Engineered allostery in light-regulated LOV-Turbo enables precise spatiotemporal control of proximity labeling in living cells.

The incorporation of light-responsive domains into engineered proteins has enabled control of protein localization, interactions and function with light. We integrated optogenetic control into proximity labeling, a cornerstone technique for high-resolution proteomic mapping of organelles and interactomes in living cells. Through structure-guided screening and directed evolution, we installed the light-sensitive LOV domain into the proximity labeling enzyme TurboID to rapidly and reversibly control its labeling activity with low-power blue light. 'LOV-Turbo' works in multiple contexts and dramatically reduces background in biotin-rich environments such as neurons. We used LOV-Turbo for pulse-chase labeling to discover proteins that traffic between endoplasmic reticulum, nuclear and mitochondrial compartments under cellular stress. We also showed that instead of external light, LOV-Turbo can be activated by bioluminescence resonance energy transfer from luciferase, enabling interaction-dependent proximity labeling. Overall, LOV-Turbo increases the spatial and temporal precision of proximity labeling, expanding the scope of experimental questions that can be addressed with proximity labeling.

Binding of USP4 to cortactin enhances cell migration in HCT116 human colon cancer cells.

Ubiquitin-specific protease 4 (USP4) is highly overexpressed in colon cancer and acts as a potent protooncogenic protein by deubiquitinating ß-catenin. However, its prominent roles in tumor formation and migration in cancer cells are not fully understood by its deubiquitinating enzyme (DUB) activity on ß-catenin. Thus, we investigated an additional role of USP4 in cancer. In this study, we identified cortactin (CTTN), an actin-binding protein involved in the regulation of cytoskeleton dynamics and a potential prognostic marker for cancers, as a new cellular interacting partner of USP4 from proximal labeling of HCT116 cells. Additionally, the role of USP4 in CTTN activation and promotion of cell dynamics and migration was investigated in HCT116 cells. We confirmed that interacting of USP4 with CTTN increased cell movement. This finding was supported by the fact that USP4 overexpression in HCT116 cells with reduced expression of CTTN was insufficient to promote cell migration. Additionally, we observed that USP4 overexpression led to a significant increase in CTTN phosphorylation, which is a requisite mechanism for cell migration, by regulating Src/focal adhesion kinase (FAK) binding to CTTN and its activation. Our results suggest that USP4 plays a dual role in cancer progression, including stabilization of ß-catenin as a DUB and interaction with CTTN to promote cell dynamics by inducing CTTN phosphorylation. Therefore, this study demonstrates that USP4 is important for cancer progression and is a good target for treating or preventing cancer.

Enhanced Microstructural Uniformity in Sulfuric-Acid-Treated Poly(3,4-Ethylenedioxythiophene):Poly(Styrene Sulfonate) Films Using Raman Map Analysis.

Poly(3,4-ethylenedioxythiophene):poly(styrene sulfonate) (PEDOT:PSS) films have emerged as potential alternatives to indium-tin oxide as transparent electrodes in optoelectronic devices because of their superior transparency, flexibility, and chemical doping stability. However, pristine PEDOT:PSS films show low conductivities because the insulating PSS-rich domains isolate the conductive PEDOT-rich domains. In this study, the conductivities and corresponding spatially resolved Raman properties of PEDOT:PSS thin films treated with various concentrations of H2SO4 are presented. After the PEDOT:PSS films are treated with the H2SO4 solutions, their electrical conductivities are significantly improved from 0.5 (nontreated) to 4358 S cm-1 (100% v/v). Raman heat maps of the peak shifts and widths of the Cα═Cß stretching mode are constructed. A blueshift and width decrease of the Cα═Cß Raman mode in PEDOT are uniformly observed in the entire measurement area (20 × 20 µm2), indicating that microstructural transitions are successfully accomplished across the area from the coiled to linear conformation and high crystallinity upon H2SO4 treatment. Thus, it is proved that comprehensive Raman map analysis can be easily utilized to clarify microstructural properties distributed in large areas induced by various dopants. These results also offer valuable insights for evaluating and optimizing the performance of other conductive thin films.

Structure and stability of polydiacetylene membrane systems: Molecular dynamics simulation studies.

