insight - Biochemistry - # Direct Ubiquitination of Single-Stranded Nucleic Acids by DTX3L Ubiquitin Ligase

DTX3L Ubiquitin Ligase Catalyzes Direct Ubiquitination of Single-Stranded Nucleic Acids

Q: How do the structural features and domain arrangements of DTX3L, compared to other Deltex family members, contribute to its unique ability to ubiquitinate single-stranded nucleic acids?

The unique ability of DTX3L to ubiquitinate single-stranded nucleic acids can be attributed to specific structural features and domain arrangements that distinguish it from other Deltex family members. DTX3L contains a distinct N-terminal region lacking WWE domains and proline-rich regions found in other Deltex family members. This N-terminal region of DTX3L contains putative domains that are structurally similar to K Homology (KH) domains and RNA recognition motifs (RRMs), which are known to bind single-stranded DNA (ssDNA) and RNA (ssRNA). This unique structural feature allows DTX3L to interact with and bind ssDNA and ssRNA, setting it apart from other Deltex family members. Furthermore, the arrangement of the RING and DELTEX C-terminal (DTC) domains in DTX3L is crucial for its ability to ubiquitinate single-stranded nucleic acids. The minimal catalytically competent fragment of DTX3L comprises the C-terminal RING and DTC domains (RD), which are essential for catalyzing the ubiquitination of ssDNA and ssRNA. The DTC domain of DTX3L contains specific residues, such as His707, Tyr719, and Glu733, which are critical for the formation of Ub-DNA and Ub-RNA products. These residues are conserved across the DTX family and are known to coordinate ADP-ribose (ADPr) binding, suggesting a shared mechanism for ubiquitination of ADPr and nucleic acids by DTX3L. In contrast, other Deltex family members may lack the specific structural motifs or catalytic residues required for binding and ubiquitination of single-stranded nucleic acids. The absence of certain structural features or key residues in the DTC domain of other Deltex family members could limit their ability to interact with and modify nucleic acids, highlighting the unique structural characteristics of DTX3L that enable its specific function in ubiquitinating ssDNA and ssRNA.

Q: Could the direct ubiquitination of single-stranded nucleic acids by DTX3L have broader implications in the regulation of RNA metabolism or the immune response to viral genetic material?

The direct ubiquitination of single-stranded nucleic acids by DTX3L could indeed have broader implications in the regulation of RNA metabolism and the immune response to viral genetic material. In terms of RNA metabolism, the ability of DTX3L to ubiquitinate single-stranded RNA (ssRNA) suggests a potential role in regulating RNA stability, processing, or turnover. Ubiquitination of RNA has been linked to various aspects of RNA metabolism, including RNA degradation, RNA quality control, and RNA processing. By ubiquitinating ssRNA, DTX3L could potentially mark specific RNA molecules for degradation or processing, influencing gene expression and cellular functions. This could contribute to the fine-tuning of RNA metabolism pathways and the maintenance of cellular homeostasis. In the context of the immune response to viral genetic material, the direct ubiquitination of viral ssRNA by DTX3L could play a critical role in antiviral defense mechanisms. Viruses often exploit host RNA metabolism pathways for their replication and survival. By targeting viral ssRNA for ubiquitination, DTX3L could mark viral genetic material for degradation or immune recognition, triggering antiviral responses and inhibiting viral replication. This process could be part of the host cell's innate immune response to viral infections, helping to eliminate viral RNA and restrict viral spread within the host. Overall, the direct ubiquitination of single-stranded nucleic acids by DTX3L opens up possibilities for its involvement in regulating RNA metabolism and modulating immune responses to viral infections, highlighting its potential significance in broader cellular processes and host defense mechanisms.

Core Concepts

DTX3L, a member of the Deltex ubiquitin ligase family, can directly ubiquitinate the 3' hydroxyl group of single-stranded DNA and RNA, expanding the repertoire of non-proteinaceous substrates that undergo ubiquitination.

Abstract

The content describes a novel function of the DTX3L ubiquitin ligase in directly ubiquitinating single-stranded nucleic acids (ssNAs), including single-stranded DNA (ssDNA) and single-stranded RNA (ssRNA). Key findings include:

DTX3L binds to ssDNA and ssRNA through its N-terminal KH-like domains, and the minimal C-terminal RING-DTC (RD) domains are sufficient to catalyze the ubiquitination reaction.
The ubiquitination occurs at the 3' hydroxyl group of the terminal nucleotide of the ssNA, forming a reversible ester linkage that can be cleaved by deubiquitinating enzymes.
The DTC domain of DTX3L binds to the ssNA and facilitates the transfer of ubiquitin from the RING-bound E2 enzyme to the 3' end. Conserved residues in the DTC domain, including His707, Tyr719, and Glu733, are critical for this catalytic activity.
While the DTC domain of DTX3L shares structural similarity with other Deltex family members, only DTX3L and DTX3 were found to be capable of ubiquitinating ssNAs, suggesting additional requirements beyond the catalytic residues.
The ubiquitination of ssNAs by DTX3L may have implications in cellular processes such as DNA damage repair and antiviral defense, where single-stranded nucleic acids are involved.

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Stats

The 3' hydroxyl group of the terminal nucleotide on single-stranded DNA and RNA is the site of ubiquitination by DTX3L.
Mutation of conserved residues His707, Tyr719, and Glu733 in the DTC domain of DTX3L abolishes or impairs the ubiquitination of single-stranded nucleic acids.
Only DTX3L and DTX3, but not other Deltex family members, were found to be capable of ubiquitinating single-stranded nucleic acids under the tested conditions.

