Genetically Encoded Fluorescent Sensor for Poly-ADP-Ribose

Serebrovskaya, Ekaterina O.; Podvalnaya, Nadezda M; Dudenkova, Varvara V.; Ефремова, А.; Gurskaya, Nadya G.; Gorbachev, D.A.; Luzhin, Artem V; Kantidze, Omar L.; Zagaynova, Elena V.; Shram, S. I.; Lukyanov, Konstantin A.

doi:10.3390/ijms21145004

Cited by 10 publications

(18 citation statements)

References 24 publications

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“…In a recent study, Serebrovskaya et al (2020) developed FRET-based PAR sensors by fusing the WWE domain from RNF146 to the Torquoise2 and Venus fluorescent proteins (Serebrovskaya et al, 2020). These sensors detect PAR formation in live cells (Krastev et al, 2018;Serebrovskaya et al, 2020). However, the aforementioned approaches for PAR sensors have the following limitations.…”

Section: Previously Developed Par Sensorsmentioning

confidence: 99%

“…March 4, 2022 resonance energy transfer (FRET) (Serebrovskaya et al, 2020). However, these tools have some limitations: (1) they can only detect PAR accumulation in vitro (Furman et al, 2011), (2) they can only detect PAR accumulation on specific target proteins (Krastev et al, 2018), or (3) they have modest dynamic ranges (Serebrovskaya et al, 2020). Moreover, none of these sensors has been shown to be capable of detecting PAR accumulation in vivo.…”

Section: Challa Et Al (Kraus)mentioning

confidence: 99%

“…To overcome this limitation, several split-protein reassembly approaches have been applied to PAR detection. With this approach, non-functional fragments of a split-fluorescent protein or luciferase are induced to reassemble through the direct interaction of fused ADPR binding domains (Furman et al, 2011; Krastev et al, 2018; Lee et al, 2021; Serebrovskaya et al, 2020). These include the PBZ modules of aprataxin PNK-like factor (APLF) (Ahel et al, 2008) attached to each half of split firefly luciferase (split-Fluc) (Furman et al, 2011), PBZ modules with split Venus GFP (Krastev et al, 2018), and WWE domains with Turquoise and Venus, allowing for Förster resonance energy transfer (FRET) (Serebrovskaya et al, 2020).…”

Section: Introductionmentioning

confidence: 99%

“…With this approach, non-functional fragments of a split-fluorescent protein or luciferase are induced to reassemble through the direct interaction of fused ADPR binding domains (Furman et al, 2011; Krastev et al, 2018; Lee et al, 2021; Serebrovskaya et al, 2020). These include the PBZ modules of aprataxin PNK-like factor (APLF) (Ahel et al, 2008) attached to each half of split firefly luciferase (split-Fluc) (Furman et al, 2011), PBZ modules with split Venus GFP (Krastev et al, 2018), and WWE domains with Turquoise and Venus, allowing for Förster resonance energy transfer (FRET) (Serebrovskaya et al, 2020). However, these tools have some limitations: (1) they can only detect PAR accumulation in vitro (Furman et al, 2011), (2) they can only detect PAR accumulation on specific target proteins (Krastev et al, 2018), or (3) they have modest dynamic ranges (Serebrovskaya et al, 2020).…”

Section: Introductionmentioning

confidence: 99%

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Development and Characterization of PAR-Trackers: New Tools for Detecting Poly(ADP-ribose) In Vitro and In Vivo

Kraus

Challa

Ryu

et al. 2021

Preprint

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ADP-ribosylation (ADPRylation) is a reversible post-translation modification resulting in the covalent attachment of ADP-ribose (ADPR) moieties on substrate proteins. Naturally-occurring protein motifs and domains, including WWEs, PBZs, and macrodomains, act as 'readers' for protein-linked ADPR. Although recombinant, antibody-like ADPR detection reagents containing these readers have facilitated the detection of ADPR, they are limited in their ability to capture the dynamic nature of ADPRylation. Herein, we describe and characterize a set of poly(ADP-ribose) (PAR) Trackers (PAR-Ts) - optimized dimerization-dependent or split-protein reassembly PAR sensors in which a naturally occurring PAR binding domain, WWE, was fused to both halves of dimerization-dependent GFP (ddGFP) or split Nano Luciferase (NanoLuc), respectively. We demonstrate that these new tools allow the detection and quantification of PAR levels in extracts, living cells, and living tissues with greater sensitivity, as well as temporal and spatial precision. Importantly, these sensors detect changes in cellular ADPR levels in response to physiological cues (e.g., hormone-dependent induction of adipogenesis without DNA damage), as well as xenograft tumor tissues in living mice. Our results indicate that PAR Trackers have broad utility for detecting ADPR in many different experimental and biological systems.

