A Nick-sensing DNA 3′-Repair Enzyme fromArabidopsis

Abstract

DNA single-strand breaks, a major cause of genome instability, often produce unconventional end groups that must be processed to restore terminal moieties suitable for reparative DNA gap filling or ligation. Here, we describe a bifunctional repair enzyme from Arabidopsis (named AtZDP) that recognizes DNA strand breaks and catalyzes the removal of 3-end-blocking lesions. The isolated C-terminal domain of AtZDP is by itself competent for 3-end processing, but not for strand break recognition. The N-terminal domain instead contains three Cys 3 -His zinc fingers and recognizes various kinds of damaged double-stranded DNA. Gapped DNA molecules are preferential targets of AtZDP, which bends them by ϳ73°upon binding, as measured by atomic force microscopy. Potential partners of AtZDP were identified in the Arabidopsis genome using the human single-strand break repairosome as a reference. These data identify a novel pathway for single-strand break repair in which a DNA-binding 3-phosphoesterase acts as a "nick sensor" for damage recognition, as the catalyst of one repair step, and possibly as a nucleation center for the assembly of a fully competent repair complex.

Solution Structure and DNA Binding of the Zinc-finger Domain from DNA Ligase IIIα

“…Instead, binding of PARP-like fingers most likely involves recognition of overall features such as the ability to form a V-shaped structure (see below). 11,22 In this context, it might be expected that the DNAbinding surface of the DL3a zinc finger would be larger than that required by GATA-1, consistent with our proposal that the surface of the b-sheet plays a role in DNA binding in addition to the flexible loops identified earlier.…”

“…9 The N terminus of DL3 contains a putative zinc finger sequence motif of the C 2 HC type, involving residues Cys18, Cys21, His52 and Cys55. 10 This sequence shows a high degree of homology with two putative zinc fingers from another protein involved in the BER and SSBR DNA repair pathways, poly(adenosine ribose)polymerase-1 (PARP-1), and also with three from the nick-sensing DNA 3 0 -phosphoesterase AtZDP from Arabidopsis thaliana; 11 in all five cases all of the zinc coordinating residues and most of the hydrophobic amino acid residues occupy analogous positions ( Figure 1). Although to our knowledge zinc binding has only been demonstrated explicitly for PARP-1, 12 the proposed zinc binding residues are absolutely conserved across the family of fingers (Figure 1), mutation of these residues abrogates function, 10,12,13 and in vitro experiments show loss of DNA binding in the absence of DTT at least in the case of AtZDP zinc fingers.…”

A tandem array of CBF/DREB1 genes is located in a major freezing tolerance QTL region on Medicago truncatula chromosome 6

“…Functional data regarding these candidate genes are still limited at present (Additional file 11). Only homologs/orthologs of MtCBF2 - 3 ; 5 - 14 , MtBAG - 1 , MTR _ 054s0019 and MtZDP have been studied in plant species [13, 15, 35, 51–54]. For MtPERLD and MTR _ 050s0020 which are members of evolutionary conserved gene families, only non-plant homologs have been described [55–57].…”

Evolutionary Genomics of the HAD Superfamily: Understanding the Structural Adaptations and Catalytic Diversity in a Superfamily of Phosphoesterases and Allied Enzymes

“…In plants, the phosphatase is fused to a Znfinger found in poly(ADP-ribose) polymerases and DNA-ligases (AtZPD _ At in Figure 9). 101,102 Another previously uncharacterized eukaryotic subfamily is found in animals and is fused to the phosphopeptide-binding forkhead-associated domain (FHA) (PNKP_Hs in Figure 9). Both the Zn-finger and FHA domain are likely to be independent means of recruiting these phosphatases to regions of DNA damage.…”