We have performed full atomistic molecular dynamics (MD) simulations to investigate structure and stability of bilayer membrane systems consisting of monomeric or polymeric 10,12-pentacosadiynoic acid (PCDA) units connected with lysine groups by amide bonds. The PCDA monomer molecules show a twisted three-rod-domain structure with two kinks but upon polymerization, they possess more elongated conformation. The resulting polydiacetylene (PDA) membrane systems have stable membrane structures at room temperature, which is similar to biological lipid bilayer membranes and maintain their gel-like membrane integrity even up to as high as 370 K. Structural properties such as area per monomer, membrane thickness, density profile, 2D pair distribution function, and orientational correlation function are also calculated to understand the membrane structure and check its stability upon thermal fluctuation with atomistic resolution. This study is expected to provide the understanding about PDA membrane systems in atomistic details as well as significant insights into designing new novel PDA sensors.

Recent Strategies to Develop Innovative Photosensitizers for Enhanced Photodynamic Therapy.

This review presents a robust strategy to design photosensitizers (PSs) for various species. Photodynamic therapy (PDT) is a photochemical-based treatment approach that involves the use of light combined with a light-activated chemical, referred to as a PS. Attractively, PDT is one of the alternatives to conventional cancer treatment due to its noninvasive nature, high cure rates, and low side effects. PSs play an important factor in photoinduced reactive oxygen species (ROS) generation. Although the concept of photosensitizer-based photodynamic therapy has been widely adopted for clinical trials and bioimaging, until now, to our surprise, there has been no relevant review article on rational designs of organic PSs for PDT. Furthermore, most of published review articles in PDT focused on nanomaterials and nanotechnology based on traditional PSs. Therefore, this review aimed at reporting recent strategies to develop innovative organic photosensitizers for enhanced photodynamic therapy, with each example described in detail instead of providing only a general overview, as is typically done in previous reviews of PDT, to provide intuitive, vivid, and specific insights to the readers.

The structure of human EXD2 reveals a chimeric 3' to 5' exonuclease domain that discriminates substrates via metal coordination.

EXD2 (3'-5' exonuclease domain-containing protein 2) is an essential protein with a conserved DEDDy superfamily 3'-5' exonuclease domain. Recent research suggests that EXD2 has two potential functions: as a component of the DNA double-strand break repair machinery and as a ribonuclease for the regulation of mitochondrial translation. Herein, electron microscope imaging analysis and proximity labeling revealed that EXD2 is anchored to the mitochondrial outer membrane through a conserved N-terminal transmembrane domain, while the C-terminal region is cytosolic. Crystal structures of the exonuclease domain in complex with Mn2+/Mg2+ revealed a domain-swapped dimer in which the central α5-α7 helices are mutually crossed over, resulting in chimeric active sites. Additionally, the C-terminal segments absent in other DnaQ family exonucleases enclose the central chimeric active sites. Combined structural and biochemical analyses demonstrated that the unusual dimeric organization stabilizes the active site, facilitates discrimination between DNA and RNA substrates based on divalent cation coordination and generates a positively charged groove that binds substrates.

NSUN2 introduces 5-methylcytosines in mammalian mitochondrial tRNAs.

Expression of human mitochondrial DNA is indispensable for proper function of the oxidative phosphorylation machinery. The mitochondrial genome encodes 22 tRNAs, 2 rRNAs and 11 mRNAs and their post-transcriptional modification constitutes one of the key regulatory steps during mitochondrial gene expression. Cytosine-5 methylation (m5C) has been detected in mitochondrial transcriptome, however its biogenesis has not been investigated in details. Mammalian NOP2/Sun RNA Methyltransferase Family Member 2 (NSUN2) has been characterized as an RNA methyltransferase introducing m5C in nuclear-encoded tRNAs, mRNAs and microRNAs and associated with cell proliferation and differentiation, with pathogenic variants in NSUN2 being linked to neurodevelopmental disorders. Here we employ spatially restricted proximity labelling and immunodetection to demonstrate that NSUN2 is imported into the matrix of mammalian mitochondria. Using three genetic models for NSUN2 inactivation-knockout mice, patient-derived fibroblasts and CRISPR/Cas9 knockout in human cells-we show that NSUN2 is necessary for the generation of m5C at positions 48, 49 and 50 of several mammalian mitochondrial tRNAs. Finally, we show that inactivation of NSUN2 does not have a profound effect on mitochondrial tRNA stability and oxidative phosphorylation in differentiated cells. We discuss the importance of the newly discovered function of NSUN2 in the context of human disease.

