Functional characterization of the transient receptor potential melastatin 2 (TRPM2) cation channel from Nematostella vectensis reconstituted into lipid bilayer
Abstract Transient receptor potential melastatin 2 (TRPM2) cation channel activity is required for insulin secretion, immune cell activation and body heat control. Channel activation upon oxidative stress is involved in the pathology of stroke and neurodegenerative disorders. Cytosolic Ca2+, ADP-rib...
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| Format: | Article |
| Language: | English |
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Nature Portfolio
2023-07-01
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| Series: | Scientific Reports |
| Online Access: | https://doi.org/10.1038/s41598-023-38640-6 |
| _version_ | 1797778997995634688 |
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| author | Andras Szollosi János Almássy |
| author_facet | Andras Szollosi János Almássy |
| author_sort | Andras Szollosi |
| collection | DOAJ |
| description | Abstract Transient receptor potential melastatin 2 (TRPM2) cation channel activity is required for insulin secretion, immune cell activation and body heat control. Channel activation upon oxidative stress is involved in the pathology of stroke and neurodegenerative disorders. Cytosolic Ca2+, ADP-ribose (ADPR) and phosphatidylinositol-4,5-bisphosphate (PIP2) are the obligate activators of the channel. Several TRPM2 cryo-EM structures have been resolved to date, yet functionality of the purified protein has not been tested. Here we reconstituted overexpressed and purified TRPM2 from Nematostella vectensis (nvTRPM2) into lipid bilayers and found that the protein is fully functional. Consistent with the observations in native membranes, nvTRPM2 in lipid bilayers is co-activated by cytosolic Ca2+ and either ADPR or ADPR-2′-phosphate (ADPRP). The physiological metabolite ADPRP has a higher apparent affinity than ADPR. In lipid bilayers nvTRPM2 displays a large linear unitary conductance, its open probability (Po) shows little voltage dependence and is stable over several minutes. Po is high without addition of exogenous PIP2, but is largely blunted by treatment with poly-l-Lysine, a polycation that masks PIP2 headgroups. These results indicate that PIP2 or some other activating phosphoinositol lipid co-purifies with nvTRPM2, suggesting a high PIP2 binding affinity of nvTRPM2 under physiological conditions. |
| first_indexed | 2024-03-12T23:24:33Z |
| format | Article |
| id | doaj.art-b8afdd8ba2504534a021e519c02c370e |
| institution | Directory Open Access Journal |
| issn | 2045-2322 |
| language | English |
| last_indexed | 2024-03-12T23:24:33Z |
| publishDate | 2023-07-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj.art-b8afdd8ba2504534a021e519c02c370e2023-07-16T11:14:00ZengNature PortfolioScientific Reports2045-23222023-07-0113111010.1038/s41598-023-38640-6Functional characterization of the transient receptor potential melastatin 2 (TRPM2) cation channel from Nematostella vectensis reconstituted into lipid bilayerAndras Szollosi0János Almássy1Department of Biochemistry, Semmelweis UniversityDepartment of Physiology, Semmelweis UniversityAbstract Transient receptor potential melastatin 2 (TRPM2) cation channel activity is required for insulin secretion, immune cell activation and body heat control. Channel activation upon oxidative stress is involved in the pathology of stroke and neurodegenerative disorders. Cytosolic Ca2+, ADP-ribose (ADPR) and phosphatidylinositol-4,5-bisphosphate (PIP2) are the obligate activators of the channel. Several TRPM2 cryo-EM structures have been resolved to date, yet functionality of the purified protein has not been tested. Here we reconstituted overexpressed and purified TRPM2 from Nematostella vectensis (nvTRPM2) into lipid bilayers and found that the protein is fully functional. Consistent with the observations in native membranes, nvTRPM2 in lipid bilayers is co-activated by cytosolic Ca2+ and either ADPR or ADPR-2′-phosphate (ADPRP). The physiological metabolite ADPRP has a higher apparent affinity than ADPR. In lipid bilayers nvTRPM2 displays a large linear unitary conductance, its open probability (Po) shows little voltage dependence and is stable over several minutes. Po is high without addition of exogenous PIP2, but is largely blunted by treatment with poly-l-Lysine, a polycation that masks PIP2 headgroups. These results indicate that PIP2 or some other activating phosphoinositol lipid co-purifies with nvTRPM2, suggesting a high PIP2 binding affinity of nvTRPM2 under physiological conditions.https://doi.org/10.1038/s41598-023-38640-6 |
| spellingShingle | Andras Szollosi János Almássy Functional characterization of the transient receptor potential melastatin 2 (TRPM2) cation channel from Nematostella vectensis reconstituted into lipid bilayer Scientific Reports |
| title | Functional characterization of the transient receptor potential melastatin 2 (TRPM2) cation channel from Nematostella vectensis reconstituted into lipid bilayer |
| title_full | Functional characterization of the transient receptor potential melastatin 2 (TRPM2) cation channel from Nematostella vectensis reconstituted into lipid bilayer |
| title_fullStr | Functional characterization of the transient receptor potential melastatin 2 (TRPM2) cation channel from Nematostella vectensis reconstituted into lipid bilayer |
| title_full_unstemmed | Functional characterization of the transient receptor potential melastatin 2 (TRPM2) cation channel from Nematostella vectensis reconstituted into lipid bilayer |
| title_short | Functional characterization of the transient receptor potential melastatin 2 (TRPM2) cation channel from Nematostella vectensis reconstituted into lipid bilayer |
| title_sort | functional characterization of the transient receptor potential melastatin 2 trpm2 cation channel from nematostella vectensis reconstituted into lipid bilayer |
| url | https://doi.org/10.1038/s41598-023-38640-6 |
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