Living HeLa Cells Research Articles

Live-Cell Imaging of RNA G-Quadruplex with a Dual-Color Fluorescence Switch-on Probe.

In-cell imaging of the G-quadruplex (G4) formation of nucleic acids remains challenging for revealing G4's functions in cells. This study describes the cell imaging method of G4 nucleic acids using our unique tripodal quinone-cyanine fluorescent dye, QCy(MeBT)3, whose 600nm and 700nm fluorescence is enhanced independently upon the binding with double-stranded DNA and G4-DNA/RNA, respectively. The use of QCy(MeBT)3 enables us to visualize cytosolic G4 RNAs in fixed and living HeLa cells with near-infrared 700nm fluorescence.

Numerical and analytical inspection of magnetic field effects in the radical pair mechanism by a simplified rate equation model.

The radical pair mechanism is by now the most prominent candidate for a biologically relevant quantum effect of magnetic fields. Recently, N. Ikeya and J. R. Woodward demonstrated a magnetic field effect for sub-extremely low frequency (ELF) fields in the mT range by investigating the autofluorescence spectrum of flavin adenine dinucleotide in living HeLa cells. We apply a simple rate equation model to show numerically and analytically that magnetic field effects can be expected to exist in the whole ELF range.

Rationally Designed G-Quadruplex Selective "Turn-On" NIR Fluorescent Probe with Large Stokes Shift for Nucleic Acid Research-Based Applications.

Guanine-rich DNA/RNA sequences can form Hoogsteen bonds to adopt noncanonical secondary structures called G-quadruplexes, and these have been associated with diverse cellular processes. There has been considerable research interest in the design of G4-interacting ligands for cellular probing of the G4 structure and understanding its associated biological function. Most of the fluorescent G4 ligands either do not have significant selectivity over other nucleic acid structures, have high Stokes shift, or are not in the near-infrared (NIR) region, which limits its cellular visualization. The current work involves the rational design and synthesis of NIR fluorescent probes comprising a (Z)-1-methyl-2-((3-methylbenzo[d]thiazol-2(3H)-ylidene)methyl)quinolin-1-ium scaffold. Among the designed molecules, 4a exhibited far-red fluorescence (λmax = 680 nm) with large Stokes shift (∼182 nm) upon selective binding to human telomeric G-quadruplexes. The dye 4a does not disturb the conformation and stability of G-quadruplexes, thereby making it suitable for nucleic acid research based applications. Interestingly, 4a showed remarkable selectivity over single- and double-stranded structures in contrast to a commercially available quadruplex binding probe, Thiazole orange (TO). The molecular docking studies indicate that 4a binds at the groove region of the telomeric DNA G-quadruplex through π-π stacking interactions with the quinoline and amine-substituted phenyl ring and with the phosphate backbone through anion-π interactions with the benzothiazole ring. The designed molecule 4a has interesting photophysical properties, cell permeability, and biocompatibility with minimal cytotoxicity. Fluorescence imaging studies in live HeLa cells showed that probe 4a binds to the transient population of the DNA G-quadruplex in the nucleus and RNA quadruplexes in the cytoplasm. In brief, G-quadruplex NIR fluorescent probe 4a with a higher signal/noise ratio has significant potential for cellular imaging studies and thus opens avenues to decipher the biological pathways for better understanding of G-quadruplex biology.

Multifunctional Au–Ag NCs for luminescence and colorimetric double signal sensing of H2S and catalytic reduction of nitrophenol

In this study, the N-Acetyl-l-Cysteine (NAC)-capped gold and silver bimetallic nanoclusters (NAC@Au–Ag NCs) was synthesized by reflux method. Due to the silver effect, the NAC@Au–Ag NCs exhibited strong photoluminescence at far-red/near-infrared regions and better catalytic performance than Au or Ag NCs. Upon addition of H2S, the Au–Ag NCs exhibited obvious fluorescence quench and color changes through the generation of metal sulfides and a static quenching process. The Au–Ag NCs displayed a wide linear luminescence response for H2S (18.5–217 μmol/L) with a detection limit of 0.269 μmol/L. Moreover, the visible color of Au–Ag NCs changed from white to brown-yellow along with increased H2S, the corresponding RGB values also displayed good linearity with the concentration of H2S (90.1–678 μmol/L). Notably, the fabrication of test strips provided a convenient and intuitive tool to screen the freshness of eggs by the color change of test strips. Au–Ag NCs could be used for living HeLa cells bioimaging and recognition of H2S abnormalities. Furthermore, it can be used as a catalyst to reduction of nitrophenols (NPhs), specific included 2NP, 3NP and 4NP. The reaction was regarded as a pseudo-first-order kinetic reaction due to the presence of excess NaBH4. At 298K, the catalytic rate constants(k) of 2NP, 3NP and 4NP were 0.1754 min−1, 0.1734 min−1 and 0.2782 min−1, respectively. The NAC@Au–Ag NCs catalyst still showed good catalytic activity and reusability after five cycles. Therefore, this study developed a H2S sensor for food samples and biological systems. And this nanocatalyst had great application potential for removed the nitrophenol pollutants in water.

