DTX3L ubiquitin ligase ubiquitinates single-stranded nucleic acids.

Area of Science:

• Biochemistry
• Molecular Biology
• Genetics

Background:

• Ubiquitination traditionally modifies protein substrates via lysine residues.
• Emerging research reveals non-proteinaceous substrates for ubiquitination, such as ADP-ribose by DELTEX E3 ligases.

Purpose of the Study:

• To investigate the substrate repertoire of the DELTEX family member DTX3L.
• To elucidate the mechanism and functional implications of DTX3L-mediated ubiquitination of non-proteinaceous substrates.

Main Methods:

• Biochemical assays to determine catalytic activity and substrate specificity.
• Nuclear Magnetic Resonance (NMR) spectroscopy to analyze domain interactions.
• Deubiquitination assays to assess reversibility of the modification.

Main Results:

• DTX3L modifies single-stranded DNA and RNA, expanding the known non-proteinaceous substrates.
• The catalytically active fragment of DTX3L includes the C-terminal RING and DTC domains (DTX3L-RD).
• DTX3L-RD catalyzes ubiquitination at the 3'-end of nucleic acids, including double-stranded DNA with overhangs.
• Deubiquitinating enzymes can reverse this nucleic acid ubiquitination.
• The DTC domain binds single-stranded DNA and facilitates ubiquitin transfer.

Conclusions:

• DTX3L directly ubiquitinates nucleic acids, a novel biological function.
• This discovery opens avenues for understanding the roles of nucleic acid ubiquitination in cellular processes.
• The findings highlight the versatility of E3 ligases beyond protein modification.