Hypoxia-Activated Adipose Mesenchymal Stem Cells Prevents Irradiation-Induced Salivary Hypofunction by Enhanced Paracrine Effect Through Fibroblast Growth Factor 10.

To explore the effects and mechanisms of paracrine factors secreted from human adipose mesenchymal stem cell (hAdMSCs) that are activated by hypoxia on radioprotection against irradiation-induced salivary hypofunction in subjects undergoing radiotherapy for head and neck cancers. An organotypic spheroid coculture model to mimic irradiation (IR)-induced salivary hypofunction was set up for in vitro experiments. Human parotid gland epithelial cells were organized to form three-dimensional (3D) acinus-like spheroids on growth factor reduced -Matrigel. Cellular, structural, and functional damage following IR were examined after cells were cocultured with hAdMSCs preconditioned with either normoxia (hAdMSCNMX ) or hypoxia (hAdMSCHPX ). A key paracrine factor secreted by hAdMSCsHPX was identified by high-throughput microarray-based enzyme-linked immunosorbent assay. Molecular mechanisms and signaling pathways on radioprotection were explored. Therapeutic effects of hAdMSCsHPX were evaluated after in vivo transplant into mice with IR-induced salivary hypofunction. In our 3D coculture experiment, hAdMSCsHPX significantly enhanced radioresistance of spheroidal human parotid epithelial cells, and led to greater preservation of salivary epithelial integrity and acinar secretory function relative to hAdMSCsNMX . Coculture with hAdMSCsHPX promoted FGFR expression and suppressed FGFR diminished antiapoptotic activity of hAdMSCsHPX . Among FGFR-binding secreted factors, we found that fibroblast growth factor 10 (FGF10) contributed to therapeutic effects of hAdMSCsHPX by enhancing antiapoptotic effect, which was dependent on FGFR-PI3K signaling. An in vivo transplant of hAdMSCsHPX into irradiated salivary glands of mice reversed IR-induced salivary hypofunction where hAdMSC-released FGF10 contributed to tissue remodeling. Our results suggest that hAdMSCsHPX protect salivary glands from IR-induced apoptosis and preserve acinar structure and functions by activation of FGFR-PI3K signaling via actions of hAdMSC-secreted factors, including FGF10. Stem Cells 2018;36:1020-1032.

Supramolecular photosensitizers rejuvenate photodynamic therapy.

Owing to its spatiotemporal selectivity and noninvasive nature, photodynamic therapy (PDT) has become a clinically promising approach for the treatment of a wide range of cancers and other diseases. However, the full potential of PDT has not been achieved thus far as a consequence of the lack of optimal photosensitizers (PSs) and/or smart transport/activation strategies. These problems, which unfortunately lie at the core of the PDT paradigm, include the oxygen reliance limits, the effect of PDT on hypoxic tumors, limitations of light penetration, and undesired skin photosensitization induced by "always on" PSs. Recently, supramolecular approaches, which rely on the use of non-covalent interactions to construct biomedical active materials, have become suitable methods for developing innovative PSs. Non-covalent interactions enable supramolecular PSs to have sensitive and controllable photoactivities, important elements needed to maximize photodynamic effects and minimize side effects. In addition, versatile supramolecular PS-assemblies can be designed so that PDT occurs synergistically with other therapeutic modalities, e.g., photothermal therapy, leading to a potential improvement of therapeutic effectiveness. In this review, recent progress made in the development of supramolecular PSs for rejuvenating PDT will be presented. Importantly, this discussion also provides a view of future advances that will likely be made in this area and their potential clinical applications.

Self-immolative colorimetric, fluorescent and chemiluminescent chemosensors.

Self-immolative chemistry features a cascade of disassembly reactions in response to external stimuli, which provides great opportunities to design new self-immolative chemosensors with advanced performance and/or functions. Self-immolative spacers in these chemosensors not only facilitate the linkage of designed triggers to various chromophores or fluorophores, but can also be used to solve inherent problems associated with native chemosensors, such as low reactivities, poor stabilities and slow response times. Their capacity for stimuli-responsive release through operation of a self-immolative reaction further enables integration of sophisticated functions into chemosensors, including signal amplification, enzyme activity localization, and drug monitoring. Significant advances have been made in the field of self-immolative chemosensors, leading to intriguing applications to sensitive detection of analytes, bioimaging and cancer theranostics. This tutorial review summarizes this recent progress with a focus on their design strategies and sensing mechanisms.

