Photodynamic Activation of Cholecystokinin 1 Receptor with Different Genetically Encoded Protein Photosensitizers and from Varied Subcellular Sites
Cholecystokinin 1 receptor (CCK1R) is activated by singlet oxygen (<sup>1</sup>O<sub>2</sub>) generated in photodynamic action with sulphonated aluminum phthalocyanine (SALPC) or genetically encoded protein photosensitizer (GEPP) KillerRed or mini singlet oxygen generator (mi...
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MDPI AG
2020-10-01
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| Series: | Biomolecules |
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| Online Access: | https://www.mdpi.com/2218-273X/10/10/1423 |
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| author | Yuan Li Zong Jie Cui |
| author_facet | Yuan Li Zong Jie Cui |
| author_sort | Yuan Li |
| collection | DOAJ |
| description | Cholecystokinin 1 receptor (CCK1R) is activated by singlet oxygen (<sup>1</sup>O<sub>2</sub>) 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 <sup>1</sup>O<sub>2</sub> 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 <i>Methylobacterium radiotolerans</i> with point mutation C71G (Mr4511<sup>C71G</sup>), and flavin-binding fluorescent protein from <i>Dinoroseobacter shibae</i> (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<sup>−2</sup>, 4’ and all others: LED 450 nm, 85 mW·cm<sup>−2</sup>, 1.5′) of GEPP<sub>PM</sub>-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<sub>PM</sub>-AR4-2J cells, light emitting diode (LED) light irradiation-induced calcium oscillations were readily inhibited by CCK1R antagonist devazepide 2 nM; miniSOG<sub>MT</sub>-AR4-2J cells were less susceptible, but miniSOG<sub>LS</sub>-AR4-2J cells were not inhibited. In conclusion, different GEPP<sub>PM</sub> could all photodynamically activate CCK1R. Intracellular GEPP photodynamic action may prove particularly suited to study intracellular GPCR. |
| first_indexed | 2024-03-10T15:47:00Z |
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| institution | Directory Open Access Journal |
| issn | 2218-273X |
| language | English |
| last_indexed | 2024-03-10T15:47:00Z |
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| spelling | doaj.art-ba4ed32bd5544058a45055963e8c72462023-11-20T16:22:45ZengMDPI AGBiomolecules2218-273X2020-10-011010142310.3390/biom10101423Photodynamic Activation of Cholecystokinin 1 Receptor with Different Genetically Encoded Protein Photosensitizers and from Varied Subcellular SitesYuan Li0Zong Jie Cui1Institute of Cell Biology, Beijing Normal University, Beijing 100875, ChinaInstitute of Cell Biology, Beijing Normal University, Beijing 100875, ChinaCholecystokinin 1 receptor (CCK1R) is activated by singlet oxygen (<sup>1</sup>O<sub>2</sub>) 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 <sup>1</sup>O<sub>2</sub> 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 <i>Methylobacterium radiotolerans</i> with point mutation C71G (Mr4511<sup>C71G</sup>), and flavin-binding fluorescent protein from <i>Dinoroseobacter shibae</i> (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<sup>−2</sup>, 4’ and all others: LED 450 nm, 85 mW·cm<sup>−2</sup>, 1.5′) of GEPP<sub>PM</sub>-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<sub>PM</sub>-AR4-2J cells, light emitting diode (LED) light irradiation-induced calcium oscillations were readily inhibited by CCK1R antagonist devazepide 2 nM; miniSOG<sub>MT</sub>-AR4-2J cells were less susceptible, but miniSOG<sub>LS</sub>-AR4-2J cells were not inhibited. In conclusion, different GEPP<sub>PM</sub> could all photodynamically activate CCK1R. Intracellular GEPP photodynamic action may prove particularly suited to study intracellular GPCR.https://www.mdpi.com/2218-273X/10/10/1423genetically encoded protein photosensitizers (GEPP)cholecystokinin 1 receptor (CCK1R)singlet oxygen (<sup>1</sup>O<sub>2</sub>)calcium oscillationsKillerRedminiSOG |
| spellingShingle | Yuan Li Zong Jie Cui Photodynamic Activation of Cholecystokinin 1 Receptor with Different Genetically Encoded Protein Photosensitizers and from Varied Subcellular Sites Biomolecules genetically encoded protein photosensitizers (GEPP) cholecystokinin 1 receptor (CCK1R) singlet oxygen (<sup>1</sup>O<sub>2</sub>) calcium oscillations KillerRed miniSOG |
| title | Photodynamic Activation of Cholecystokinin 1 Receptor with Different Genetically Encoded Protein Photosensitizers and from Varied Subcellular Sites |
| title_full | Photodynamic Activation of Cholecystokinin 1 Receptor with Different Genetically Encoded Protein Photosensitizers and from Varied Subcellular Sites |
| title_fullStr | Photodynamic Activation of Cholecystokinin 1 Receptor with Different Genetically Encoded Protein Photosensitizers and from Varied Subcellular Sites |
| title_full_unstemmed | Photodynamic Activation of Cholecystokinin 1 Receptor with Different Genetically Encoded Protein Photosensitizers and from Varied Subcellular Sites |
| title_short | Photodynamic Activation of Cholecystokinin 1 Receptor with Different Genetically Encoded Protein Photosensitizers and from Varied Subcellular Sites |
| title_sort | photodynamic activation of cholecystokinin 1 receptor with different genetically encoded protein photosensitizers and from varied subcellular sites |
| topic | genetically encoded protein photosensitizers (GEPP) cholecystokinin 1 receptor (CCK1R) singlet oxygen (<sup>1</sup>O<sub>2</sub>) calcium oscillations KillerRed miniSOG |
| url | https://www.mdpi.com/2218-273X/10/10/1423 |
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