vPOL, encoded by ORF9 , and the viral DNA polymerase processivity factor, vPPF, encoded by ORF59 . MHV68 vUNG co-localized with vPOL and vPPF in subnuclear structures consistent with viral replication compartments. In reciprocal co-immunoprecipitations, the vUNG formed a complex with the vPOL and vPPF upon transfection with either factor alone or in combination. Lastly, we determined that key catalytic residues of vUNG are not required for interactions with vPOL and vPPF upon transfection or in the context of infection. We conclude that the vUNG of MHV68 associates with vPOL and vPPF independently of its catalytic activity. IMPORTANCE Gammaherpesviruses encode a uracil-DNA glycosylase (vUNG) that is presumed to excise uracil residues from viral genomes. We previously identified the vUNG enzymatic activity, but not the protein itself, as dispensable for gammaherpesvirus replication in vivo . In this study, we report a non-enzymatic role for the viral UNG of a murine gammaherpesvirus in forming a complex with two key components of the viral DNA replication machinery. Understanding the role of the vUNG in this viral DNA replication complex may inform the development of antiviral drugs that combat gammaherpesvirus-associated cancers.

show abstract

Sequencing of Kaposi’s Sarcoma Herpesvirus (KSHV) genomes from persons of diverse ethnicities and provenances with KSHV-associated diseases demonstrate multiple infections, novel polymorphisms, and low intra-host variance

Marshall,

Cornejo Castro,

Goodman

et al. 2024

PLoS Pathog

Self Cite

View full text Add to dashboard Cite

Recently published near full-length KSHV genomes from a Cameroon Kaposi sarcoma case-control study showed strong evidence of viral recombination and mixed infections, but no sequence variations associated with disease. Using the same methodology, an additional 102 KSHV genomes from 76 individuals with KSHV-associated diseases have been sequenced. Diagnoses comprise all KSHV-associated diseases (KAD): Kaposi sarcoma (KS), primary effusion lymphoma (PEL), KSHV-associated large cell lymphoma (KSHV-LCL), a type of multicentric Castleman disease (KSHV-MCD), and KSHV inflammatory cytokine syndrome (KICS). Participants originated from 22 different countries, providing the opportunity to obtain new near full-length sequences of a wide diversity of KSHV genomes. These include near full-length sequence of genomes with KSHV K1 subtypes A, B, C, and F as well as subtype E, for which no full sequence was previously available. High levels of recombination were observed. Fourteen individuals (18%) showed evidence of infection with multiple KSHV variants (from two to four unique genomes). Twenty-six comparisons of sequences, obtained from various sampling sites including PBMC, tissue biopsies, oral fluids, and effusions in the same participants, identified near complete genome conservation between different biological compartments. Polymorphisms were identified in coding and non-coding regions, including indels in the K3 and K15 genes and sequence inversions here reported for the first time. One such polymorphism in KSHV ORF46, specific to the KSHV K1 subtype E2, encoded a mutation in the leucine loop extension of the uracil DNA glycosylase that results in alteration of biochemical functions of this protein. This confirms that KSHV sequence variations can have functional consequences warranting further investigation. This study represents the largest and most diverse analysis of KSHV genome sequences to date among individuals with KAD and provides important new information on global KSHV genomics.

show abstract

Divergent structures of Mammalian and gammaherpesvirus uracil DNA glycosylases confer distinct DNA binding and substrate activity

Cited by 3 publications

References 67 publications

UNG-RPA interaction governs the choice between high-fidelity and mutagenic uracil repair

UNG-RPA interaction governs the choice between high-fidelity and mutagenic uracil repair

Uracil-DNA glycosylase of murine gammaherpesvirus 68 binds cognate viral replication factors independently of its catalytic residues

Sequencing of Kaposi’s Sarcoma Herpesvirus (KSHV) genomes from persons of diverse ethnicities and provenances with KSHV-associated diseases demonstrate multiple infections, novel polymorphisms, and low intra-host variance

Contact Info

Product

Resources

About