Disease relevance of CCNU

• We have previously reported that the host uracil DNA glycosylase enzyme UNG2 is packaged into HIV-1 viral particles via a specific association with the integrase domain of the Gag-Pol precursor [1].
• We demonstrate that virion-associated UNG2 catalytic activity can be replaced by the packaging of heterologous dUTPase into virion, indicating that UNG2 acts to counteract dUTP misincorporation in the viral genome [2].
• Our findings indicate that pharmacologic strategies aimed toward blocking UNG2 packaging should be explored as potential HIV/AIDS therapeutics [2].
• In this study, we investigated whether UNG2 was packaged into two phylogenetically closely related primate lentiviruses, HIV-2(ROD) and SIV(MAC239) [3].
• We then showed by Western blotting that highly purified HIV-2 and SIV(MAC) viral particles did not incorporate host UNG2, contrasting with the presence of UNG2 in HIV-1 viral particles [3].

High impact information on CCNU

• Therefore, HIV-1 prevents incorporation of dUTP in viral cDNA by UNG2-mediated uracil excision followed by a dNTP-dependent, reverse transcriptase-mediated endonucleolytic cleavage and finally by strand-displacement polymerization [2].
• Thus, PPM1D may inhibit BER by dephosphorylating UNG2 to facilitate its inactivation after completion of DNA repair [4].
• We have identified UNG2 phosphorylation sites at threonines 6 and 126 that exhibit enhanced phosphorylation following UV irradiation [4].
• PPM1D dephosphorylation of UNG2 at phosphothreonine 6 is associated with reduced UNG2 activity [4].
• Our results may explain why SMUG1 cannot compensate the UNG2 deficiency in human B cells, and are fully consistent with the DNA deamination model that requires active nuclear UNG2 [5].

Biological context of CCNU

• In both cases, we find UNG2 activity and protein to be completely dispensable for virus replication [6].
• Overexpression of ung1 or human UNG2 induces a DNA checkpoint-dependent cell cycle delay and causes cell death which is enhanced when the checkpoints are inactive [7].
• Major substrates of these enzymes, a uracil opposite an adenine for UNG2 and an adenine opposite an 8-oxoguanine for MYH, are formed during DNA replication [8].
• In this work, we aimed to investigate the replication of HIV-1 viruses rendered deficient in virion-associated UNG2 by single or double point mutations in the region 170-180 of IN [9].
• During each of these processes, UNG2 recognizes uracilated DNA and excises the uracil base by flipping it into the enzyme active site [10].

Anatomical context of CCNU

• We have found the highest levels of UNG1 mRNA in skeletal muscle, heart and testis and the highest UNG2 mRNA levels in testis, placenta, colon, small intestine and thymus, all of which contain proliferating cells [11].
• In antigen-stimulated B-lymphocytes UNG2 removes uracil from U/G mispairs as part of somatic hypermutation and class switch recombination processes [12].
• Vpr-mediated incorporation of UNG2 into HIV-1 particles is required to modulate the virus mutation rate and for replication in macrophages [13].
• However, no major effect on TR-mediated DNA replication was observed when a UNG2-deficient (UNG(-/-)) cell line was used [14].
• Examination of electrophoretically resolved randomly generated PCR amplicons from mature murine oocytes revealed the presence of a short sequence with partial homology to a cyclin-like human uracil DNA glycosylase (UDG2), a member of an important group of base-excision enzymes that remove misincorporated or cytosine-derived uracil from nascent DNA [15].

Associations of CCNU with chemical compounds

• The UV-induced phosphorylated forms of UNG2 are more active than nonphosphorylated forms in mediating uracil-associated DNA cleavage [4].
• The resulting triskelion oximes were directly screened for inhibitory activity and the most potent of these showed micromolar binding affinities to UNG2 and dUTPase [16].
• Human nuclear uracil DNA glycosylase (UNG2) and deoxyuridine triphosphate nucleotidohydrolase (dUTPase) are the primary enzymes that prevent the incorporation and accumulation of deoxyuridine in genomic DNA [16].
• We propose a model in which hUNG2 is responsible for both prereplicative removal of deaminated cytosine and postreplicative removal of misincorporated uracil at the replication fork [17].

Regulatory relationships of CCNU

• Addition of PCNA (30-810 pmol) to standard uracil-DNA glycosylase reactions containing linear [uracil-(3)H]DNA stimulated UNG2 catalytic activity up to 2.6-fold [18].

Other interactions of CCNU

• Here, we demonstrate that PPM1D interacts with the nuclear isoform of uracil DNA glycosylase, UNG2, and suppresses base excision repair (BER) [4].
• Association of UNG2 with PCNA was abolished by the addition of 100 mM NaCl, and significantly decreased in the presence of 10 mM MgCl(2) [18].
• We report that the major nuclear uracil-DNA glycosylase (UNG2) increases in S phase, during which it co-localizes with incorporated BrdUrd in replication foci [19].
• Synthesis and high-throughput evaluation of triskelion uracil libraries for inhibition of human dUTPase and UNG2 [16].
• RESULTS: Three novel missense variants were identified NEIL2 C367A, TDG3 A196G and UNG2 C262T in patients, which were not observed in 188 healthy control DNAs [20].

Analytical, diagnostic and therapeutic context of CCNU

• UNG2 co-eluted with PCNA during size exclusion chromatography and bound to a PCNA affinity column [18].
• SDS-PAGE analysis of Ugi affinity-purified and immunoprecipitated UDG2 reveals two closely migrating phosphate-containing species, indicating that UDG2 either contains multiple phosphorylation sites (resulting in heterogeneous migration) or that two distinct forms of UDG2 exist in the cell [21].

References