Abstract
BackgroundPoly (ADP-ribosyl) ation (PARylation) is an important posttranslational modification that regulates DNA repair, gene transcription, stress responses and developmental processes in multicellular organisms. Poly (ADP-ribose) polymerase (PARP) catalyzes PARylation by consecutively adding ADP-ribose moieties from NAD+ to the amino acid receptor residues on target proteins. Arabidopsis has three canonical PARP members, and two of these members, AtPARP1 and AtPARP2, have been demonstrated to be bona fide poly (ADP-ribose) polymerases and to regulate DNA repair and stress response processes. However, it remains unknown whether AtPARP3, a member that is highly expressed in seeds, has similar biochemical activity to that of AtPARP1 and AtPARP2. Additionally, although both the phylogenetic relationships and structural similarities indicate that AtPARP1 and AtPARP2 correspond to animal PARP1 and PARP2, respectively, two previous studies have indicated that AtPARP2, and not AtPARP1, accounts for most of the PARP activity in Arabidopsis, which is contrary to the knowledge that PARP1 is the predominant PARP in animals.ResultsIn this study, we obtained both in vitro and in vivo evidence demonstrating that AtPARP3 does not act as a typical PARP in Arabidopsis. Domain swapping and point mutation assays indicated that AtPARP3 has lost NAD+-binding capability and is inactive. In addition, our results showed that AtPARP1 was responsible for most of the PARP enzymatic activity in response to the DNA damage-inducing agents zeocin and methyl methanesulfonate (MMS) and was more rapidly activated than AtPARP2, which supports that AtPARP1 remains the predominant PARP member in Arabidopsis. AtPARP1 might first become activated by binding to damaged sites, and AtPARP2 is then poly (ADP-ribosyl) ated by AtPARP1 in vivo.ConclusionsCollectively, our biochemical and genetic analysis results strongly support the notion that AtPARP3 has lost poly (ADP-ribose) polymerase activity in plants and performs different functions from those of AtPARP1 and AtPARP2. AtPARP1, instead of AtPARP2, plays the predominant role in PAR synthesis in both seeds and seedlings. These data bring new insights into our understanding of the physiological functions of plant PARP family members.
Highlights
(ADP-ribosyl) ation (PARylation) is an important posttranslational modification that regulates DNA repair, gene transcription, stress responses and developmental processes in multicellular organisms
HsPARP1 is thought to be the founding member of the Poly (ADP-ribose) polymerase (PARP) family in humans [1, 3, 11]. Both HsPARP1 and AtPARP1 have five important domains with known functions arranged from the N to the C terminus: three N-terminal zinc fingers responsible for DNA damage detection, the BRCA-1 C-terminus (BRCT) domain for phospho-protein binding, the WGR domain with the conserved Trp-Gly-Arg (WGR) motif for nucleic acid binding, the PARP regulatory domain (PRD) or helical subdomain (HD) believed to regulate PAR-branching, and the C-terminal characteristic PARP domain with catalytic activity [3, 33] (Additional file 1: Figure S1A)
The sequence analysis revealed that both AtPARP1 and AtPARP2 have a typical histidine-tyrosine-glutamic acid (H-Y-E) catalytic triad, whereas AtPARP3 has an alternative histidine-valineglutamic acid (C-V-E) triad in its catalytic core (Additional file 1: Figure S1B)
Summary
(ADP-ribosyl) ation (PARylation) is an important posttranslational modification that regulates DNA repair, gene transcription, stress responses and developmental processes in multicellular organisms. (ADP-ribose) polymerase (PARP) catalyzes PARylation by consecutively adding ADP-ribose moieties from NAD+ to the amino acid receptor residues on target proteins. Arabidopsis has three canonical PARP members, and two of these members, AtPARP1 and AtPARP2, have been demonstrated to be bona fide poly (ADP-ribose) polymerases and to regulate DNA repair and stress response processes. It remains unknown whether AtPARP3, a member that is highly expressed in seeds, has similar biochemical activity to that of AtPARP1 and AtPARP2. Seventeen members in humans are involved in various physiological processes, such as DNA repair, cell death, transcriptional regulation, energy metabolism, and chromatin remodeling [2, 11, 12]
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