Mammalian N1-adenosine PARylation is a reversible DNA modification

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• Poly(ADP-ribosyl)ation (PARylation) was traditionally considered a protein-specific post-translational modification.
• Recent in vitro studies suggested poly(ADP-ribose) polymerases (PARPs) might modify DNA termini.

Purpose of the Study:

• To investigate the physiological occurrence and characteristics of DNA poly(ADP-ribosyl)ation in mammalian tissues.
• To determine the enzymes involved and the reversibility of DNA PARylation.
• To identify the specific site and product of DNA PARylation in vivo.

Main Methods:

• Utilized ultrasensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS).
• Employed anti-PAR antibodies and anti-PAR reagents for detection.
• Performed partial hydrolysis of genomic DNA (gDNA) to identify modification products.

Main Results:

• Demonstrated that mammalian DNA is physiologically poly(ADP-ribosyl)ated, with varying levels in primary tissues.
• Confirmed DNA PARylation requires PARP1 and is reversible, as inhibiting PAR glycohydrolase (PARG) increases modification.
• Identified the modification occurring at the N1-position of adenosine, forming N1-Poly(ADP-ribosyl)-deoxyadenosine, and confirmed its in vivo presence via a deamination product.

Conclusions:

• Establishes N1-adenosine PARylation as a novel DNA modification in vivo.
• Extends the known functions of PARP enzymes beyond protein modification.
• Provides a framework for future research into the biological significance of DNA PARylation.