Reversible Photoregulation Research Articles

Host-guest assembly triggered lower critical solution temperature behavior with reversible photo-regulation

The less frequent lower critical solution temperature (LCST) phenomenon has attracted extensive attention due to the foresighted application in smart materials. Nevertheless, the existing LCST-type materials are mainly dominated by a few polymers with specific moieties due to the damped hydration at a higher temperature and the lack of reversible regulations of the corresponding temperature sensitivity. Herein, an unanticipated and reversibly photo-responsive LCST behavior was uncovered in the BSC4/Azo-4 assembly solution. Such behavior arose from the host-guest binding between BSC4 and Azo-4 in an aqueous solution since neither the guest nor host alone exhibited an LCST property. Ascribing to the distinct π-π stacking between the trans- and cis-isomers of Azo-4, a reversible regulation of LCST behavior was achieved by irradiating UV and blue light alternately. Our study demonstrated the complexity and possibility of supramolecular self-assembly, opening a new avenue in the fabrication and adjustment of LCST-type materials in the future.

Reversible Photoregulation of Cell-Cell Adhesions With Opto-E-cadherin.

The cell-cell adhesion molecule E-cadherin has been intensively studied due to its prevalence in tissue function and its spatiotemporal regulation during epithelial-to-mesenchymal cell transition. Nonetheless, regulating and studying the dynamics of it has proven challenging. We developed a photoswitchable version of E-cadherin, named opto-E-cadherin, which can be toggled OFF with blue light illumination and back ON in the dark. Herein, we describe easy-to-use methods to test and characterise opto-E- cadherin cell clones for downstream experiments. Key features • This protocol describes how to implement optogenetic cell-cell adhesion molecules effectively (described here on the basis of opto-E-cadherin), while highlighting possible pitfalls. • Utilises equipment commonly found in most laboratories with high ease of use. • Phenotype screening is easy and done within a few hours (comparison of cell clusters in the dark vs. blue light in an aggregation assay). • Three different functionality assay systems are described. • After the cell line is established, all experiments can be performed within three days.

Pd(ii)-catalyzed enantioselective C-H olefination and photoregulation of sterically hindered diarylethenes.

Sterically hindered diarylethenes with intrinsic chirality have shown great potential in chiral signal regulation, light-controlled liquid crystals (LCs), etc. Their unique enantiospecific phototransformation between axial chirality of ring-open isomers and central chirality of ring-closed isomers can break through the bottleneck of interference between multiple chiral centers in traditional chiral diarylethenes. However, these intrinsic chiral diarylethenes require necessary chiral resolution through preparative chiral HPLC, typically resulting in limited separation efficiency and production scale. Here, we present an enantioselective olefination strategy to directly construct intrinsic chiral diarylethenes from a prochiral sterically hindered diarylethene, achieving high yields and enantioselectivity. The resulting isomers can be further decorated by incorporating mesogenic units, and the derivatives enable the successful reversible photoregulation of blue, green, and red reflection colors of LCs with excellent thermal stability, fatigue resistance, and little texture disorderliness, demonstrating the practical application potential of direct enantioselective olefination in photoregulation with intrinsic chiral diarylethenes.

Reversible photoregulation of cell-cell adhesions with opto-E-cadherin

E-cadherin-based cell-cell adhesions are dynamically and locally regulated in many essential processes, including embryogenesis, wound healing and tissue organization, with dysregulation manifesting as tumorigenesis and metastasis. However, the lack of tools that would provide control of the high spatiotemporal precision observed with E-cadherin adhesions hampers investigation of the underlying mechanisms. Here, we present an optogenetic tool, opto-E-cadherin, that allows reversible control of E-cadherin-mediated cell-cell adhesions with blue light. With opto-E-cadherin, functionally essential calcium binding is photoregulated such that cells expressing opto-E-cadherin at their surface adhere to each other in the dark but not upon illumination. Consequently, opto-E-cadherin provides remote control over multicellular aggregation, E-cadherin-associated intracellular signalling and F-actin organization in 2D and 3D cell cultures. Opto-E-cadherin also allows switching of multicellular behaviour between single and collective cell migration, as well as of cell invasiveness in vitro and in vivo. Overall, opto-E-cadherin is a powerful optogenetic tool capable of controlling cell-cell adhesions at the molecular, cellular and behavioural level that opens up perspectives for the study of dynamics and spatiotemporal control of E-cadherin in biological processes.

