Intracellular Macromolecular Crowding within Individual Stress Fibers Analyzed by Fluorescence Correlation Spectroscopy

Buenaventura, Aria; Saito, Takumi; Kanao, Taiga; Matsunaga, Daiki; Matsui, Tsubasa S.; Deguchi, Shinji

doi:https://doi.org/10.1007/s12195-024-00803-4

Intracellular Macromolecular Crowding within Individual Stress Fibers Analyzed by Fluorescence Correlation Spectroscopy

Buenaventura, Aria; Saito, Takumi; Kanao, Taiga et al.

Cellular and Molecular Bioengineering, 2024, 17(3), 165-176

Number of Access:507（2025-07-18 14:18 Counts）

Identifier to cite or link to this item: https://hdl.handle.net/11094/97601

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Title	Intracellular Macromolecular Crowding within Individual Stress Fibers Analyzed by Fluorescence Correlation Spectroscopy
Creator	CreatorName Buenaventura, Aria
	CreatorName Saito, Takumi
	CreatorName Kanao, Taiga
	NRID 1000040833794 1000040833794 ORCID 0000-0002-6407-7766 0000-0002-6407-7766 researchmap permalink daikimatsunaga daikimatsunaga Scopus Author ID 55497164000 55497164000 CreatorName Matsunaga, Daiki
	NRID 1000050638707 1000050638707 ORCID 0000-0002-6699-9131 0000-0002-6699-9131 researchmap permalink ts.mat29 ts.mat29 Scopus Author ID 35248391600 35248391600 CreatorName Matsui, Tsubasa S.
	NRID 1000030379713 1000030379713 ORCID 0000-0002-0556-4599 0000-0002-0556-4599 researchmap permalink read0138586 read0138586 Scopus Author ID 8645720800 8645720800 CreatorName Deguchi, Shinji
Subject	Autocorrelation
	FCS
	Macromolecular crowding
	Nucleus
	Protein diffusion
	Stress fibers
Description	The version of record of this article, first published in Cellular and Molecular Bioengineering, is available online at Publisher’s website: https://doi.org/10.1007/s12195-024-00803-4.
Abstract	Introduction: The diffusion of cell components such as proteins is crucial to the function of all living cells. The abundance of macromolecules in cells is likely to cause a state of macromolecular crowding, but its effects on the extent of diffusion remain poorly understood. Methods: Here we investigate the diffusion rate in three distinct locations in mesenchymal cell types, namely the open cytoplasm, the stress fibers in the open cytoplasm, and those below the nucleus using three kinds of biologically inert green fluorescent proteins (GFPs), namely a monomer, dimer, and trimer GFP. Fluorescence correlation spectroscopy (FCS) was used to determine the diffusion coefficients. Results: We show that diffusion tends to be lowered on average in stress fibers and is significantly lower in those located below the nucleus. Our data suggest that the diffusive properties of GFPs, and potentially other molecules as well, are hindered by macromolecular crowding. However, although the size dependence on protein diffusion was also studied for monomer, dimer, and trimer GFPs, there was no significant difference in the diffusion rates among the GFPs of these sizes. These results could be attributed to the lack of significant change in protein size among the selected GFP multimers. Conclusion: The data presented here would provide a basis for better understanding of the complex protein diffusion in the nonuniform cytoplasm, shedding light on cellular responses to mechanical stress, their local mechanical properties, and reduced turnover in senescent cells.
Publisher	Springer
Source Title	Cellular and Molecular Bioengineering
Volume (Issue)	17(3)
Page	165-176
Date of Issued	2024-06-07
Language	English
Handle URL	https://hdl.handle.net/11094/97601
PISSN	18655025
eISSN	18655033
Relation.isIdenticalTo	DOI (Publisher Version) https://doi.org/10.1007/s12195-024-00803-4
Access Rights	open access
CopyRight	This article is licensed under a Creative Commons Attribution 4.0 International License.
oaire:version	Version of Record
Category	学術雑誌論文 Journal Article

Resource Type	journal article
Local Type	Journal Article
DCTERMS.bibliographicCitation	Cellular and Molecular Bioengineering. 2024, 17(3), p. 165-176
DC.title	Intracellular Macromolecular Crowding within Individual Stress Fibers Analyzed by Fluorescence Correlation Spectroscopy
DC.creator	Buenaventura, Aria
	Saito, Takumi
	Kanao, Taiga
	Matsunaga, Daiki
	Matsui, Tsubasa S.
	Deguchi, Shinji
DC.publisher	Springer
DC.language' scheme='DCTERMS.RFC1766	English
DCTERMS.issued' scheme='DCTERMS.W3CDTF	2024-06-07
DC.identifier	https://doi.org/10.1007/s12195-024-00803-4
DC.subject	Autocorrelation
	FCS
	Macromolecular crowding
	Nucleus
	Protein diffusion
	Stress fibers
DC.description	The version of record of this article, first published in Cellular and Molecular Bioengineering, is available online at Publisher’s website: https://doi.org/10.1007/s12195-024-00803-4.
DCTERMS.abstract	Introduction: The diffusion of cell components such as proteins is crucial to the function of all living cells. The abundance of macromolecules in cells is likely to cause a state of macromolecular crowding, but its effects on the extent of diffusion remain poorly understood. Methods: Here we investigate the diffusion rate in three distinct locations in mesenchymal cell types, namely the open cytoplasm, the stress fibers in the open cytoplasm, and those below the nucleus using three kinds of biologically inert green fluorescent proteins (GFPs), namely a monomer, dimer, and trimer GFP. Fluorescence correlation spectroscopy (FCS) was used to determine the diffusion coefficients. Results: We show that diffusion tends to be lowered on average in stress fibers and is significantly lower in those located below the nucleus. Our data suggest that the diffusive properties of GFPs, and potentially other molecules as well, are hindered by macromolecular crowding. However, although the size dependence on protein diffusion was also studied for monomer, dimer, and trimer GFPs, there was no significant difference in the diffusion rates among the GFPs of these sizes. These results could be attributed to the lack of significant change in protein size among the selected GFP multimers. Conclusion: The data presented here would provide a basis for better understanding of the complex protein diffusion in the nonuniform cytoplasm, shedding light on cellular responses to mechanical stress, their local mechanical properties, and reduced turnover in senescent cells.
DC.rights	This article is licensed under a Creative Commons Attribution 4.0 International License.
DC.format	97601
citation_title	Intracellular Macromolecular Crowding within Individual Stress Fibers Analyzed by Fluorescence Correlation Spectroscopy
citation_author	Buenaventura, Aria
	Saito, Takumi
	Kanao, Taiga
	Matsunaga, Daiki
	Matsui, Tsubasa S.
	Deguchi, Shinji
citation_publisher	Springer
citation_language	English
citation_date	2024-06-07
citation_journal_title	Cellular and Molecular Bioengineering
citation_volume	17
citation_issue	3
citation_firstpage	165
citation_lastpage	176
citation_doi	https://doi.org/10.1007/s12195-024-00803-4
citation_public_url	https://hdl.handle.net/11094/97601

Intracellular Macromolecular Crowding within Individual Stress Fibers Analyzed by Fluorescence Correlation Spectroscopy

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