A new coumarin-based “OFF–ON” fluorescent sensor for H2S detection in HeLa cells

A new “OFF–ON” coumarin-based fluorescent probe for H2S detection was designed and successfully developed through O-sulfonylation between a dabsyl quencher and 7-hydroxy-4-methylcoumarin as a fluorescent reporter, based on a FRET approach. This H2S responsive probe, utilizing H2S assisted thiolysis of a sulfonate ester as the sensing strategy, demonstrated excellent performance towards H2S with a limit of detection (LoD) of 1.64 µM, along with superb selectivity, good stability and high specificity towards H2S without interference from other biomarkers and analytes. Moreover, dabsyl-7-hydroxy-4-methylcoumarin (dabsylcoumarin) is capable of permeating the cell membrane and effectively visualizing the level of H2S in the living HeLa cells without cytotoxicity.

Targeting Mitochondrial Guanine Quadruplexes for Photoactivatable Chemotherapy in Hypoxic Environments.

A first example of a mitochondrial G-quadruplex (mitoG4s) targeted Ru(II) photooxidant complex is reported. The complex, Ru-TAP-PDC3 induces photodamage toward guanine quadruplexes (G4s) located in the mitochondrial genome under hypoxic and normoxic conditions. Ru-TAP-PDC3 shows high affinity for mitoG4s and localises within mitochondria of live HeLa cells. Immunolabelling with anti-G4 antibody, BG4, confirms Ru-TAP-PDC3 associates with G4s within the mitochondria of fixed cells. The complex induces depletion of mtDNA in live cells under irradiation at 405 nm, confirmed by loss of PicoGreen signal from mitochondria. Biochemical studies confirm this process induces apoptosis. The complex shows low dark toxicity and an impressive phototoxicity index (PI) of >89 was determined in Hela under very low intensity irradiation, 5 J/cm2. The phototoxicity is thought to operate through both Type II singlet oxygen and Type III pathways depending on normoxic or hypoxic conditions, from live cell assays and plasmid DNA cleavage. Overall, we demonstrate targeting mitoG4s and mtDNA with a photooxidant is a potent route to achieving apoptosis under hypoxic conditions that can be extended to phototherapy.

Targeting Mitochondrial Guanine Quadruplexes for Photoactivatable Chemotherapy in Hypoxic Environments

AbstractA first example of a mitochondrial G‐quadruplex (mitoG4s) targeted Ru(II) photooxidant complex is reported. The complex, Ru‐TAP‐PDC3 induces photodamage toward guanine quadruplexes (G4s) located in the mitochondrial genome under hypoxic and normoxic conditions. Ru‐TAP‐PDC3 shows high affinity for mitoG4s and localises within mitochondria of live HeLa cells. Immunolabelling with anti‐G4 antibody, BG4, confirms Ru‐TAP‐PDC3 associates with G4s within the mitochondria of fixed cells. The complex induces depletion of mtDNA in live cells under irradiation at 405 nm, confirmed by loss of PicoGreen signal from mitochondria. Biochemical studies confirm this process induces apoptosis. The complex shows low dark toxicity and an impressive phototoxicity index (PI) of >89 was determined in Hela under very low intensity irradiation, 5 J/cm2. The phototoxicity is thought to operate through both Type II singlet oxygen and Type III pathways depending on normoxic or hypoxic conditions, from live cell assays and plasmid DNA cleavage. Overall, we demonstrate targeting mitoG4s and mtDNA with a photooxidant is a potent route to achieving apoptosis under hypoxic conditions that can be extended to phototherapy.