Architecture Mapping of the Inner Mitochondrial Membrane Proteome by Chemical Tools in Live Cells.

The inner mitochondrial membrane (IMM) proteome plays a central role in maintaining mitochondrial physiology and cellular metabolism. Various important biochemical reactions such as oxidative phosphorylation, metabolite production, and mitochondrial biogenesis are conducted by the IMM proteome, and mitochondria-targeted therapeutics have been developed for IMM proteins, which is deeply related for various human metabolic diseases including cancer and neurodegenerative diseases. However, the membrane topology of the IMM proteome remains largely unclear because of the lack of methods to evaluate it in live cells in a high-throughput manner. In this article, we reveal the in vivo topological direction of 135 IMM proteins, using an in situ-generated radical probe with genetically targeted peroxidase (APEX). Owing to the short lifetime of phenoxyl radicals generated in situ by submitochondrial targeted APEX and the impermeability of the IMM to small molecules, the solvent-exposed tyrosine residues of both the matrix and intermembrane space (IMS) sides of IMM proteins were exclusively labeled with the radical probe in live cells by Matrix-APEX and IMS-APEX, respectively and identified by mass spectrometry. From this analysis, we confirmed 58 IMM protein topologies and we could determine the topological direction of 77 IMM proteins whose topology at the IMM has not been fully characterized. We also found several IMM proteins (e.g., LETM1 and OXA1) whose topological information should be revised on the basis of our results. Overall, our identification of structural information on the mitochondrial inner-membrane proteome can provide valuable insights for the architecture and connectome of the IMM proteome in live cells.

Specific Inhibition of the Distribution of Lobeglitazone to the Liver by Atorvastatin in Rats: Evidence for a Rat Organic Anion Transporting Polypeptide 1B2-Mediated Interaction in Hepatic Transport.

Cytochrome P450 enzymes and human organic anion transporting polypeptide (OATP) 1B1 are reported to be involved in the pharmacokinetics of lobeglitazone (LB), a new peroxisome proliferator-activated receptor γ agonist. Atorvastatin (ATV), a substrate for CYP3A and human OATP1B1, is likely to be coadministered with LB in patients with the metabolic syndrome. We report herein on a study of potential interactions between LB and ATV in rats. When LB was administered intravenously with ATV, the systemic clearance and volume of distribution at steady state for LB remained unchanged (2.67 ± 0.63 ml/min per kg and 289 ± 20 ml/kg, respectively), compared with that of LB without ATV (2.34 ± 0.37 ml/min per kg and 271 ± 20 ml/kg, respectively). Although the tissue-to-plasma partition coefficient (Kp) of LB was not affected by ATV in most major tissues, the liver Kp for LB was decreased by ATV coadministration. Steady-state liver Kp values for three levels of LB were significantly decreased as a result of ATV coadministration. LB uptake was inhibited by ATV in rat OATP1B2-overexpressing Madin-Darby canine kidney cells and in isolated rat hepatocytes in vitro. After incorporating the kinetic parameters for the in vitro studies into a physiologically based pharmacokinetics model, the characteristics of LB distribution to the liver were consistent with the findings of the in vivo study. It thus appears that the distribution of LB to the liver is mediated by the hepatic uptake of transporters such as rat OATP1B2, and carrier-mediated transport is involved in the liver-specific drug-drug interaction between LB and ATV in vivo.

HIV Peptide-Mediated Binding Behaviors of Nanoparticles on a Lipid Membrane.

The bioinspired design of ligands for nanoparticle coating with remarkable precision in controlling anisotropic connectivity and with universal binding efficiency to the membrane has made a great impact on nanoparticle self-assembly. We utilize the HIV-1-derived trans-activator of transcription peptide (TAT), a member of the cell-penetrating peptides, as a soft shell coating on gold nanoparticles (GNPs) and characterize TAT pepide-mediated binding behaviors of GNPs on the lipid membrane. Whereas the peptides enable GNPs to firmly attach to the membrane, the binding structures are driven by two electrostatic forces: the interparticle peptide repulsion and the peptide-membrane attraction. Although transmission electron microscopy images showed that the densities of membrane-embedded GNPs were almost equal, X-ray reflectivity revealed a significant difference in binding structures of GNPs along the surface normal upon the increase of charge densities (ϕ) of the membrane. In particular, GNPs were densely suspended at ϕ = 70% while they adopted an additional well-defined layer underneath the membrane at ϕ = 100%, in addition to a translocation of the initially bound particles into the membrane. The observed behaviors of GNPs manifest a 3D to 2D transformation of the self-assembled structures from the diffused state to the closely packed state with the increase in the charge density of the membrane. The present study also provides insights on the binding mechanisms of the cell-penetrating peptide-coated nanoparticles to the lipid membranes, which is a common theme of delivery systems in pharmaceutical research.

