Yang Chun-Song, Jividen Kasey, Spencer Adam, Dworak Natalia, Ni Li, Oostdyk Luke T, Chatterjee Mandovi, Kuśmider Beata, Reon Brian, Parlak Mahmut, Gorbunova Vera, Abbas Tarek, Jeffery Erin, Sherman Nicholas E, Paschal Bryce M
Center for Cell Signaling, University of Virginia, West Complex, 1335 Lee Street, Charlottesville, VA 22908, USA.
Center for Cell Signaling, University of Virginia, West Complex, 1335 Lee Street, Charlottesville, VA 22908, USA; Department of Biochemistry and Molecular Genetics, University of Virginia, PO Box 800733, Charlottesville, VA 22908, USA.
Mol Cell. 2017 May 18;66(4):503-516.e5. doi: 10.1016/j.molcel.2017.04.028.
ADP-ribosylation of proteins is emerging as an important regulatory mechanism. Depending on the family member, ADP-ribosyltransferases either conjugate a single ADP-ribose to a target or generate ADP-ribose chains. Here we characterize Parp9, a mono-ADP-ribosyltransferase reported to be enzymatically inactive. Parp9 undergoes heterodimerization with Dtx3L, a histone E3 ligase involved in DNA damage repair. We show that the Dtx3L/Parp9 heterodimer mediates NAD-dependent mono-ADP-ribosylation of ubiquitin, exclusively in the context of ubiquitin processing by E1 and E2 enzymes. Dtx3L/Parp9 ADP-ribosylates the carboxyl group of Ub Gly76. Because Gly76 is normally used for Ub conjugation to substrates, ADP-ribosylation of the Ub carboxyl terminus precludes ubiquitylation. Parp9 ADP-ribosylation activity therefore restrains the E3 function of Dtx3L. Mutation of the NAD binding site in Parp9 increases the DNA repair activity of the heterodimer. Moreover, poly(ADP-ribose) binding to the Parp9 macrodomains increases E3 activity. Dtx3L heterodimerization with Parp9 enables NAD and poly(ADP-ribose) regulation of E3 activity.
蛋白质的ADP核糖基化正成为一种重要的调节机制。根据家族成员的不同，ADP核糖基转移酶要么将单个ADP核糖连接到靶标上，要么生成ADP核糖链。在这里，我们对Parp9进行了表征，Parp9是一种据报道无酶活性的单ADP核糖基转移酶。Parp9与Dtx3L发生异源二聚化，Dtx3L是一种参与DNA损伤修复的组蛋白E3连接酶。我们表明，Dtx3L/Parp9异源二聚体仅在E1和E2酶进行泛素加工的背景下介导泛素的NAD依赖性单ADP核糖基化。Dtx3L/Parp9将泛素Gly76的羧基进行ADP核糖基化。由于Gly76通常用于泛素与底物的结合，泛素羧基末端的ADP核糖基化排除了泛素化。因此，Parp9的ADP核糖基化活性抑制了Dtx3L的E3功能。Parp9中NAD结合位点的突变增加了异源二聚体的DNA修复活性。此外，聚（ADP-核糖）与Parp9大结构域的结合增加了E3活性。Dtx3L与Parp9的异源二聚化使NAD和聚（ADP-核糖）能够调节E3活性。