Copper-bridged fluorescent turn-on molecular system for high-performance identification and bioimaging of carbon monoxide

Novel high-performance fluorescent strategies have always been in great demand for carbon monoxide (CO) detection and cell-imaging at room temperature, which have two sides of a coin: its high toxicity even at low concentrations and crucial role in physiological and pathophysiological processes. Herein, we constructed a copper-bridged fluorescent turn-on molecular system, Nap-BC-Cu2+, in monitoring and cell-imaging of vaporous CO at room temperature. The absence of expensive palladium salts and the capability to detect both CORM-3 and CO are two major advantages of this strategy compared to previous reports. The sensing performance of the system is exceptional, demonstrated by less than 1 s response time and high sensitivity with remarkable fluorescence enhancement. The detection limit is calculated to be as low as approximately 82 nM and ∼ 14 ppb for CORM-3 and CO vapor, respectively. Besides, the presence of potential interferences had minimal impact on the detection. Moreover, the quantitative detection of CO vapor was also realized by naked-eye observation via paper tests, which provide a low-cost portable strategy for the real-time CO detection. Furthermore, the fluorescent turn-on molecular system exhibited low cell toxicity and high cellular uptake, and was effectively utilized for imaging CO in living Hela cells.

A lysosome-targeted fluorescent probe for thiol detection in drug analysis and multiple biological systems.

Biothiols, characterized by their unique sulfhydryl (-SH) groups, possess excellent antioxidant properties, effectively neutralizing the damage to cellular structures caused by reactive oxygen species (ROS) in living organisms. Additionally, lysosomes play a crucial role in decomposing damaged biomolecules through the action of their internal enzymes, regulating the cellular redox state, and mitigating oxidative stress. To facilitate rapid monitoring of intracellular biothiols, particularly within lysosomes, we constructed a lysosome-targeted biothiol fluorescent probe, PHL-DNP, in this study. PHL-DNP exhibited excellent photophysical properties in an aqueous test system, including strong fluorescence enhancement response, excellent selectivity, and low detection limits (Cys 16.5nM, Hcy 16.8nM, GSH 21.3nM, Cap 26.6nM). These attributes enabled easy and efficient qualification of Cys on test strips and accurate determination of the effective content of captopril tablets. Notably, PHL-DNP demonstrated low cytotoxicity and precise lysosomal targeting. Through bioimaging, PHL-DNP not only monitored changes in biothiol levels under oxidative stress but also assessed biothiols in complex biological systems such as live HeLa cells, zebrafish, tumor tissue sections, and radish roots. This provides a promising tool for quantitative analysis of biothiols, disease marker detection, and drug testing.

Ultrafast and Chemoselective Biotinylation of Living Cell Surfaces for Time-Resolved Surfaceome Analysis.

Cell surface proteins participate in many important biological processes, such as cell-to-cell interaction, signal transduction, cell adhesion, and protein transportation. In-depth study of the cell surface protein group is of great significance. Nevertheless, detection and analysis of the surfaceome remain a significant challenge due to their low abundance and hydrophobicity. Herein, we reported an ultrafast and chemoselective labeling method using our newly developed trifunctional probe, the OPA-S-S-alkyne, which labeled cell surface lysine residues, and then established a novel cell surfaceome profiling approach. According to our experimental results, the OPA-S-S-alkyne probe can react extremely fast with living cells, labeling cells in only 1 min, while traditional NHS (labeling cell surface lysine with N-hydroxysuccinimide ester probe) and CSC (labeling cell surface glycan with hydrazide biotin probe) methods normally take longer time of more than 30 min and 1 h, respectively. Taking advantage of this ultrafast property of the method, we highlight the utility of this method by exploring the temporal dynamic changes of surfaceome upon EGF stimulation in living Hela cells and reported "early" and "late" EGF-regulated cell surface proteins, which are difficult to be distinguished by the current cell surface profiling approaches.

Organelle-Targeting Polymer Dots Exhibiting Thermally Activated Delayed Fluorescence for Subcellular Imaging.

