{"title":"Expansion microscopy reveals thylakoid organisation alterations due to genetic mutations and far-red light acclimation\\.","authors":"Jarne Berentsen, Peter R Bos, Emilie Wientjes","doi":"10.1016/j.bbabio.2025.149552","DOIUrl":null,"url":null,"abstract":"<p><p>The thylakoid membrane is the site of the light-dependent reactions of photosynthesis. It is a continuous membrane, folded into grana stacks and the interconnecting stroma lamellae. The CURVATURE THYLAKOID1 (CURT1) protein family is involved in the folding of the membrane into the grana stacks. The thylakoid membrane remodels its architecture in response to light conditions, but its 3D organisation and dynamics remain incompletely understood. To resolve these details, an imaging technique is needed that provides high-resolution 3D images in a high-throughput manner. Recently, we have used expansion microscopy, a technique that meets these criteria, to visualise the thylakoid membrane isolated from spinach. Here, we show that this protocol can also be used to visualise enveloped spinach chloroplasts. Additionally, we present an improved protocol for resolving the thylakoid structure of Arabidopsis thaliana. Using this protocol, we show the changes in thylakoid architecture in response to long-term far-red light acclimation and due to knocking out CURT1A. We show that far-red light acclimation results in higher grana stacks that are packed closer together. In addition, the distance between stroma lamellae, which are wrapped around the grana, decreases. In the curt1a mutant, grana have an increased diameter and height, and the distance between grana is increased. Interestingly, in this mutant, the stroma lamellae occasionally approach the grana stacks from the top. These observations show the potential of expansion microscopy to study the thylakoid membrane architecture.</p>","PeriodicalId":50731,"journal":{"name":"Biochimica et Biophysica Acta-Bioenergetics","volume":" ","pages":"149552"},"PeriodicalIF":3.4000,"publicationDate":"2025-03-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Biochimica et Biophysica Acta-Bioenergetics","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1016/j.bbabio.2025.149552","RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
The thylakoid membrane is the site of the light-dependent reactions of photosynthesis. It is a continuous membrane, folded into grana stacks and the interconnecting stroma lamellae. The CURVATURE THYLAKOID1 (CURT1) protein family is involved in the folding of the membrane into the grana stacks. The thylakoid membrane remodels its architecture in response to light conditions, but its 3D organisation and dynamics remain incompletely understood. To resolve these details, an imaging technique is needed that provides high-resolution 3D images in a high-throughput manner. Recently, we have used expansion microscopy, a technique that meets these criteria, to visualise the thylakoid membrane isolated from spinach. Here, we show that this protocol can also be used to visualise enveloped spinach chloroplasts. Additionally, we present an improved protocol for resolving the thylakoid structure of Arabidopsis thaliana. Using this protocol, we show the changes in thylakoid architecture in response to long-term far-red light acclimation and due to knocking out CURT1A. We show that far-red light acclimation results in higher grana stacks that are packed closer together. In addition, the distance between stroma lamellae, which are wrapped around the grana, decreases. In the curt1a mutant, grana have an increased diameter and height, and the distance between grana is increased. Interestingly, in this mutant, the stroma lamellae occasionally approach the grana stacks from the top. These observations show the potential of expansion microscopy to study the thylakoid membrane architecture.
期刊介绍:
BBA Bioenergetics covers the area of biological membranes involved in energy transfer and conversion. In particular, it focuses on the structures obtained by X-ray crystallography and other approaches, and molecular mechanisms of the components of photosynthesis, mitochondrial and bacterial respiration, oxidative phosphorylation, motility and transport. It spans applications of structural biology, molecular modeling, spectroscopy and biophysics in these systems, through bioenergetic aspects of mitochondrial biology including biomedicine aspects of energy metabolism in mitochondrial disorders, neurodegenerative diseases like Parkinson''s and Alzheimer''s, aging, diabetes and even cancer.