Reversible photoregulation of morphological structure for porous coumarin-graphene composite and the removal of heavy metal ions

In this study, a photoreversible graphene oxide (GO) - coumarin (GC) composite is fabricated by electrostatic self-assembly of positively charged nitrogenous coumarin surfactant and negatively charged GO. It is found that the introduction of coumarin not only results in a large amount of micropores and mesopores on the surface of GO, but photo-regulates its structure transformation between close-packed structure and 3D pleat-like structure, solves the separation problem, improves and reversibly controls its adsorption capacity, under 365/254 nm light irradiation. The maximum adsorption capacity of the GC composite and the Irradiated GC composite (the GC composite irradiated by 365 nm light for 30 min) for Pb2+ are 403.97 and 492.51 mg/g, respectively, under room temperature at pH 6. The adsorption kinetic processes of these two composites are described by pseudo-second-order kinetic model, and the equilibrium data are well fitted with the Langmuir isotherm model. Furthermore, adsorption-desorption experiments show that these two composites have effective reusability, and display desirable adsorption capacities toward Cd2+, Cu2+ and Zn2+ heavy metal ions. The results indicate that the GC composite providetheoreticaland experimentalbasisfor structurallyphotocontrollable materials, and can be served as photoreversible adsorbents for effective removal of heavy metal ions from wastewater.

Reversible photo-regulation on the folding/unfolding of telomere G-quadruplexes with solid-state nanopores.

The formation of G-quadruplexes (G4) in human telomere and other important biological regions inhibits the replication and transcription of DNA, thereby influencing further cell proliferation. The investigation of G4 formation and unfolding is vital for understanding their modulation in biological processes and life science. Photo regulation is a facile and sensitive approach for monitoring the structures of biomacromolecules and material surface properties. The nanopore-based technique is also prevalent for label-free single-molecule characterization with high accuracy. This study provides a combination of solid-state nanopore technology with light-switch as a platform for the modulation of human telomere G4 formation and splitting under switchable light exposure. The introduction of molecular switch, namely azobenzene moiety at different positions of the DNA sequence influences the formation and stability of G4. Three azobenzenes immobilized on each of the G-quartet plane (hTelo-3azo-p) or four azobenzenes on the same plane (hTelo-4azo-4p) of the human telomere G4 sequence realized the reversible control of G4 folding/unfolding at the temporal scale upon photo regulation, and the formation and splitting of G4 with hTelo-4azo-4p is slower and not thorough compared to that with hTelo-3azo-p due to the coplanar steric hindrance. Moreover, the G4 formation recorded with the combined nanopore and photo-responsive approach was also characterized with fluorescence, and the variation in the fluorescence intensity of the NMM and G4 complex exhibited a different tendency under reverse light irradiation due to the distinct interactions of NMM with the azobenzene-modified G4. Our study demonstrated a controllable and sensitive way for the manipulation of G4 structures, which will be inspiring for the intervention of G4-related cell senescence, cancer diagnosis and drug exploration.

Regulation of mRNA translation by a photoriboswitch.

Optogenetic tools have revolutionized the study of receptor-mediated processes, but such tools are lacking for RNA-controlled systems. In particular, light-activated regulatory RNAs are needed for spatiotemporal control of gene expression. To fill this gap, we used in vitro selection to isolate a novel riboswitch that selectively binds the trans isoform of a stiff-stilbene (amino-tSS)-a rapidly and reversibly photoisomerizing small molecule. Structural probing revealed that the RNA binds amino-tSS about 100-times stronger than the cis photoisoform (amino-cSS). In vitro and in vivo functional analysis showed that the riboswitch, termed Werewolf-1 (Were-1), inhibits translation of a downstream open reading frame when bound to amino-tSS. Photoisomerization of the ligand with a sub-millisecond pulse of light induced the protein expression. In contrast, amino-cSS supported protein expression, which was inhibited upon photoisomerization to amino-tSS. Reversible photoregulation of gene expression using a genetically encoded RNA will likely facilitate high-resolution spatiotemporal analysis of complex RNA processes.

Transcription Driven by Reversible Photocontrol of Hyperstable G-Quadruplexes

G-quadruplexes occur in promoter regions, 5'-untranslated regions of mRNA and telomeric regions, and they function as regulatory elements for various key biological events, such as transcription, translation, and telomere elongation. As the stability of G-quadruplexes dramatically impacts these biological processes, controlling G-quadruplex stability via external stimuli such as light enables regulation of important biological phenomena with high spatial and temporal resolution. Here, we report a method for reversible photoregulation of transcription by controlling the stability of G-quadruplexes via cis- trans photoisomerization of photochromic nucleobase (PCN). Transcription was effectively inhibited when the PCN-modified G-quadruplex was in a hyperstable state, whereas transcription activity recovered markedly when the G-quadruplex changed to an unstable state induced by trans to cis PCN photoisomerization. Moreover, a reversibly photoactivatable plasmid was constructed by introducing PCN-modified G-quadruplexes downstream of the cytomegalovirus promoter of the pCS2 plasmid, which was used to demonstrate photoregulation of gene expression in zebrafish embryos.

