Search

ZFP57 is necessary for maintaining repressive epigenetic modifications at Imprinting control regions (ICRs). In mouse embryonic stem cells (ESCs), ZFP57 binds ICRs (ICRBS) and many other loci (non-ICRBS). To address the role of ZFP57 on all its target sites, we performed high-throughput and multi-locus analyses of inbred and hybrid mouse ESC lines carrying different gene knockouts. By using an allele-specific RNA-seq approach, we demonstrate that ZFP57 loss results in derepression of the imprinted allele of multiple genes in the imprinted clusters. We also find marked epigenetic differences between ICRBS and non-ICRBS suggesting that different cis-acting regulatory functions are repressed by ZFP57 at these two classes of target loci. Overall, these data demonstrate that ZFP57 is pivotal to maintain the allele-specific epigenetic modifications of ICRs that in turn are necessary for maintaining the imprinted expression over long distances. At non-ICRBS, ZFP57 inactivation results in acquisition of epigenetic features that are characteristic of poised enhancers, suggesting that another function of ZFP57 in early embryogenesis is to repress cis-acting regulatory elements whose activity is not yet required.

ADP-ribosylation, a modification of proteins, nucleic acids, and metabolites, confers broad functions, including roles in stress responses elicited, for example, by DNA damage and viral infection and is involved in intra- and extracellular signaling, chromatin and transcriptional regulation, protein biosynthesis, and cell death. ADP-ribosylation is catalyzed by ADP-ribosyltransferases (ARTs), which transfer ADP-ribose from NAD(+) onto substrates. The modification, which occurs as mono- or poly-ADP-ribosylation, is reversible due to the action of different ADP-ribosylhydrolases. Importantly, inhibitors of ARTs are approved or are being developed for clinical use. Moreover, ADP-ribosylhydrolases are being assessed as therapeutic targets, foremost as antiviral drugs and for oncological indications. Due to the development of novel reagents and major technological advances that allow the study of ADP-ribosylation in unprecedented detail, an increasing number of cellular processes and pathways are being identified that are regulated by ADP-ribosylation. In addition, characterization of biochemical and structural aspects of the ARTs and their catalytic activities have expanded our understanding of this protein family. This increased knowledge requires that a common nomenclature be used to describe the relevant enzymes. Therefore, in this viewpoint, we propose an updated and broadly supported nomenclature for mammalian ARTs that will facilitate future discussions when addressing the biochemistry and biology of ADP-ribosylation. This is combined with a brief description of the main functions of mammalian ARTs to illustrate the increasing diversity of mono- and poly-ADP-ribose mediated cellular processes.

Mono-ADP-ribosylation is a reversible post-translational modification that is catalysed by either bacterial pathogenic toxins or endogenous cellular ADP- ribosyltransferases (ARTs), and that can modulate the functions of target proteins. This latter aspect has only recently emerged as an important mechanism in mammalian cells for the control of a vast array of cellular processes. Some of the members of the poly-ADP-ribosyl-polymerase (PARP) family are among the mammalian enzymes that can catalyse mono-ADP-ribosylation. Here, based on biochemical data, I provide evidence that the PARP family member PARP12 is a mono-ART (mART) that can modify acidic amino acid residues. Using an affinity purification method based on a protein module that recognises ADP-ribosylated proteins (the macro domain), together with liquid chromatography coupled to tandem mass spectrometry, I have identified 25 putative PARP12 substrates that are involved in different cellular processes. The identification of this range of substrates suggests that PARP12 is involved in multiple cellular functions, ranging from regulation of small-GTPases to the control of newly synthesized RNA. To determine its biological role, I analysed its intracellular localisation. Data reported here show that PARP12 is a mART that is localised to the Golgi complex, where it can use the NAD+ pool present on this organelle to modify Golgi-localised proteins. In line with these findings, among the PARP12 substrates, I identified Rab1A, a small GTPase that controls both . transport from the endoplasmic reticulum to the Golgi- complex, and the structure of the Golgi itself My data suggest a role for PARP12 in maintenance of Golgi structure, potentially through ADP-ribosylation of Rab1A. A further aspect I have analysed is the ADP-ribosylation of the dual function protein C-terminal binding protein 1/BFA- dependent ADP-ribosylated substrate (CtBP1/BARS) as a mechanism of action of the toxin brefeldin A (BFA). BFA-ind

ADP-ribosylation, a modification of proteins, nucleic acids, and metabolites, confers broad functions, including roles in stress responses elicited, for example, by DNA damage and viral infection and is involved in intra- and extracellular signaling, chromatin and transcriptional regulation, protein biosynthesis, and cell death. ADP-ribosylation is catalyzed by ADP-ribosyltransferases (ARTs), which transfer ADP-ribose from NAD(+) onto substrates. The modification, which occurs as mono- or poly-ADP-ribosylation, is reversible due to the action of different ADP-ribosylhydrolases. Importantly, inhibitors of ARTs are approved or are being developed for clinical use. Moreover, ADP-ribosylhydrolases are being assessed as therapeutic targets, foremost as antiviral drugs and for oncological indications. Due to the development of novel reagents and major technological advances that allow the study of ADP-ribosylation in unprecedented detail, an increasing number of cellular processes and pathways are being identified that are regulated by ADP-ribosylation. In addition, characterization of biochemical and structural aspects of the ARTs and their catalytic activities have expanded our understanding of this protein family. This increased knowledge requires that a common nomenclature be used to describe the relevant enzymes. Therefore, in this viewpoint, we propose an updated and broadly supported nomenclature for mammalian ARTs that will facilitate future discussions when addressing the biochemistry and biology of ADP-ribosylation. This is combined with a brief description of the main functions of mammalian ARTs to illustrate the increasing diversity of mono- and poly-ADP-ribose mediated cellular processes.

Mono-ADP-ribosylation is a reversible post-translational modification that is catalysed by either bacterial pathogenic toxins or endogenous cellular ADP- ribosyltransferases (ARTs), and that can modulate the functions of target proteins. This latter aspect has only recently emerged as an important mechanism in mammalian cells for the control of a vast array of cellular processes. Some of the members of the poly-ADP-ribosyl-polymerase (PARP) family are among the mammalian enzymes that can catalyse mono-ADP-ribosylation. Here, based on biochemical data, I provide evidence that the PARP family member PARP12 is a mono-ART (mART) that can modify acidic amino acid residues. Using an affinity purification method based on a protein module that recognises ADP-ribosylated proteins (the macro domain), together with liquid chromatography coupled to tandem mass spectrometry, I have identified 25 putative PARP12 substrates that are involved in different cellular processes. The identification of this range of substrates suggests that PARP12 is involved in multiple cellular functions, ranging from regulation of small-GTPases to the control of newly synthesized RNA. To determine its biological role, I analysed its intracellular localisation. Data reported here show that PARP12 is a mART that is localised to the Golgi complex, where it can use the NAD+ pool present on this organelle to modify Golgi-localised proteins. In line with these findings, among the PARP12 substrates, I identified Rab1A, a small GTPase that controls both . transport from the endoplasmic reticulum to the Golgi- complex, and the structure of the Golgi itself My data suggest a role for PARP12 in maintenance of Golgi structure, potentially through ADP-ribosylation of Rab1A. A further aspect I have analysed is the ADP-ribosylation of the dual function protein C-terminal binding protein 1/BFA- dependent ADP-ribosylated substrate (CtBP1/BARS) as a mechanism of action of the toxin brefeldin A (BFA). BFA-ind