A human 5′-tyrosyl DNA phosphodiesterase that repairs topoisomerase-mediated DNA damage

Abstract

Topoisomerases regulate DNA topology and are fundamental to many aspects of chromosome metabolism. Their activity involves the transient cleavage of DNA, which, if it occurs near sites of endogenous DNA damage or in the presence of topoisomerase poisons, can result in abortive topoisomerase-induced DNA strand breaks. These breaks feature covalent linkage of the enzyme to the DNA termini by a 3'- or 5'-phosphotyrosyl bond and are implicated in hereditary human disease, chromosomal instability and cancer, and underlie the clinical efficacy of an important class of anti-tumour poisons. The importance of liberating DNA termini from trapped topoisomerase is illustrated by the progressive neurodegenerative disease observed in individuals containing a mutation in tyrosyl-DNA phosphodiesterase 1 (TDP1), an enzyme that cleaves 3'-phosphotyrosyl bonds. However, a complementary human enzyme that cleaves 5'-phosphotyrosyl bonds has not been reported, despite the effect of DNA double-strand breaks containing such termini on chromosome instability and cancer. Here we identify such an enzyme in human cells and show that this activity efficiently restores 5'-phosphate termini at DNA double-strand breaks in preparation for DNA ligation. This enzyme, TTRAP, is a member of the Mg(2+)/Mn(2+)-dependent family of phosphodiesterases. Cellular depletion of TTRAP results in increased susceptibility and sensitivity to topoisomerase-II-induced DNA double-strand breaks. TTRAP is, to our knowledge, the first human 5'-tyrosyl DNA phosphodiesterase to be identified, and we suggest that this enzyme is denoted tyrosyl DNA phosphodiesterase-2 (TDP2).

Involvement of the host DNA-repair enzyme TDP2 in formation of the covalently closed circular DNA persistence reservoir of hepatitis B viruses

“…2B). The preference of TDP2 for 5′ phosphotyrosyl bonds is consistent with recent X-ray data (27)(28)(29), but TDP2 also is active on 3′-substrates (20,26,30). Although TDP2 likely is the only enzyme of its kind (30), complete knockout of TDP2 in chicken DT40 cells and in mice was not lethal (20), possibly reflecting the power of redundancy in DNA repair.…”

“…Alternatively, trapped TOP1 can be removed nucleolytically (21,22). For repair of the 5′-phosphotyrosyl-linked TOP2 adducts, only nucleolytic mechanisms were considered (23,24) until a protein known as "TRAF and TNF receptor-associated protein" (TTRAP) or "ETS1-associated protein II" (EAPII) (25) was found to have such an activity (26); that protein has been renamed "TDP2" (Fig. 2B).…”

The PML nuclear bodies-associated protein TTRAP regulates ribosome biogenesis in nucleolar cavities upon proteasome inhibition

“…siTTRAP cells were then reconstituted with constructs for phosphodiesterase mutants E152A and D262A. 1 No detectable changes in pre-rRNA or processing intermediates were observed in the absence of MG132 treatment with all transfected constructs (Supplementary Figure S4). Notably, phosphodiesterase inactive TTRAP (both E152A and D262A mutants) was able to rescue the alterations in rRNA biogenesis and this effect was comparable to that of wt protein (Figure 7b), thus suggesting that the enzymatic activity of TTRAP is dispensable for its control on rRNA maturation.…”

“…1 It is a member of the endonuclease/exonuclease/ phosphatase family of proteins containing an Mg(2 þ )-dependent apurinic/apyrimidinic endonuclease Ape1-like domain. 2 Various yeast-two hybrid (Y2H) screenings and functional studies have unveiled TTRAP ability to interact with TNF receptors (TNFR) family members and TNFR-associated factors (TRAFs), especially TRAF6.…”

TDP2, TOP2, and SUMO: what is ZATT about?

“…The enzyme 5′-tyrosyl DNA phosphodiesterase 2 (TDP2) can initiate the repair of TOP2cc by hydrolytic removal of trapped topoisomerase peptide from the 5′-termini of DSBs, thereby allowing the DSB to be religated [3]. The importance of this enzyme is illustrated by its proposed role in oncogenic translocations, resistance to chemotherapeutic topoisomerase 'poisons' and by causative mutations in the TDP2 gene in Spinocerebellar Ataxia Autosomal Recessive 23 (SCAR23) [4,5].…”