Selective imaging of specific subcellular structures provides valuable information about the cellular microenvironment. Materials exhibiting thermally activated delayed fluorescence (TADF) are rapidly emerging as metal-free probes with long-lived emission for intracellular time-gated imaging applications. Polymers incorporating TADF emitters can self-assemble into luminescent nanoparticles, termed polymer dots (Pdots), and this strategy enables them to circumvent the limitations of commercial organelle trackers and small molecule TADF emitters. In this study, diblock copolymers comprised of a hydrophilic block containing organelle-targeting monomers and a hydrophobic TADF-active block were synthesized by ring-opening metathesis polymerization (ROMP). Oxanorbornene-based monomers incorporating morpholine and triphenylphosphonium groups for lysosome and mitochondria targeting, respectively, were also synthesized. ROMP by sequential addition yielded well-defined diblock copolymers with dispersities <1.28. To analyze the effect of tuning the hydrophilic corona on cellular viability and uptake, we prepared Pdots with poly(ethylene glycol) (PEG) and bis-guanidinium (BGN) coronas, resulting in limited and efficient cellular uptake, respectively. Red-emissive Pdots with BGN-based coronas and organelle-targeting functionality were obtained with quantum yields up to 12% in water under air. Colocalization analysis confirmed that lysosome and mitochondria labeling in live HeLa cells was accomplished within 2 h of incubation, affording Pearson's correlation coefficients of 0.37 and 0.70, respectively. The potential application of these Pdots for time-resolved imaging is highlighted by a proof of concept using time-gated spectroscopy, which effectively separates the delayed emission of the TADF Pdots from the background autofluorescence of biological serum.

Development of a fluorescent sensor for dual visual detection of Cr3+ and H2S using a fluorescein framework and bio-imaging applications

A novel responsive fluorescent sensor derived from a fluorescein framework has been successfully fabricated for highly selective dual recognition of Cr3+ and H2S in aqueous solution. Bisanthraflu demonstrated excellent selectivity and great sensitivity towards Cr3+ with a non-fluorescent complex of bisanthraflu-Cr3+ formation via a 2:1 binding mode with minimum detectable amount of 1.7 μM. The fluorescent intensity of the bisanthraflu-Cr3+ complex was markedly restored in the presence of H2S via the displacement approach, potentially applicable as a highly responsive probe for H2S detection with a low detectable concentration of 1.95 μM. This dual functional fluorescent sensor with an On-Off-On approach demonstrated potential for detecting H2S in live HeLa cells with not significant cytotoxicity.

Dicyanisophorone-based viscosity fluorescent probe and its fluorescence imaging in cells and zebrafish

Cellular viscosity is a critical parameter for maintaining normal cellular functions, and its abnormalities are associated with the occurrence and development of various diseases. Therefore, real-time monitoring of cellular viscosity is of significant importance for understanding viscosity-related biological functions and revealing the pathological mechanisms of diseases. In this study, we developed a novel fluorescence probe (IP-HBO) by hybridizing dicyanisophorone with 2-(2-hydroxyphenyl)benzoxazole for monitoring of cellular microenvironment and viscosity changes in zebrafish. The probe IP-HBO combines the principles of molecular rotors and enhanced solid-state fluorescence properties, demonstrating highly specific responsiveness to viscosity, long-wavelength emission, large Stokes shift, and excellent photostability. It has been successfully employed for fluorescence imaging of viscosity in live HeLa cells and zebrafish. The construction of this probe provides a new tool for gaining insights into viscosity-related biological processes and disease mechanisms within cells.

Single-point substitution F97M leads to in cellulo crystallization of the biphotochromic protein moxSAASoti

To enhance the photoconversion performance of biphotochromic moxSAASoti protein, a substitution F97 M was introduced. In addition to enhancing the target properties, this substitution also resulted in the crystallization of the recombinant protein within living HeLa cells (also referred to as in cellulo crystallization). The phenomenon of protein crystallization in living cells is not unique, yet the mechanisms and application of in cellulo crystallization remain significant for further research. However, in cellulo crystallization is atypical for fluorescent proteins and detrimental for their biotechnological application. The objective of this study was to elucidate the underlying mechanisms responsible for the crystallization of moxSAASotiF97Min cellulo. For this purpose, the crystal structure of the green form of biphotochromic protein moxSAASotiF97M was determined at high resolution, which surprisingly has a space group, different from those of parent mSAASotiC21N. The analysis provided allowed to propose a mechanism of new crystal contacts formation, which might be a cause of in cellulo protein crystallization.

