The special case of far-red vision of mesopelagic dragonfish Malacosteus niger facilitated by the presence in the rod outer segment of photosensitizer chlorin e6 from diet has drawn considerable attention both in vision research and in photodynamic action. Rhodopsin binding of Ce6 from either the extracellular or intracellular loops may exert different effects. Theoretical works predict that the extracellularly bound Ce6 upon absorption of red light produces a singlet oxygen, which could via an oxygen tunnel reach the Lys-tethered 11-cis-retinal, by way of peroxy-dioxetane intermediates, to enhance 11-cis- to all-trans-retinal isomerization, therefore triggering the ultrafast phototransduction process. Recent works on the permanent photodynamic activation of some A-class G protein-coupled receptors suggest that the singlet oxygen generated by Ce6 photodynamic action might also oxidize the scotopsin moiety of rhodopsin, leading to direct oxidative rhodopsin activation. More attention needs to be paid to the latter respects of the far-red vision process of the deep-sea dragonfish, with potential translational values.

Different from reversible agonist-stimulated receptor activation, singlet oxygen oxidation activates permanently G protein-coupled receptor (GPCR) cholecystokinin 1 (CCK1R) in type II photodynamic action, with soluble photosensitizer dyes (sulphonated aluminum phthalocyanine, λmax 675 nm) or genetically encoded protein photosensitizers (KillerRed λmax 585 nm; mini singlet oxygen generator λmax 450 nm), together with a pulse of light (37 mW/cm2, 1-2 minutes). Three lines of evidence shed light on the mechanism of GPCR activated by singlet oxygen (GPCR-ABSO): (1) CCK1R is quantitatively converted from dimer to monomer; (2) Transmembrane domain 3, a pharmacophore for permanent photodynamic CCK1R activation, can be transplanted to non-susceptible M3 acetylcholine receptor; and (3) Larger size of disordered region in intracellular loop 3 correlates with higher sensitivity to photodynamic CCK1R activation. GPCR-ABSO will add to the arsenal of engineered designer GPCR such as receptors activated solely by synthetic ligands and designer receptors exclusively activated by designer drugs, but show some clear advantages: Enhanced selectivity (double selectivity of localized photosensitizer and light illumination), long-lasting activation with no need for repeated drug administration, antagonist-binding site remains intact when needed, ease to apply to multiple GPCR. This type of permanent photodynamic activation may be applied to functional proteins other than GPCR.

Reduced nicotinamide adenine dinucleotide phosphate [NAD(P)H] oxidases (NOX) are a major cellular source of reactive oxygen species, regulating vital physiological functions, whose dys-regulation leads to a plethora of major diseases. Much effort has been made to develop varied types of NOX inhibitors, but biotechnologies for spatially and temporally controlled NOX activation, however, are not readily available. We previously found that ultraviolet A (UVA) irradiation activates NOX2 in rodent mast cells, to elicit persistent calcium spikes. NOX2 is composed of multiple subunits, making studies of its activation rather complicated. Here we show that the single-subunit nonrodent-expressing NOX5, when expressed ectopically in CHO-K1 cells, is activated by UVA irradiation (380 nm, 0.1-12 mW/cm2, 1.5 min) inducing repetitive calcium spikes, as monitored by Fura-2 fluorescent calcium imaging. UVA-elicited calcium oscillations are inhibited by NOX inhibitor diphenyleneiodonium chloride (DPI) and blocked by singlet oxygen (1O2) quencher Trolox-C (300 μM). A brief pulse of photodynamic action (1.5 min) with photosensitizer sulfonated aluminum phthalocyanine (SALPC 2 μM, 675 nm, 85 mW/cm2) in NOX5-CHO-K1 cells, or with genetically encoded protein photosensitizer miniSOG fused to N-terminus of NOX5 (450 nm, 85 mW/cm2) in miniSOG-NOX5-CHO-K1 cells, induces persistent calcium oscillations, which are blocked by DPI. In the presence of Trolox-C, miniSOG photodynamic action no longer induces any calcium increases in miniSOG-NOX5-CHO-K1 cells. DUOX2 in human thyroid follicular cells SW579 and in DUOX2-CHO-K1 cells is similarly activated by UVA irradiation and SALPC photodynamic action. These data together suggest that NOX   is activated with a brief pulse of photodynamic action.

Light emission by living organisms in the visible spectrum range is called bioluminescence [...].

Acute pancreatitis (AP) is a major, globally increasing gastrointestinal disease and a biliary origin is the most common cause. However, the effects of bile acids (BAs), given systemically, on the pancreas and on disease severity remains elusive. In this study, we have investigated the roles of different circulating BAs in animal models for AP to elucidate their impact on disease severity and the underlying pathomechanisms. BAs were incubated on isolated acini and AP was induced through repetitive injections of caerulein or L-arginine; pancreatic duct ligation (PDL); or combined biliopancreatic duct ligation (BPDL). Disease severity was assessed using biochemical and histological parameters. Serum cholecystokinin (CCK) concentrations were determined via enzyme immunoassay. The binding of the CCK1 receptor was measured using fluorescence-labeled CCK. In isolated acini, hydrophobic BAs mitigated the damaging effects of CCK. The same BAs further enhanced pancreatitis in L-arginine- and PDL-based pancreatitis, whereas they ameliorated pancreatic damage in the caerulein and BPDL models. Mechanistically, the binding affinity of the CCK1 receptor was significantly reduced by hydrophobic BAs. The hydrophobicity of BAs and the involvement of CCK seem to be relevant in the course of AP. Systemic BAs may affect the severity of AP by interfering with the CCK1 receptor.

