Characterization of TCDD-inducible poly-ADP-ribose polymerase (TIPARP/ARTD14) catalytic activity.

Here, we report the biochemical characterization of the mono-ADP-ribosyltransferase 2,3,7,8-tetrachlorodibenzo-p-dioxin poly-ADP-ribose polymerase which is known to repress aryl hydrocarbon receptor (AHR)-dependent transcription. We found that the nuclear localization of was dependent on a short N-terminal sequence and its zinc finger domain. Deletion and in vitro ADP-ribosylation studies identified amino acids 400-657 as the minimum catalytically active region, which retained its ability to mono-ADP-ribosylate AHR. However, the ability of TIPduanyu37 to ADP-ribosylate and repress AHR in cells was dependent on both its catalytic activity and zinc finger domain. The catalytic activity of TIPduanyu37 was resistant to meta-iodobenzylguanidine but sensitive to iodoacetamide and hydroxylamine, implicating cysteines and acidic side chains as ADP-ribosylated target residues. Mass spectrometry identified multiple ADP-ribosylated peptides in TIPduanyu37 and AHR. Electron transfer dissociation analysis of the TIPduanyu37 peptide ³³ITPLKTCFK⁴¹ revealed cysteine 39 as a site for mono-ADP-ribosylation. Mutation of cysteine 39 to alanine resulted in a small, but significant, reduction in TIPduanyu37 autoribosylation activity, suggesting that additional amino acid residues are modified, but loss of cysteine 39 did not prevent its ability to repress AHR. Our findings characterize the subcellular localization and mono-ADP-ribosyltransferase activity of identify cysteine as a mono-ADP-ribosylated residue targeted by this enzyme, and confirm the mono-ADP-ribosylation of other protein targets, such as AHR. © 2018 The Author(s).

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