Comparison of UTI antibiograms stratified by ED patient disposition.

OBJECTIVE: Institutional antibiograms guide Emergency Department (ED) clinicians' empiric antibiotic selection. For this study, we created and compared antibiograms of ED patients stratified by disposition (admitted or discharged). METHODS: We conducted a cross-sectional study at two hospitals for 2014, comparing antibiograms limited to Escherichia coli urinary tract infections. Study-Specific Antibiograms, created for the study, excluded polymicrobial samples and multiple cultures from the same patient. Study-Specific Antibiograms were arranged by patient disposition: admitted (IP-Only) vs discharged from the ED (ED-Only). Antibiogram data were presented as average antibiotic sensitivities with 95% confidence intervals and demographic data as medians with interquartile ranges. Sensitivities between Study-Specific Antibiograms were compared by Fisher's Exact Test, alpha=0.05, 2 tails. RESULTS: For Hospital A, 13 antibiotics were compared between Study-Specific ED-Only (n=313) vs IP-Only (n=244). We found that sensitivities to all four antibiotics appropriate for empiric outpatient therapy by Infectious Disease Society of America guidelines were significantly (p<0.0001) higher in the ED-Only compared to IP-Only groups: ciprofloxacin 80% (76-90%) vs 60% (53-69%), levofloxacin 81% (77-91%) vs 63% (57-72%), nitrofurantoin 75% (70-84%) vs 51% (44-58%), and trimethoprim/sulfamethoxazole 73% (68-82%) vs 58% (52-67%). For Hospital B, 14 antibiotics were compared between Study-Specific ED-Only (n=256) and IP-Only (n=168). Two out of the five appropriate empiric outpatient antibiotics had significantly (p<0.0001) higher sensitivities for ED-Only compared to IP-Only: ciprofloxacin 87% (83-91%) vs 71% (64-78%) and levofloxacin 86% (82-91%) vs 71% (65-78%). CONCLUSIONS: We found higher antibiotic sensitivities in ED-Only than the IP-Only Study-Specific Antibiograms. Our Study-Specific Antibiograms offer an alternative guide for antibiotic selection in the ED.

A Dual Colorimetric/Fluorescence System for Determining pH Based on the Nucleophilic Addition Reaction of an o-Hydroxymerocyanine Dye.

Owing to their ability to monitor pH in a precise and rapid manner, optical probes have widely been developed for biological and nonbiological applications. The strategies thus far employed to determine pH rely on two types of processes including reversible protonation of amine nitrogen atoms and deprotonation of phenols. We have developed a novel dual, colorimetric/fluorescence system for determining the pH of a solution. This system utilizes an o-hydroxymerocyanine dye that undergoes a nucleophilic addition reaction that subsequently causes reversible structural changes interconverting a merocyanine to a spirocyanine and a spirocyanine to a spiropyran. It was demonstrated that the dye can be employed to measure the pH of solutions in the 2.5-5.75 and 9.6-11.8 ranges with color changes from yellow to dark blue and then to lavender. Moreover, the fluorescence response associated with the spirocyanine-spiropyran transformation of the dye occurring in alkaline solutions provides a precise method.

A review: the trend of progress about pH probes in cell application in recent years.

Intracellular pH values are some of the most important factors that govern biological processes and the acid-base homeostasis in cells, body fluids and organs sustains the normal operations of the body. Subcellular organelles including the acidic lysosomes and the alkalescent mitochondria undergo various processes such as intracellular digestion, ATP production and apoptosis. Due to their precise imaging capabilities, fluorescent probes have attracted great attention for the illustration of pH modulated processes. Furthermore, based on the unique acidic extracellular environment of acidic lysosomes, fluorescent probes can specifically be activated in cancer cells or tumors. In this review, recently reported lysosome and mitochondria specific pH imaging probes as well as pH-activatable cancer cell-targetable probes have been discussed.