Reversible photoregulation in biological systems via introduction of azobenzene into nucleic acid

As photoisomerization mechanism and characteristic of azobenzene was deeper and deeper, reversible photoregulation of related processes via introduction of azobenzene into nucleic acids has received a lot of attention. Azobenzene, as a photoresponsive unit, was not only used to synthesize artificial intelligent materials or molecular machines, but also infiltrating rapidly into analysis and modulation of chemical biological systems. In view of information diversity provided by nucleic acid molecules, the modular introduction of these switchable units into nucleic acids has allowed diverse applications ranging from reversible formation of duplex, photoswitchable RNA silencing, reversible control of gene expression, reversible aptamer recognition, to reversible photocontrol of DNAzymes, as well as obtaining structural information and intermolecular interaction using the introduced azobenzene as an internal probe. Thus, the combination of functional azobenzene group with nucleic acids to apply in biological systems will be a hot topic in the field of nucleic acid chemistry. Here, the design principles and characteristics of four different combining ways between azobenzene and nucleic acids were mainly elaborated, and reversible regulation of structure and function of nucleic acid in biological systems was reviewed due to photo-isomerization properties of azobenzene photosensitizers by screening some representative examples. Finally, the problems that may exist in the biosystems, and looks into the future and trends of the research were also depicted.

Reversible photoregulation of DNA B-Z transition by a photochromic nucleoside

We designed a system using a photochromic nucleoside, which can convert duplex DNA between right-handed and left-handed (B-Z transition) response to light stimuli. Our system DNA can be switched between B-conformation and Z-conformation by irradiating with monochromatic 254nm and 365nm light. The transition of DNA against photo was completely reversible.

Recent developments in reversible photoregulation of oligonucleotide structure and function.

There is a growing interest in the photoregulation of biological functions, due to the high level of spatiotemporal precision achievable with light. Additionally, light is non-invasive and waste-free. In particular, the photoregulation of oligonucleotide structure and function is a rapidly developing study field with relevance to biological, physical and material sciences. Molecular photoswitches have been incorporated in oligonucleotides for 20 years, and the field has currently grown beyond fundamental studies on photochemistry of the switches and DNA duplex stability, and is moving towards applications in chemical biology, nanotechnology and material science. Moreover, the currently emerging field of photopharmacology indicates the relevance of photocontrol in future medicine. In recent years, a large number of publications has appeared on photoregulation of DNA and RNA structure and function. New strategies are evaluated and novel, exciting applications are shown. In this comprehensive review, the key strategies for photoswitch inclusion in oligonucleotides are presented and illustrated with recent examples. Additionally the applications that have emerged in recent years are discussed, including gene regulation, drug delivery and materials design. Finally, we identify the challenges that the field currently faces and look forward to future applications.

Reversible Photoregulation of Gene Expression and Translation.

Several methods for controlling gene expression by light illumination have been reported. Most of these methods control transcription by regulating the interaction between DNA and transcription factors. The use of a photolabile protecting compound (cage compound) is another promising approach for controlling gene expression, although typically in an irreversible manner. We here describe a new approach for reversibly controlling translation using a photoresponsive 8-styryl cap (8ST-cap) that can be reversibly isomerized by illumination with light of a specific wavelength.

Control of cellular function by reversible photoregulation of translation.

The use of light as an external stimulus offers the potential for spatiotemporal control and is thus ideal for controlling gene expression in living cells. In commonly used caging systems, once the caging compound is removed, protein expression cannot be stopped, due to the irreversibility of the uncaging reaction. We have developed a reversible method for regulating protein expression with the aid of a photoresponsive cap that can control the translation of mRNA in a reversible manner through triggering of cis-trans photoisomerization of the cap. In its trans form, the photoresponsive cap completely inhibited translation, whereas the cis form yielded protein (12.7 times more translated protein than in the trans form). Moreover, we succeeded in controlling the levels, timing and duration of protein expression in living mammalian cells. Additionally, neuronal differentiation of PC12 cells was photoinduced by controlling constitutively active H-Ras 61L protein expression.

Photoswitch Nucleic Acid Catalytic Activity by Regulating Topological Structure with a Universal Supraphotoswitch

We demonstrated the generality of a strategy for photoswitching the activity of functional oligonucleotides by modulating their topological structure. Our strategy was proved to be versatile because it can be used to photoregulate functional oligonucleotides, e.g., ribozymes and DNAzymes, which have two binding arms and a catalytic loop. Repeated reversible photoregulation of RNA cleavage by a ribozyme or a DNAzyme was achieved by attaching two photoresponsive strands, artificial oligomers involving azobenzene moieties and nucleobases capable of forming a duplex as the supraphotoswitch. Individual strands were attached to the 3' and 5' ends of a RNA-cleavage oligonucleotide. Thus, the topological structure of the ribozyme or DNAzyme was constrained, and RNA cleavage was greatly suppressed when the supraphotoswitch duplex formed (OFF state). In contrast, RNA cleavage resumed when the supraphotoswitch duplex dissociated (ON state). Light irradiation was used to repeatedly switch the supraphotoswitch between the ON and OFF states so that RNA cleavage activity could be efficiently photoregulated. Analysis of the regulatory mechanism showed that topological constraints suppressed the RNA cleavage by causing both structural changes at the catalytic site and lower binding affinity between the RNA substrates and the functional oligonucleotides.