Side-chain-engineered fluorescent dyes for 3D and long-term dynamic tracking of the plasma membrane in living cells

The plasma membrane involves in many important biological events such as cell fusion and programmed cell death, but most of current plasma membrane probes cannot meet the requirement of long-term specific anchoring to the plasma membrane. Herein, we propose a molecular side-chain engineering strategy to modulate the long-term imaging performance of fluorescent dyes to the plasma membrane by regulating the cell permeability and anchoring ability. A series of FMR dyes with different lengths of side chains were designed and synthesized, and their transmembrane behaviours and staining performance were evaluated in living HeLa cells. We found that short-chain and medium-chain FMR dyes have excellent cell permeability without the labeling ability to the plasma membrane while the long-chain FMR dyes specifically stain the plasma membrane and can be firmly anchored to the plasma membrane for a long period of time. These long-chain FMR dyes have high stain specificality to the plasma membrane, and C10-FMR can be anchored to the plasma membrane of living cells for 2 h, which enables it to continuously monitor dynamic changes of the plasma membrane. The three-dimensional precision imaging of various cells was achieved using C10-FMR, which provides an opportunity to obtain complete information on the three-dimensional spatial morphology of the plasma membrane. The PEG-induced cell fusion of chicken red blood cells and H2O2-induced apoptosis of HeLa cells were monitored by real-time tracking of dynamic changes of the plasma membrane during these processes, which provide solid examples to prove the usefulness of these fluorescent dyes as long-term imaging tools. This work validates the hypothesis that cell permeability of membrane dyes can be readily regulated by tuning the side chains, and provides the effective design strategy of fluorescent dyes for 3D and long-term dynamic tracking of the plasma membrane of diverse animal cells.

Novel phenothiazine appended thiophene and pyridine for highly selective and sensitive detection of hypochlorite in aqueous and living cells

A novel colorimetric chemosensor, (Z)-3-(5-(10H-phenothiazin-10-yl)thiophen-2-yl)-2-(pyridin-2-yl)acrylonitrile (PTH-TH-PY), was synthesized through Buchwald coupling followed by Knoevenagel condensation for the detection of hypochlorite ions (OCl-). This compound demonstrates exceptional selectivity and sensitivity towards OCl- ions with a low detection limit of 0.2 nM. PTH-TH-PY's remarkable sensitivity is attributed to its effective inhibition of the internal charge transfer (ICT) mechanism. High-resolution mass spectrometry (HR-MS) and Job's plot analysis confirmed a 1:1 binding ratio between the compound and OCl-. The probe also changes from yellow to colorless upon interaction with OCl- in naked-eye. The practical utility of PTH-TH-PY was validated through TLC plate analysis and its ability to detect OCl- in various water samples. Furthermore, the compound's capacity to detect hypochlorite in live HeLa cells indicates its potential application in biological systems. These findings highlight PTH-TH-PY as a promising sensor for real-time OCl- monitoring in diverse settings, including environmental and medical contexts.

Behaviors of nucleosomes with mutant histone H4s in euchromatic domains of living human cells.

Since Robert Feulgen first stained DNA in the cell, visualizing genome chromatin has been a central issue in cell biology to uncover how chromatin is organized and behaves in the cell. To approach this issue, we have developed single-molecule imaging of nucleosomes, a basic unit of chromatin, to unveil local nucleosome behavior in living cells. In this study, we investigated behaviors of nucleosomes with various histone H4 mutants in living HeLa cells to address the role of H4 tail acetylation, including H4K16Ac and others, which are generally associated with more transcriptionally active chromatin regions. We ectopically expressed wild-type (wt) or mutated H4s (H4K16 point; H4K5,8,12,16 quadruple; and H4 tail deletion) fused with HaloTag in HeLa cells. Cells that expressed wtH4-Halo, H4K16-Halo mutants, and multiple H4-Halo mutants had euchromatin-concentrated distribution. Consistently, the genomic regions of the wtH4-Halo nucleosomes corresponded to Hi-C contact domains (or topologically associating domains, TADs)with active chromatin marks (A-compartment). Utilizing single-nucleosome imaging, we found that none of the H4 deacetylation or acetylation mimicked H4 mutants altered the overall local nucleosome motion. This finding suggests that H4 mutant nucleosomes embedded in the condensed euchromatic domains with excess endogenous H4 nucleosomes cannot cause an observable change in the local motion. Interestingly, H4 with four lysine-to-arginine mutations displayed a substantial freely diffusing fraction in the nucleoplasm, whereas H4 with a truncated N-terminal tail was incorporated in heterochromatic regions as well as euchromatin. Our study indicates the power of single-nucleosome imaging to understand individual histone/nucleosome behavior reflecting chromatin environments in living cells.