Quotes

"DTX3L catalyses ubiquitination of the 3'-end of single-stranded DNA and RNA, as well as double-stranded DNA with a 3' overhang of two or more nucleotides."
"Mutation of the residues previously proposed to form the catalytic site for ubiquitination of ADPr abolished the ubiquitination of ssDNA, indicating a conserved site for this reaction."

Key Insights Distilled From

DTX3L ubiquitin ligase ubiquitinates single-stranded nucleic acids

by Dearlove,E. ... at www.biorxiv.org 04-02-2024

https://www.biorxiv.org/content/10.1101/2024.04.02.587769v1

Deeper Inquiries

What are the potential biological functions of DTX3L-mediated ubiquitination of single-stranded nucleic acids in cellular processes such as DNA damage repair and antiviral defense?

DTX3L-mediated ubiquitination of single-stranded nucleic acids could play crucial roles in cellular processes such as DNA damage repair and antiviral defense. In DNA damage repair, the ability of DTX3L to ubiquitinate single-stranded DNA (ssDNA) suggests a potential role in recognizing and marking damaged DNA for repair. This could involve targeting DNA lesions or breaks with 3' overhangs, which are common in DNA damage sites, for ubiquitination. By ubiquitinating these damaged DNA regions, DTX3L could facilitate the recruitment of repair factors to the site, aiding in the efficient and accurate repair of DNA damage.
In the context of antiviral defense, the direct ubiquitination of single-stranded RNA (ssRNA) by DTX3L could have implications in the immune response to viral genetic material. Viruses often contain ssRNA as part of their genetic material, and the ability of DTX3L to ubiquitinate ssRNA suggests a potential mechanism for targeting and marking viral RNA for degradation or immune recognition. This process could be part of the host cell's defense mechanism against viral infections, helping to eliminate viral genetic material and inhibit viral replication.
Overall, the ubiquitination of single-stranded nucleic acids by DTX3L could contribute to maintaining genomic integrity, regulating immune responses to viral infections, and potentially influencing various cellular processes involved in DNA damage repair and antiviral defense mechanisms.

How do the structural features and domain arrangements of DTX3L, compared to other Deltex family members, contribute to its unique ability to ubiquitinate single-stranded nucleic acids?

The unique ability of DTX3L to ubiquitinate single-stranded nucleic acids can be attributed to specific structural features and domain arrangements that distinguish it from other Deltex family members. DTX3L contains a distinct N-terminal region lacking WWE domains and proline-rich regions found in other Deltex family members. This N-terminal region of DTX3L contains putative domains that are structurally similar to K Homology (KH) domains and RNA recognition motifs (RRMs), which are known to bind single-stranded DNA (ssDNA) and RNA (ssRNA). This unique structural feature allows DTX3L to interact with and bind ssDNA and ssRNA, setting it apart from other Deltex family members.
Furthermore, the arrangement of the RING and DELTEX C-terminal (DTC) domains in DTX3L is crucial for its ability to ubiquitinate single-stranded nucleic acids. The minimal catalytically competent fragment of DTX3L comprises the C-terminal RING and DTC domains (RD), which are essential for catalyzing the ubiquitination of ssDNA and ssRNA. The DTC domain of DTX3L contains specific residues, such as His707, Tyr719, and Glu733, which are critical for the formation of Ub-DNA and Ub-RNA products. These residues are conserved across the DTX family and are known to coordinate ADP-ribose (ADPr) binding, suggesting a shared mechanism for ubiquitination of ADPr and nucleic acids by DTX3L.
In contrast, other Deltex family members may lack the specific structural motifs or catalytic residues required for binding and ubiquitination of single-stranded nucleic acids. The absence of certain structural features or key residues in the DTC domain of other Deltex family members could limit their ability to interact with and modify nucleic acids, highlighting the unique structural characteristics of DTX3L that enable its specific function in ubiquitinating ssDNA and ssRNA.

Could the direct ubiquitination of single-stranded nucleic acids by DTX3L have broader implications in the regulation of RNA metabolism or the immune response to viral genetic material?

The direct ubiquitination of single-stranded nucleic acids by DTX3L could indeed have broader implications in the regulation of RNA metabolism and the immune response to viral genetic material.
In terms of RNA metabolism, the ability of DTX3L to ubiquitinate single-stranded RNA (ssRNA) suggests a potential role in regulating RNA stability, processing, or turnover. Ubiquitination of RNA has been linked to various aspects of RNA metabolism, including RNA degradation, RNA quality control, and RNA processing. By ubiquitinating ssRNA, DTX3L could potentially mark specific RNA molecules for degradation or processing, influencing gene expression and cellular functions. This could contribute to the fine-tuning of RNA metabolism pathways and the maintenance of cellular homeostasis.
In the context of the immune response to viral genetic material, the direct ubiquitination of viral ssRNA by DTX3L could play a critical role in antiviral defense mechanisms. Viruses often exploit host RNA metabolism pathways for their replication and survival. By targeting viral ssRNA for ubiquitination, DTX3L could mark viral genetic material for degradation or immune recognition, triggering antiviral responses and inhibiting viral replication. This process could be part of the host cell's innate immune response to viral infections, helping to eliminate viral RNA and restrict viral spread within the host.
Overall, the direct ubiquitination of single-stranded nucleic acids by DTX3L opens up possibilities for its involvement in regulating RNA metabolism and modulating immune responses to viral infections, highlighting its potential significance in broader cellular processes and host defense mechanisms.