Photo-Regulation of Cell Division using Photochromic Inhibitors for Kinesin Eg5

Previously it has been demonstrated that some kinesins known as an ATP driven motor protein are directly involved in regulation of the cell division cycle. Eg5 is one of the kinesin and its biochemical properties and structure were well studied. It is suggested that that Eg5 is activated in M phase of cell cycle and performs stabilizing and positioning of spindle. It is also known that Eg5 is overexpressed in tumor cell and induce significant cell division. Monastrol and STLC, which are potent inhibitor specific for kinesin Eg5, shut off mitotic division and result in apotosis. Therefore, these inhibitors are attracting as anti-cancer drug. Azobenzene and spiropyran, a widely studied photochromic compound, can be reversibly isomerized between the cis and trans forms by ultra-violet (UV) and visible (VIS) light irradiation, respectively. We have recently demonstrated the Monastrol and STLC analogues composed of photochromic molecules inhibit ATPase activity of Eg5 reversibly upon UV and VIS light irradiation in vitro. Therefore, it is strongly expected that the photochromic inhibitors are applicable to reversible photo-regulation of cell mitosis. In this study, we have examined the effects of the photochromic inhibitors for mammalian cells. Effects of the photochromic inhibitors were evaluated by survival number of mammalian cells, HeLa cells or A172 cells. Survival numbers of cells were quantitatively analyzed by MTS reagent and cell divisions were observed by phase-contrast microscope. spindle behavior of the cells cultured in the presence of photochromic inhibitors were also examined using immunofluorescent staining. These experimental results suggested that the effects of photochromic inhibitors for the cell division were controlled by UV and VIS light irradiations reversibly.

Photoresponsive 5′-cap for the reversible photoregulation of gene expression

Photoresponsive 5'-caps that can be reversibly cis-trans isomerized by light irradiation were developed for the reversible photoregulation of gene expression. The 8-naphthylvinyl cap (8NV-cap) in the trans form completely inhibited translation of mRNA, whereas the cis form yielded protein with the same efficiency as mRNA capped with the normal-cap, a 26-fold higher efficiency than that of the trans form. The 8NV-capped mRNA could be switched between a translating state (ON state) and a non-translating state (OFF state) in a reversible fashion by alternately irradiating with monochromatic 410 nm or 310 nm light.

Straightforward and Reversible Photoregulation of Hybridization by Using a Photochromic Nucleoside

Straightforward and Reversible Photoregulation of Hybridization by Using a Photochromic Nucleoside

Straightforward and Reversible Photoregulation of Hybridization by Using a Photochromic Nucleoside

Straightforward and Reversible Photoregulation of Hybridization by Using a Photochromic Nucleoside

A thermostable azo-linker for reversible photoregulation of DNA replication

Small molecules such as azobenzenes, one of the best reversible photo-switches, can be covalently incorporated into DNA to regulate its structures and functions with irradiation of the specific wavelengths. Using this strategy, a thermostable azobenzene linker was employed to construct modified oligodeoxynucleotides, and we successfully achieve reversible photoregulation of DNA replication in vitro with short irradiation time. Five minutes UV irradiation for regulating trans→cis transformation can minimize DNA damage, still ensure the polymerase reaction of cis-form. Formation of DNA hairpin structure can also be controlled by photoregulation using this linker.

Reversible Photoregulation of Binding of α-Chymotrypsin to a Gold Surface

An ability to optically modulate the interactions of surfaces with functional biomolecules provides an important basis for generating new technologies including reversible biosensors, advanced medical implants, and biomolecular computers. Here we report the first example of reversible photoregulation of binding of a protease to a functional surface. A modular approach is presented with a surface-bound inhibitor containing a photoisomerizable azobenzene core to which is attached (i) appropriate protease binding functionality and (ii) a tether for surface attachment. The principle is demonstrated for alpha-chymotrypsin using a phenylalanine-based trifluoromethylketone inhibitor containing an azobenzene core and an alkyne-functionalized ethylene glycol tether, which is attached to the surface using click chemistry. UV/vis irradiation of the functional surface leads to a significant, reversible change in the amount of alpha-chymotrypsin that attaches to the surface, as measured by surface plasmon resonance.