Pixel-super-resolved lens-free quantitative phase microscopy with partially coherent illumination

Lens-free on-chip microscopy (LFOCM) has been widely utilized in digital pathology, drug screening, point-of-care testing (POCT), and quantitative phase imaging (QPI) due to its high throughput imaging capability and compactness. Initially, coherent laser sources were used in LFOCM to generate interference fringes to reconstruct the intensity and phase information of an object. The use of partially coherent light-emitting diodes (LEDs) in LFOCM offers a more portable and cost-effective alternative to conventional coherent illumination sources. However, the coherence-gating effect from a relatively low degree of coherence may cause a blur of high-frequency information in holograms, leading to an inaccurate object recovery. Thus, we present a pixel-super-resolved lens-free quantitative phase microscopy (PSR-LFQPM) with partially coherent illumination, which not only compensates for the impact of low coherence without increasing the volume of the system but also suppresses the theoretical Nyquist-Shannon sampling resolution limit imposed by the sensor pixel size (0.9 μm). Based on the partially coherent imaging model, we integrate the spatial coherence transfer function (SCTF) obtained from the pre-calibrated LED source distribution during the iteration process to obtain an accurate high-resolution recovery. Applying PSR-LFQPM to image living HeLa cells in vitro, we achieve real-time dynamic high-throughput QPI performance (half-pitch resolution of 780 nm with a 1.41-fold improvement compared to results without considering the effect of coherence) across a wide FOV (19.53 mm2). The proposed method provides a compact, low-cost, and high-throughput lens-free on-chip microscopy system for biomedical and POCT applications.

Bright ESIPT emission from 2,6-di(thiazol/oxazol/imidazol-2-yl)phenol derivatives in solution, aggregation and solid states

Most luminophores often suffer from the problem of aggregation-caused quenching (ACQ) or fluorescence disappearance in dilute solution. It is significant to bridge the gap between ACQ and AIE. In this work, a facile but effective strategy was proposed for the fabrication of always-on luminophores based on the excited state intramolecular proton transfer (ESIPT) mechanism, and six luminophores emitting bright fluorescence in solution, aggregation and solid states were synthesized from 5-tert-butyl-2-hydroxyisophthalaldehyde. All these ESIPT systems show only keto emission owing to their congested structures which block the breakage of intramolecular hydrogen bond (O–H⋯N) by solvation, and subsequently make enol emission impossible. Three of these luminophores are prone to convert into the corresponding phenolate anions emitting blue-shifted emission, which enable them to sense pH variation in the weakly basic range. Furthermore, white-light emission was achieved by combining two of them which show complementary-color fluorescence, and one of them was utilized for bioimaging of living Hela cells and the high-resolution image was obtained.

Bioinspired Hg2+-sensing fluorogenic probe based on amino acid–functionalized rhodamine

Inspired by the suitability of rhodamine spirolactam switching for superresolution imaging and fluorescent probe development, we herein synthesized rhodamine-based probes with sulfur-bearing amine and amino acid moieties and screened their abilities to respond to Hg2+ in the presence of other metal ions. The probe based on S-methylcysteine methyl ester–substituted rhodamine 6G (Rho6G-6) was identified as optimal because of its straightforward synthesis and rapid, selective, and sensitive (detection limit = 8 × 10–9 M) response to Hg2+ manifested by absorption spectrum and emission color changes. The complexation of Hg2+ by Rho6G-6 was examined by absorbance and fluorescence analyses, electrospray ionization–mass spectrometry, and 1H nuclear magnetic resonance (NMR) titration experiments. Furthermore, the practical applicability of the above probe was demonstrated by naked-eye colorimetric Hg2+ detection using paper test strips and the fluorescence-based qualitative mapping of the Hg2+ distribution in living HeLa cells and the organs of live mice. Thus, unlike other probe design strategies, the use of flexible S-methyl amino acids for Hg2+ binding provides new opportunities and strategies for photopromoting the ring switching of the rhodamine spirothiolactam.