Protein ADP-ribosylation is a reversible post-translational modification of cellular proteins that is catalysed by enzymes that transfer one (mono) or several (poly) units of ADP-ribose from β-NAD+
to a specific amino acid of the target protein. The most studied member of the ADP-ribosyltransferase family is PARP1 (also known as ADP-ribosyltransferase diphtheria toxin-like 1, ARTD1), which is directly activated by DNA strand breaks and is involved in DNA damage repair, chromatin remodelling and transcriptional regulation. Much less is known about the further 16 members of this family. Among these, ARTD10/PARP10 has been previously characterised as a mono-ADP-ribosyltransferase with a role in cell proliferation and in NF-kB signalling. In the present study, we identified the glycolytic enzyme GAPDH as an interactor and a novel cellular target for ARTD10/PARP10. Moreover, we detected the co-localisation of GAPDH and ARTD10/PARP10 in well-defined cytosolic bodies, which we show here to be membrane-free, rounded structures using immunogold labelling and electron microscopy. Using the cognitive binding module macro
domain to visualise ADP-ribosylated proteins by immunofluorescence microscopy in cells over-expressing the ARTD10/PARP10 enzyme, we show that the staining of the ARTD10/PARP10-dependent cytosolic bodies was lost when the cells were treated with compounds that inhibit ARTD10/PARP10, either by directly inhibiting the enzyme or by reducing the cellular NAD+
levels. In parallel, the same treatment affected the co-localisation of GAPDH and ARTD10/PARP10, as GAPDH disappeared from the cytosolic cell bodies, which indicates that its presence there depends on the catalytic activity of ARTD10/PARP10. In line with this, in cells over-expressing the ARTD10/PARP10 catalytic domain alone, which we show here to form stress granules, GAPDH was recruited into stress granules. These data identify ARTD10/PARP10 as the enzyme that modifies and recruits GAPDH into cytosolic structures.