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Near-Infrared Imaging of Steroid Hormone Activities Using Bright BRET Templates

Bioluminescence (BL) is an excellent optical readout for bioassays and molecular imaging. Herein, we accomplished new near infrared bioluminescence resonance energy transfer (NIR-BRET) templates for monitoring molecular events in cells with higher sensitivity. We first identified the best resonance...

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Autores principales: Kim, Sung-Bae, Nishihara, Ryo, Paulmurugan, Ramasamy
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9820568/
https://www.ncbi.nlm.nih.gov/pubmed/36614119
http://dx.doi.org/10.3390/ijms24010677
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author Kim, Sung-Bae
Nishihara, Ryo
Paulmurugan, Ramasamy
author_facet Kim, Sung-Bae
Nishihara, Ryo
Paulmurugan, Ramasamy
author_sort Kim, Sung-Bae
collection PubMed
description Bioluminescence (BL) is an excellent optical readout for bioassays and molecular imaging. Herein, we accomplished new near infrared bioluminescence resonance energy transfer (NIR-BRET) templates for monitoring molecular events in cells with higher sensitivity. We first identified the best resonance energy donor for the NIR-BRET templates through the characterization of many coelenterazine (CTZ)–marine luciferase combinations. As a result, we found that NLuc–DBlueC and ALuc47–nCTZ combinations showed luminescence in the blue emission wavelength with excellent BL intensity and stability, for example, the NLuc–DBlueC and ALuc47–nCTZ combinations were 17-fold and 22-fold brighter than their second highest combinations, respectively, and were stably bright in living mammalian cells for at least 10 min. To harness the excellent BL properties to the NIR-BRET systems, NLuc and ALuc47 were genetically fused to fluorescent proteins (FPs), allowing large “blue-to-red” shifts, such as LSSmChe, LSSmKate2, and LSSmNep (where LSS means Large Stokes Shift). The excellent LSSmNep–NLuc combination showed approximately 170 nm large resonance energy shift from blue to red. The established templates were further utilized in the development of new NIR-BRET systems for imaging steroid hormone activities by sandwiching the ligand-binding domain of a nuclear receptor (NR-LBD) between the luciferase and the FP of the template. The NIR-BRET systems showed a specific luminescence signal upon exposure to steroid hormones, such as androgen, estrogen, and cortisol. The present NIR-BRET templates are important additions for utilizing their advantageous imaging of various molecular events with high efficiency and brightness in physiological samples.
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spelling pubmed-98205682023-01-07 Near-Infrared Imaging of Steroid Hormone Activities Using Bright BRET Templates Kim, Sung-Bae Nishihara, Ryo Paulmurugan, Ramasamy Int J Mol Sci Article Bioluminescence (BL) is an excellent optical readout for bioassays and molecular imaging. Herein, we accomplished new near infrared bioluminescence resonance energy transfer (NIR-BRET) templates for monitoring molecular events in cells with higher sensitivity. We first identified the best resonance energy donor for the NIR-BRET templates through the characterization of many coelenterazine (CTZ)–marine luciferase combinations. As a result, we found that NLuc–DBlueC and ALuc47–nCTZ combinations showed luminescence in the blue emission wavelength with excellent BL intensity and stability, for example, the NLuc–DBlueC and ALuc47–nCTZ combinations were 17-fold and 22-fold brighter than their second highest combinations, respectively, and were stably bright in living mammalian cells for at least 10 min. To harness the excellent BL properties to the NIR-BRET systems, NLuc and ALuc47 were genetically fused to fluorescent proteins (FPs), allowing large “blue-to-red” shifts, such as LSSmChe, LSSmKate2, and LSSmNep (where LSS means Large Stokes Shift). The excellent LSSmNep–NLuc combination showed approximately 170 nm large resonance energy shift from blue to red. The established templates were further utilized in the development of new NIR-BRET systems for imaging steroid hormone activities by sandwiching the ligand-binding domain of a nuclear receptor (NR-LBD) between the luciferase and the FP of the template. The NIR-BRET systems showed a specific luminescence signal upon exposure to steroid hormones, such as androgen, estrogen, and cortisol. The present NIR-BRET templates are important additions for utilizing their advantageous imaging of various molecular events with high efficiency and brightness in physiological samples. MDPI 2022-12-30 /pmc/articles/PMC9820568/ /pubmed/36614119 http://dx.doi.org/10.3390/ijms24010677 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Kim, Sung-Bae
Nishihara, Ryo
Paulmurugan, Ramasamy
Near-Infrared Imaging of Steroid Hormone Activities Using Bright BRET Templates
title Near-Infrared Imaging of Steroid Hormone Activities Using Bright BRET Templates
title_full Near-Infrared Imaging of Steroid Hormone Activities Using Bright BRET Templates
title_fullStr Near-Infrared Imaging of Steroid Hormone Activities Using Bright BRET Templates
title_full_unstemmed Near-Infrared Imaging of Steroid Hormone Activities Using Bright BRET Templates
title_short Near-Infrared Imaging of Steroid Hormone Activities Using Bright BRET Templates
title_sort near-infrared imaging of steroid hormone activities using bright bret templates
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9820568/
https://www.ncbi.nlm.nih.gov/pubmed/36614119
http://dx.doi.org/10.3390/ijms24010677
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