Repair Synthesis and Ligation

TCR refers to a repair pathway that preferentially and rapidly repairs lesions on the transcribed strand of active genes (less than 1% of human DNA) [47,48]. In TCR-NER the ability of a lesion to block RNA polymerases seems critical. The stalled polymerase recruits at least two TRC-specific factors CSA and CSB. The remainder of the TCR-NER pathway (i.e., unwinding, incision, excision, DNA synthesis, and ligation) may be identical to the GGR-NER pathway. The breast and ovarian cancer susceptibility gene BRCAl is involved in TCR [49,50] however, the exact biological function of the protein remains uncertain (for review see [51]). 

The NER pathway is also present in prokaryotic organisms, such as Escherichia coli, as well as in eukaryotes from yeast to mammals. Many of the basic steps of NER are evolutionarily conserved, including damage recognition and dual incisions to excise the lesion, helicase activity to displace the excised oligonucleotide and repair factors, and synthesis and ligation enzymes to seal the nick [19, 20], Nevertheless, the biochemical features in prokaryotes and eukaryotes are distinct. 

Matsumoto, Y., and Bogenhagen, D. F. (1991). Repair of a synthetic abasic site involves concerted reactions of DNA synthesis followed by excision and ligation. Mol. Cell. Biol. 11, 4441-4447. 

Figure 6 Nucleotide excision repair in (A) E. coli and (B) humans. There are five basic steps of nucieotide excision repair (1) damage recognition, (2) dual incisions, (3) release of the excised oligomer, (4) repair synthesis to fill in the gap, and (5) ligation.

Both of these processes require multiple enzymes acting either sequentially or, as indicated above, as molecular complexes. The hallmarks of both NER and BER are strand scission, removal of a segment of DNA containing the adducted base, 5 to 3 -oriented DNA patch synthesis through the action of a polymerase, using the intact strand as a template, and ligation of the free ends. The distinctions between these two mechanisms are the proteins involved and the types of adducts that are repaired. Bulky adducts, like those of aromatic compounds or... 

Figure 12. (A) DNA ligase reaction (B) DNA synthesis by repair, (C) and (D) blimt-end ligation to give synthetic duplex (E) containing restiictian enzyme sites trimming by restriction enzymes to give synthetic duplex with exposed recognition sites (F) for insertion in plasmid.

The published data indicate that ADPRT inhibitors do not block early steps in DNA repair, because the single-strand breaks appear in the presence of the enzyme inhibitor, as they do in its absence. DNA repair synthesis is not inhibited by the ADPRT inhibitors [11], as was first reported from the laboratories of N. Berger and M. Miwa. These observations surest that ADPRT activity is not needed for the early steps in DNA excision repair. We therefore turn our attention to the DNA ligase reaction. The available published data has led us to propose that ADP-ribosylation regulates the rejoining or the ligation step of DNA repair. 

In summary then, it has been demonstrated that ADP-ribosylation reactions regulate diverse cellular processes. Poly(ADP-ribosylation) participates in DNA repair in eukaryotic organisms, possibly by regulating the ligation step (Fig. 13). In addition, ADP-ribosylation reactions modulate cell differentiation, protein synthesis and membrane adenyl cyclase reactions. No doubt, as time passes, the list of cellular processes modulated by ADP-ribosylation reactions will grow. 

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