Cholecystokinin 1 receptor (CCK1R) is activated photodynamically. For this to happen in situ, genetically encoded protein photosensitizers (GEPP) may be tagged to natively expressed CCK1R, but how to best tag GEPP has not been examined. Therefore, GEPP (miniSOG or KillerRed) was tagged to CCK1R and light-driven photodynamic CCK1R activation was monitored by Fura-2 fluorescent calcium imaging, to screen for optimized tagging patterns. Blue light-emitting diode irradiation of CHO-K1 cells expressing miniSOG fused to N- or C-terminus of CCK1R was found to both trigger persistent calcium oscillations-a hallmark of permanent photodynamic CCK1R activation. Photodynamic CCK1R activation was accomplished also with miniSOG fused to N-terminus of CCK1R via linker (GlySerGly)_{4 or 8} , but not linker (GSG)₁₂ or an internal ribosomal entry site insert. KillerRed fused to N- or C-terminus of CCK1R after white light irradiation resulted in similar activation of in-frame CCK1R. Photodynamic CCK1R activation in miniSOG-CCK1R-CHO-K1 cells was blocked by singlet oxygen (¹ O₂ ) quencher uric acid or Trolox C, corroborating the role of ¹ O₂ as the reactive intermediate. It is concluded that photodynamic CCK1R activation can be achieved either with direct GEPP fusion to CCK1R or fusion via a short linker, fusion via long linkers might serve as the internal control.

Cholecystokinin 1 receptor (CCK1R) is activated in photodynamic action by singlet oxygen, but detailed molecular mechanisms are not elucidated. To identify the pharmacophore(s) in photodynamic CCK1R activation, we examined photodynamic activation of point mutants CCK1R^M121/3.32A, CCK1R^M121/3.32Q, and a chimeric receptor with CCK1R transmembrane domain 3 (TM3) transplanted to muscarinic ACh receptor 3 (M3R) which is unaffected by photodynamic action. These engineered receptors were tagged at the N-terminus with genetically encoded protein photosensitizer miniSOG, and their light-driven photodynamic activation was compared to wild type CCK1R and M3R, as monitored by Fura-2 fluorescent calcium imaging. Photodynamic activations of miniSOG-CCK1R^M121/3.32A and miniSOG-CCK1R^M121/3.32Q were found to be 55% and 73%, respectively, when compared to miniSOG-CCK1R (100%), whereas miniSOG-M3R was not affected (0% activation). Notably, the chimeric receptor miniSOG-M3R-TM3_CCK1R was effectively activated photodynamically (65%). These data suggest that TM3 is an important pharmacophore in photodynamic CCK1R activation, readily transplantable to nonsusceptible M3R for photodynamic activation.

Cholecystokinin 1 receptor (CCK1R) is activated by singlet oxygen (¹O₂) generated in photodynamic action with sulphonated aluminum phthalocyanine (SALPC) or genetically encoded protein photosensitizer (GEPP) KillerRed or mini singlet oxygen generator (miniSOG). A large number of GEPP with varied ¹O₂ quantum yields have appeared recently; therefore, in the present work, the efficacy of different GEPP to photodynamically activate CCK1R was examined, as monitored by Fura-2 calcium imaging. KillerRed, miniSOG, miniSOG2, singlet oxygen protein photosensitizer (SOPP), flavin-binding fluorescent protein from Methylobacterium radiotolerans with point mutation C71G (Mr4511^C71G), and flavin-binding fluorescent protein from Dinoroseobacter shibae (DsFbFP) were expressed at the plasma membrane (PM) in AR4-2J cells, which express endogenous CCK1R. Light irradiation (KillerRed: white light 85.3 mW‧cm^-2, 4' and all others: LED 450 nm, 85 mW·cm^-2, 1.5') of GEPP_PM-expressing AR4-2J was found to all trigger persistent calcium oscillations, a hallmark of permanent photodynamic CCK1R activation; DsFbFP was the least effective, due to poor expression. miniSOG was targeted to PM, mitochondria (MT) or lysosomes (LS) in AR4-2J in parallel experiments; LED light irradiation was found to all induce persistent calcium oscillations. In miniSOG_PM-AR4-2J cells, light emitting diode (LED) light irradiation-induced calcium oscillations were readily inhibited by CCK1R antagonist devazepide 2 nM; miniSOG_MT-AR4-2J cells were less susceptible, but miniSOG_LS-AR4-2J cells were not inhibited. In conclusion, different GEPP_PM could all photodynamically activate CCK1R. Intracellular GEPP photodynamic action may prove particularly suited to study intracellular GPCR.