Functional Roles of PARP2 in Assembling Protein–Protein Complexes Involved in Base Excision DNA Repair

Abstract

Poly(ADP-ribose) polymerase 2 (PARP2) participates in base excision repair (BER) alongside PARP1, but its functions are still under study. Here, we characterize binding affinities of PARP2 for other BER proteins (PARP1, APE1, Polβ, and XRCC1) and oligomerization states of the homo- and hetero-associated complexes using fluorescence-based and light scattering techniques. To compare PARP2 and PARP1 in the efficiency of PAR synthesis, in the absence and presence of protein partners, the size of PARP2 PARylated in various reaction conditions was measured. Unlike PARP1, PARP2 forms more dynamic complexes with common protein partners, and their stability is effectively modulated by DNA intermediates. Apparent binding affinity constants determined for homo- and hetero-oligomerized PARP1 and PARP2 provide evidence that the major form of PARP2 at excessive PARP1 level is their heterocomplex. Autoregulation of PAR elongation at high PARP and NAD+ concentrations is stronger for PARP2 than for PARP1, and the activity of PARP2 is more effectively inhibited by XRCC1. Moreover, the activity of both PARP1 and PARP2 is suppressed upon their heteroPARylation. Taken together, our findings suggest that PARP2 can function differently in BER, promoting XRCC1-dependent repair (similarly to PARP1) or an alternative XRCC1-independent mechanism via hetero-oligomerization with PARP1.

Dual function of HPF1 in the modulation of PARP1 and PARP2 activities

“…Notably, the initial rate of automodification in the absence of HPF1 depends on NAD + concentration more strongly for PARP1- than PARP2-catalyzed reaction (27-fold increase at 10 μM vs. 1 μM NAD + concentration for PARP1 and 5‐fold for PARP2). This difference, which can be explained by a higher activity of PARP1 at the elongation step 10 , 16 , disappeared in the presence of HPF1 (6- and 8-fold increase at 10 μM vs. 1 μM NAD + concentration for PARP1 and PARP2, respectively). These data suggest that HPF1 controls the balance between initiation and elongation events upon the ADP-ribosylation reaction.…”

“…PARP2 was also discovered as an enzyme that catalyzes the synthesis of PAR 5 . The role of PARP2 and its cooperation with PARP1 is under intensive investigation 6 – 10 . In human cells, the majority of PARP activity is exerted by PARP1 (around 90%) and by PARP2 (10‒15%) 7 .…”

“…Difference in the affinities of PARP1 and PARP2 for various DNA structures in the nucleosome context was lately revealed 18 . PARP2 synthesizes shorter PAR chains than PARP1 does and also inhibits PARP1-catalyzed PAR synthesis via hetero-oligomerization of the two PARPs 8 , 10 . It has been shown that removal of histone H3 is largely suppressed in PARP2-deficient cells 20 .…”

A sePARate phase? Poly(ADP-ribose) versus RNA in the organization of biomolecular condensates

“…It is possible that PARylation can alter a conformation and therefore interaction profile of the modified protein similar to ( 145 ). Moreover, PAR covalently attached to the protein may be a significantly more specifically organized structure than free PAR ( 146 ), and represent a specific platform for initiating the assembly of macromolecules ( 147 ).…”

HPF1 can promote opposite effects on different stages of PARP1 and PARP2 autoPARylation and histone modification

“…Overlapping functions of PARP1 and PARP2 in regulation of these processes were shown by using model DNA duplexes as well as nucleosomes [6,7]. PARP2 synthesizes shorter PAR chains than PARP1 does and also inhibits PARP1catalyzed PAR synthesis via hetero-oligomerization of the two PARPs [8]. It has been shown that removal of histone H3 is largely suppressed in PARP2-deficient cells21.…”