Journal
NATURE CHEMICAL BIOLOGY
Volume 13, Issue 3, Pages 302-+Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/NCHEMBIO.2280
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- NIH-NIGMS [R01 GM087544]
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ADP-ribosylation is a post-translational modification that is known to be involved in cellular homeostasis and stress but has been challenging to analyze biochemically. To facilitate the detection of ADP-ribosylated proteins, we show that an alkyne-adenosine analog, N-6-propargyl adenosine (N(6)pA), is metabolically incorporated in mammalian cells and enables fluorescence detection and proteomic analysis of ADP-ribosylated proteins. Notably, our analysis of N(6)pA-labeled proteins that are upregulated by oxidative stress revealed differential ADP-ribosylation of small GTPases. We discovered that oxidative stress induced ADP-ribosylation of Hras on Cys181 and Cys184 in the C-terminal hypervariable region, which are normally S-fatty-acylated. Downstream Hras signaling is impaired by ADP-ribosylation during oxidative stress, but is rescued by ADP-ribosyltransferase inhibitors. Our study demonstrates that ADP-ribosylation of small GTPases not only is mediated by bacterial toxins but is endogenously regulated in mammalian cells. N(6)pA provides a useful tool to characterize ADP-ribosylated proteins and their regulatory mechanisms in cells.
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