Electronic relaxation dynamics of PCDA-PDA studied by transient absorption spectroscopy.

Photo-curable polymers originating from 10,12-pentacosadiynoic acid (PCDA-PDA) are commonly used polydiacetylenes (PDAs). PCDA-PDA exhibits thermochromic properties undergoing a unique colorimetric transition from blue to red as the temperature is increased from low to high. In this work, we have carefully studied the temperature-dependent optical properties of PCDA-PDA by using UV-visible absorption, FTIR, Raman, and transient absorption (TA) spectroscopy in combination with quantum chemical calculations. Temperature-dependent UV-visible absorption spectra indicate that PCDA-PDA exhibits reversible thermochromic properties up to 60 °C and its thermochromic properties become irreversible above 60 °C. Such distinct thermochromic properties are also manifested in TA signals so that the electronically excited PCDA-PDA relaxes to the ground state via an intermediate state at 20 °C (blue form) but it relaxes directly back to the ground state at 80 °C (red form). The electronic relaxation dynamics of PCDA-PDA are comprehensively analyzed based on different kinetic models by using the global fitting analysis method. The intermediate state in the blue form of PCDA-PDA is clearly found to be responsible for fluorescence quenching. FTIR and Raman spectroscopy and quantum chemical calculations confirm that the H-bonds between the carboxylic acid groups in PCDA-PDA are broken at high temperatures leading to an irreversible structural change of PCDA-PDA.

Fluorescent and colorimetric chemosensors for pyrophosphate.

Pyrophosphate anions play key roles in various biological and chemical processes. During the last few years, many exciting results have emerged regarding the development of fluorescent and colorimetric sensors for this biologically important species. In this review, we will cover the fluorescent and colorimetric chemosensors developed for the detection of pyrophosphate (PPi) since 2010.

Super-resolution proximity labeling with enhanced direct identification of biotinylation sites.

Promiscuous labeling enzymes, such as APEX2 or TurboID, are commonly used in in situ biotinylation studies of subcellular proteomes or protein-protein interactions. Although the conventional approach of enriching biotinylated proteins is widely implemented, in-depth identification of specific biotinylation sites remains challenging, and current approaches are technically demanding with low yields. A novel method to systematically identify specific biotinylation sites for LC-MS analysis followed by proximity labeling showed excellent performance compared with that of related approaches in terms of identification depth with high enrichment power. The systematic identification of biotinylation sites enabled a simpler and more efficient experimental design to identify subcellular localized proteins within membranous organelles. Applying this method to the processing body (PB), a non-membranous organelle, successfully allowed unbiased identification of PB core proteins, including novel candidates. We anticipate that our newly developed method will replace the conventional method for identifying biotinylated proteins labeled by promiscuous labeling enzymes.

Charge Transfer-Promoted Excited State of a Heavy-Atom-Free Photosensitizer for Efficient Application of Mitochondria-Targeted Fluorescence Imaging and Hypoxia Photodynamic Therapy.

Conventional photosensitizers (PSs) used in photodynamic therapy (PDT) have shown preliminary success; however, they are often associated with several limitations including potential dark toxicity in healthy tissues, limited efficacy under acidic and hypoxic conditions, suboptimal fluorescence imaging capabilities, and nonspecific targeting during treatment. In response to these challenges, we developed a heavy-atom-free PS, denoted as Cz-SB, by incorporating ethyl carbazole into a thiophene-fused BODIPY core. A comprehensive investigation into the photophysical properties of Cz-SB was conducted through a synergistic approach involving experimental and computational investigations. The enhancement of intersystem crossing (kISC) and fluorescence emission (kfl) rate constants was achieved through a donor-acceptor pair-mediated charge transfer mechanism. Consequently, Cz-SB demonstrated remarkable efficiency in generating reactive oxygen species (ROS) under acidic and low-oxygen conditions, making it particularly effective for hypoxic cancer PDT. Furthermore, Cz-SB exhibited good biocompatibility, fluorescence imaging capabilities, and a high degree of localization within the mitochondria of living cells. We posit that Cz-SB holds substantial prospects as a versatile PS with innovative molecular design, representing a potential "one-for-all" solution in the realm of cancer phototheranostics.