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Section: Previously Developed Par Sensorsmentioning

confidence: 99%

Section: Challa Et Al (Kraus)mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Development and Characterization of PAR-Trackers: New Tools for Detecting Poly(ADP-ribose) In Vitro and In Vivo

Kraus

Challa

Ryu

et al. 2021

Preprint

View full text Add to dashboard Cite

show abstract

“…To overcome this limitation, several split-protein reassembly approaches have been applied to PAR detection. With this approach, nonfunctional fragments of a split-fluorescent protein or luciferase are induced to reassemble through the direct interaction of fused ARBDs ( Furman et al, 2011 ; Krastev et al, 2018 ; Lee et al, 2021 ; Serebrovskaya et al, 2020 ). These include the PBZ modules of aprataxin PNK-like factor (APLF) ( Ahel et al, 2008 ) attached to each half of split firefly luciferase (split-Fluc) ( Furman et al, 2011 ), PBZ modules with split Venus GFP ( Krastev et al, 2018 ), and WWE domains with Turquoise and Venus, allowing for Förster resonance energy transfer (FRET) ( Serebrovskaya et al, 2020 ).…”

Section: Introductionmentioning

confidence: 99%

Development and characterization of new tools for detecting poly(ADP-ribose) in vitro and in vivo

Challa

Ryu

Whitaker

et al. 2022

eLife

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ADP-ribosylation (ADPRylation) is a reversible post-translation modification resulting in the covalent attachment of ADP-ribose (ADPR) moieties on substrate proteins. Naturally occurring protein motifs and domains, including WWEs, PBZs, and macrodomains, act as ‘readers’ for protein-linked ADPR. Although recombinant, antibody-like ADPR detection reagents containing these readers have facilitated the detection of ADPR, they are limited in their ability to capture the dynamic nature of ADPRylation. Herein, we describe and characterize a set of poly(ADP-ribose) (PAR) Trackers (PAR-Ts)—optimized dimerization-dependent or split-protein reassembly PAR sensors in which a naturally occurring PAR binding domain, WWE, was fused to both halves of dimerization-dependent GFP (ddGFP) or split Nano Luciferase (NanoLuc), respectively. We demonstrate that these new tools allow the detection and quantification of PAR levels in extracts, living cells, and living tissues with greater sensitivity, as well as temporal and spatial precision. Importantly, these sensors detect changes in cellular ADPR levels in response to physiological cues (e.g., hormone-dependent induction of adipogenesis without DNA damage), as well as xenograft tumor tissues in living mice. Our results indicate that PAR Trackers have broad utility for detecting ADPR in many different experimental and biological systems.

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Poly‐ADP‐ribosylation dynamics, signaling, and analysis

Al‐Rahahleh,

Sobol

2024

Environ and Mol Mutagen

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ADP‐ribosylation is a reversible post‐translational modification that plays a role as a signaling mechanism in various cellular processes. This modification is characterized by its structural diversity, highly dynamic nature, and short half‐life. Hence, it is tightly regulated at many levels by cellular factors that fine‐tune its formation, downstream signaling, and degradation that together impacts cellular outcomes. Poly‐ADP‐ribosylation is an essential signaling mechanism in the DNA damage response that mediates the recruitment of DNA repair factors to sites of DNA damage via their poly‐ADP‐ribose (PAR)‐binding domains (PBDs). PAR readers, encoding PBDs, convey the PAR signal to mediate cellular outcomes that in some cases can be dictated by PAR structural diversity. Several PBD families have been identified, each with variable PAR‐binding affinity and specificity, that also recognize and bind to distinct parts of the PAR chain. PARylation signaling has emerged as an attractive target for the treatment of specific cancer types, as the inhibition of PAR formation or degradation can selectively eliminate cancer cells with specific DNA repair defects and can enhance radiation or chemotherapy response. In this review, we summarize the key players of poly‐ADP‐ribosylation and its regulation and highlight PBDs as tools for studying PARylation dynamics and the expanding potential to target PARylation signaling in cancer treatment.

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Genetically Encoded Fluorescent Sensor for Poly-ADP-Ribose

Cited by 10 publications

References 24 publications

Development and Characterization of PAR-Trackers: New Tools for Detecting Poly(ADP-ribose) In Vitro and In Vivo

Development and Characterization of PAR-Trackers: New Tools for Detecting Poly(ADP-ribose) In Vitro and In Vivo

Development and characterization of new tools for detecting poly(ADP-ribose) in vitro and in vivo

Poly‐ADP‐ribosylation dynamics, signaling, and analysis

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