Engineering fluorescent protein chromophores with an internal reference for high-fidelity ratiometric G4 imaging in living cells

Han, Jiao-Na; Zhong, Caijun; Ge, Mingmin; Kuang, Shi; Nie, Zhou

doi:10.1039/d3sc00022b

Cited by 12 publications

(4 citation statements)

References 70 publications

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“…Apoptosis and ferroptosis are two common mechanisms of cell death, and differentiation between them is essential for understanding cell biology and developing corresponding therapeutic strategies. − Apoptosis is typically accompanied by DNA degradation, whereas ferroptosis is caused by the disruption of the cellular redox balance (Figure a). − Apoptosis is characterized by nuclear changes, such as chromatin condensation, nuclear shrinkage, DNA fragmentation, and the formation of apoptotic bodies; in contrast, ferroptosis does not involve these nuclear changes. Therefore, we investigated whether SPP could track DNA damage in mitochondria and nuclei during apoptosis and ferroptosis.…”

Section: Resultsmentioning

confidence: 99%

Light-Driven Mitochondrion-to-Nucleus DNA Cascade Fluorescence Imaging and Enhanced Cancer Cell Photoablation

Xia,

Tang

et al. 2024

J. Am. Chem. Soc.

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Nucleic acids are mainly found in the mitochondria and nuclei of cells. Detecting nucleic acids in the mitochondrion and nucleus in cascade mode is crucial for understanding diverse biological processes. This study introduces a novel nucleic acidbased fluorescent styrene dye (SPP) that exhibits light-driven cascade migration from the mitochondrion to the nucleus. By introducing N-arylpyridine on one side of the styrene dye skeleton and a bis(2-ethylsulfanyl-ethy)-amino unit on the other side, we found that SPP exhibits excellent DNA specificity (16-fold, F DNA / F free ) and a stronger binding force to nuclear DNA (−5.09 kcal/ mol) than to mitochondrial DNA (−2.59 kcal/mol). SPP initially accumulates in the mitochondrion and then migrates to the nucleus within 10 s under light irradiation. By tracking the damage to nucleic acids in apoptotic cells, SPP allows the successful visualization of the differences between apoptosis and ferroptosis. Finally, a triphenylamine segment with photodynamic effects was incorporated into SPP to form a photosensitizer (MTPA-SPP), which targets the mitochondria for photosensitization and then migrates to the nucleus under light irradiation for enhanced photodynamic cancer cell treatment. This innovative nucleic acid-based fluorescent molecule with light-triggered mitochondrion-to-nucleus migration ability provides a feasible approach for the in situ identification of nucleic acids, monitoring of subcellular physiological events, and efficient photodynamic therapy.

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Section: Resultsmentioning

confidence: 99%

Light-Driven Mitochondrion-to-Nucleus DNA Cascade Fluorescence Imaging and Enhanced Cancer Cell Photoablation

Xia,

Tang

et al. 2024

J. Am. Chem. Soc.

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“…G-quadruplexes (G4s) are a cation-dependent (including K + , Na + , and Li + ) secondary structure that is usually formed by single-stranded G-rich sequence of nucleic acid, which present quite high stability and resistance to nuclease degradation. , Thanks to the development of extensive small molecular tools that selectively recognize G4 structures of nDNA, − the biological function of G4s in the nuclear genome has been widely studied, and it has been proven that G4s are closely related to genome transcription, recombination, and replication. , Unfortunately, although a mitochondrial genome can also form G4s, research based on mitochondrial G4s (mtG4s) is still limited due to the lack of suitable tools, − especially for mtG4s in sperms.…”

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confidence: 99%

Construction of Near-Infrared Probes with Remarkable Large Stokes Shift Based on a Novel Purine Platform for the Visualization of mtG4 Upregulation during Mitochondrial Disorder in Somatic Cells and Human Sperms

Yu,

Li,

Wang

et al. 2024

Anal. Chem.

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G-quadruplex structures within the nuclear genome (nG4) is an important regulatory factor, while the function of G4 in the mitochondrial genome (mtG4) still needs to be explored, especially in human sperms. To gain a better understanding of the relationship between mtG4 and mitochondrial function, it is crucial to develop excellent probes that can selectively visualize and track mtG4 in both somatic cells and sperms. Herein, based on our previous research on purine frameworks, we attempted for the first time to extend the conjugated structure from the C-8 site of purine skeleton and discovered that the purine derivative modified by the C-8 aldehyde group is an ideal platform for constructing near-infrared probes with extremely large Stokes shift (>220 nm). Compared with the compound substituted with methylpyridine (PAP), the molecule substituted with methylthiazole orange (PATO) showed better G4 recognition ability, including longer emission (∼720 nm), more significant fluorescent enhancement (∼67-fold), lower background, and excellent photostability. PATO exhibited a sensitive response to mtG4 variation in both somatic cells and human sperms. Most importantly, PATO helped us to discover that mtG4 was significantly increased in cells with mitochondrial respiratory chain damage caused by complex I inhibitors (6-OHDA and rotenone), as well as in human sperms that suffer from oxidative stress. Altogether, our study not only provides a novel ideal molecular platform for constructing high-performance probes but also develops an effective tool for studying the relationship between mtG4 and mitochondrial function in both somatic cells and human sperms.

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“…To date, a series of fluorescent molecules have been designed and developed as optical probes for the visualization of G4s. 12–16 However, most of them target the nucleus, and there are few probes, especially with two-photon fluorescence properties and fluorescence lifetime responses, that can specifically target cytoplasmic G4s, such as mitochondrial DNA-G4s and RNA-G4s. 17–19 We previously designed an organic small-molecule fluorescent probe, NBTE , which has been shown to have significantly distinguishable differences in fluorescence quantum yield ( Φ ) and fluorescence lifetime responses between DNA G4s and other DNA topologies.…”

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confidence: 99%

Detection and tracking of cytoplasmic G-quadruplexes in live cells

Rao,

Hao,

Liu

et al. 2023

Chem. Commun.

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Engineering fluorescent protein chromophores with an internal reference for high-fidelity ratiometric G4 imaging in living cells

Abstract: G-quadruplexes (G4s) are significant nucleic acid secondary structures formed by guanine-rich sequences. Many single-emission G4s fluorescent probes that are lighted up by inhibiting intramolecular rotation have been reported. However, they...

Cited by 12 publications

References 70 publications

Light-Driven Mitochondrion-to-Nucleus DNA Cascade Fluorescence Imaging and Enhanced Cancer Cell Photoablation

Light-Driven Mitochondrion-to-Nucleus DNA Cascade Fluorescence Imaging and Enhanced Cancer Cell Photoablation

Construction of Near-Infrared Probes with Remarkable Large Stokes Shift Based on a Novel Purine Platform for the Visualization of mtG4 Upregulation during Mitochondrial Disorder in Somatic Cells and Human Sperms

Detection and tracking of cytoplasmic G-quadruplexes in live cells

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