De novo labeling and trafficking of individual lipid species in live cells
Objective: Lipids exert dynamic biological functions which are determined both by their fatty acyl compositions and spatiotemporal distributions inside the cell. However, it remains a daunting task to investigate any of these features for each of the more than 1000 lipid species due to a lack of a u...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2022-07-01
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| Series: | Molecular Metabolism |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2212877822000801 |
| _version_ | 1818254048723206144 |
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| author | Jun Zhang Jia Nie Haoran Sun Jie Li John-Paul Andersen Yuguang Shi |
| author_facet | Jun Zhang Jia Nie Haoran Sun Jie Li John-Paul Andersen Yuguang Shi |
| author_sort | Jun Zhang |
| collection | DOAJ |
| description | Objective: Lipids exert dynamic biological functions which are determined both by their fatty acyl compositions and spatiotemporal distributions inside the cell. However, it remains a daunting task to investigate any of these features for each of the more than 1000 lipid species due to a lack of a universal labeling method for individual lipid moieties in live cells. Here we report a de novo lipid labeling method for individual lipid species with precise acyl compositions in live cells. The method is based on the principle of de novo lipid remodeling of exogenously added lysolipids with fluorescent acyl-CoA, leading to the re-synthesis of fluorescence-labeled lipids which can be imaged by confocal microscopy. Methods: The cells were incubated with lysolipids and a nitro-benzoxadiazolyl (NBD) labeled acyl-CoA. The newly remodeled NBD-labeled lipids and their subcellular localization were analyzed by confocal imaging in live cells. Thin layer chromatography was carried out to verify the synthesis of NBD-labeled lipids. The mitochondrial trafficking of NBD-labeled lipids was validated in live cells with targeted deletion of phospholipids transporters, including TRIAP1/PRELI protein complex and StarD7. Results: Incubation cells with lysolipids and NBD-acyl-CoA successfully labeled major lipid species with precise acyl compositions, including phospholipids, cholesterol esters, and neutral lipids, which can be analyzed by confocal imaging in live cells. In contrast to exogenously labeled lipids, the de novo labeled lipids retained full biological properties of their endogenous counterparts, including subcellular localization, trafficking, and recognition by lipid transporters. This method also uncovered some unexpected features of newly remodeled lipids and their transporters. Conclusions: The de novo lipid labeling method not only provides a powerful tool for functional analysis of individual lipid species and lipid transporters, but also calls for re-evaluation of previously published results using exogenously labeled lipids. |
| first_indexed | 2024-12-12T16:49:46Z |
| format | Article |
| id | doaj.art-2957dccba462469faafb2498d102e814 |
| institution | Directory Open Access Journal |
| issn | 2212-8778 |
| language | English |
| last_indexed | 2024-12-12T16:49:46Z |
| publishDate | 2022-07-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Molecular Metabolism |
| spelling | doaj.art-2957dccba462469faafb2498d102e8142022-12-22T00:18:23ZengElsevierMolecular Metabolism2212-87782022-07-0161101511De novo labeling and trafficking of individual lipid species in live cellsJun Zhang0Jia Nie1Haoran Sun2Jie Li3John-Paul Andersen4Yuguang Shi5Sam and Ann Barshop Institute for Longevity and Aging Studies, Department of Pharmacology, University of Texas Health Science Center at San Antonio, 4939 Charles Katz Drive, San Antonio, TX, 78229, USA; Perenna Pharmceuticals Inc., 14785 Omicron Drive, Ste 100, San Antonio, TX, 78245, USASam and Ann Barshop Institute for Longevity and Aging Studies, Department of Pharmacology, University of Texas Health Science Center at San Antonio, 4939 Charles Katz Drive, San Antonio, TX, 78229, USADepartment of Biochemistry and Molecular Biology, Nanjing Medical University, Nanjing, People's Republic of ChinaDepartment of Biochemistry and Molecular Biology, Nanjing Medical University, Nanjing, People's Republic of ChinaSam and Ann Barshop Institute for Longevity and Aging Studies, Department of Pharmacology, University of Texas Health Science Center at San Antonio, 4939 Charles Katz Drive, San Antonio, TX, 78229, USASam and Ann Barshop Institute for Longevity and Aging Studies, Department of Pharmacology, University of Texas Health Science Center at San Antonio, 4939 Charles Katz Drive, San Antonio, TX, 78229, USA; Department of Biochemistry and Molecular Biology, Nanjing Medical University, Nanjing, People's Republic of China; Corresponding author. Joe R. & Teresa Lozano Long Distinguished Chair in Metabolic Biology, Professor of Pharmacology, Barshop Institute for Longevity and Aging Studies, University of Texas Health Science Center at San Antonio, 4939 Charles Katz Drive, San Antonio, TX 78229. fax: +210 562 6150.Objective: Lipids exert dynamic biological functions which are determined both by their fatty acyl compositions and spatiotemporal distributions inside the cell. However, it remains a daunting task to investigate any of these features for each of the more than 1000 lipid species due to a lack of a universal labeling method for individual lipid moieties in live cells. Here we report a de novo lipid labeling method for individual lipid species with precise acyl compositions in live cells. The method is based on the principle of de novo lipid remodeling of exogenously added lysolipids with fluorescent acyl-CoA, leading to the re-synthesis of fluorescence-labeled lipids which can be imaged by confocal microscopy. Methods: The cells were incubated with lysolipids and a nitro-benzoxadiazolyl (NBD) labeled acyl-CoA. The newly remodeled NBD-labeled lipids and their subcellular localization were analyzed by confocal imaging in live cells. Thin layer chromatography was carried out to verify the synthesis of NBD-labeled lipids. The mitochondrial trafficking of NBD-labeled lipids was validated in live cells with targeted deletion of phospholipids transporters, including TRIAP1/PRELI protein complex and StarD7. Results: Incubation cells with lysolipids and NBD-acyl-CoA successfully labeled major lipid species with precise acyl compositions, including phospholipids, cholesterol esters, and neutral lipids, which can be analyzed by confocal imaging in live cells. In contrast to exogenously labeled lipids, the de novo labeled lipids retained full biological properties of their endogenous counterparts, including subcellular localization, trafficking, and recognition by lipid transporters. This method also uncovered some unexpected features of newly remodeled lipids and their transporters. Conclusions: The de novo lipid labeling method not only provides a powerful tool for functional analysis of individual lipid species and lipid transporters, but also calls for re-evaluation of previously published results using exogenously labeled lipids.http://www.sciencedirect.com/science/article/pii/S2212877822000801Lipid remodelingPhospholipid transportersLipid traffickingNBD |
| spellingShingle | Jun Zhang Jia Nie Haoran Sun Jie Li John-Paul Andersen Yuguang Shi De novo labeling and trafficking of individual lipid species in live cells Molecular Metabolism Lipid remodeling Phospholipid transporters Lipid trafficking NBD |
| title | De novo labeling and trafficking of individual lipid species in live cells |
| title_full | De novo labeling and trafficking of individual lipid species in live cells |
| title_fullStr | De novo labeling and trafficking of individual lipid species in live cells |
| title_full_unstemmed | De novo labeling and trafficking of individual lipid species in live cells |
| title_short | De novo labeling and trafficking of individual lipid species in live cells |
| title_sort | de novo labeling and trafficking of individual lipid species in live cells |
| topic | Lipid remodeling Phospholipid transporters Lipid trafficking NBD |
| url | http://www.sciencedirect.com/science/article/pii/S2212877822000801 |
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