Androgen signaling through the androgen receptor (AR) directs gene expression in both normal and prostate cancer cells. Androgen regulates multiple aspects of the AR life cycle, including its localization and post-translational modification, but understanding how modifications are read and integrated with AR activity has been difficult. Here, we show that ADP-ribosylation regulates AR through a nuclear pathway mediated by Parp7. We show that Parp7 mono-ADP-ribosylates agonist-bound AR, and that ADP-ribosyl-cysteines within the N-terminal domain mediate recruitment of the E3 ligase Dtx3L/Parp9. Molecular recognition of ADP-ribosyl-cysteine is provided by tandem macrodomains in Parp9, and Dtx3L/Parp9 modulates expression of a subset of AR-regulated genes. Parp7, ADP-ribosylation of AR, and AR-Dtx3L/Parp9 complex assembly are inhibited by Olaparib, a compound used clinically to inhibit poly-ADP-ribosyltransferases Parp1/2.
Fig: a PC3-AR cells were treated with the synthetic androgen R1881 for the indicated times. AR was isolated by IP, and the protein complexes were examined by SDS-PAGE and silver staining. The proteins migrating slightly faster than Hsp90 were identified as Dtx3L and Parp9 by mass spectrometry (MS). b AR IP’s from R1881-treated PC3-AR cells immunoblotted for AR, Dtx3L, and Parp9. c Dtx3L IP’s from R1881-treated PC3-AR cells immunoblotted for AR, Dtx3L, and Parp9. d Biochemical assays for ubiquitin E3 and ADP-ribosyltransferase activities in AR-Dtx3L/Parp9 complexes. Lanes 1–5 contain negative and positive controls (buffer, recombinant Dtx3L/Parp9, E1, E2, biotin-NAD+) for detection of Ub-modified Parp9 and ADP-ribosylated ubiquitin. Lanes 6–9 contain AR IP’s from control and R1881-treated cells incubated at 30 °C, or on ice to block the reaction. Lane 8 shows that Dtx3L/Parp9 bound to AR has E3 activity (Ub-induced mobility shift of Parp9) and mono-ADP-ribosyltransferase activity towards Ub detected via ADPr-biotin. Lane 10 shows the levels of E3 and ADP-ribosyltransferase activity detected in the Dtx3L/Parp9 directly isolated by IP. Source data are provided as a Source data file.
Our study reveals the components of an androgen signaling axis that uses a writer and reader of ADP-ribosylation to regulate protein-protein interactions and AR activity.
Yang, CS., Jividen, K., Kamata, T. et al. Androgen signaling uses a writer and a reader of ADP-ribosylation to regulate protein complex assembly. Nat Commun 12, 2705 (2021). https://doi.org/10.1038/s41467-021-23055-6