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The T cell receptor resides in small ordered plasma membrane domains that aggregate upon T cell activation
Stockholm University, Faculty of Science, The Wenner-Gren Institute .
Uppsala universitet, Institutionen för medicinsk cellbiologi .
Stockholm University, Faculty of Science, The Wenner-Gren Institute .
(English)Manuscript (preprint) (Other academic)
Abstract [en]

T cell signaling emanates from large lipid raft platforms. Whether lipid rafts form upon T cell receptor (TCR) engagement or exist in resting T cells was the focus of this study. Plasma membrane order was followed in live T cells at 37°C using laurdan to report on lipid packing. Patching of the TCR in both Jurkat and human primary CD4+ T cells resulted in higher fractions of ordered plasma domains in the patches but did not increase the overall membrane order. The TCR colocalized with actin filaments in unstimulated Jurkat T cells and this colocalization was most prominent for cells in G1 phase. Moreover, the TCR located to the nuclear envelope, in addition to the plasma membrane, in cells in S and G2/M phase. Our study suggests that the TCR resides in ordered plasma membrane domains/lipid rafts that are linked to actin filament and aggregate upon T cell activation.

Keyword [en]
actin, generalised polarisation, lipid rafts, membrane order, T cell receptor
National Category
Cell Biology
Research subject
Cellbiology
Identifiers
URN: urn:nbn:se:su:diva-62288OAI: oai:DiVA.org:su-62288DiVA: diva2:441172
Available from: 2011-09-14 Created: 2011-09-13 Last updated: 2011-09-16Bibliographically approved
In thesis
1. Plasma membrane order; the role of cholesterol and links to actin filaments:  
Open this publication in new window or tab >>Plasma membrane order; the role of cholesterol and links to actin filaments:  
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The connection between T cell activation, plasma membrane order and actin filament dynamics was the main focus of this study. Laurdan and di-4-ANEPPDHQ, membrane order sensing probes, were shown to report only on lipid packing rather than being influenced by the presence of membrane-inserted peptides justifying their use in membrane order studies. These dyes were used to follow plasma membrane order in live cells at 37°C. Disrupting actin filaments had a disordering effect while stabilizing actin filaments had an ordering effect on the plasma membrane, indicating there is a basal level of ordered domains in resting cells. Lowering PI(4,5)P2 levels decreased the proportion of ordered domains strongly suggesting that the connection of actin filaments to the plasma membrane is responsible for the maintaining the level of ordered membrane domains. Membrane blebs, which are detached from the underlying actin filaments, contained a low fraction of ordered domains. Aggregation of membrane components resulted in a higher proportion of ordered plasma membrane domains and an increase in cell peripheral actin polymerization. This strongly suggests that the attachment of actin filaments to the plasma membrane induces the formation of ordered domains. Limited cholesterol depletion with methyl-beta-cyclodextrin triggered peripheral actin polymerization. Cholesterol depleted cells showed an increase in plasma membrane order as a result of actin filament accumulation underneath the membrane. Moderate cholesterol depletion also induced membrane domain aggregation and activation of T cell signaling events. The T cell receptor (TCR) aggregation caused redistribution of domains resulting in TCR patches of higher order and the bulk membrane correspondingly depleted of ordered domains. This suggests the preexistence of small ordered membrane domains in resting T cells that aggregate upon cell activation. Increased actin polymerization at the TCR aggregation sites showed that actin polymerization is strongly correlated with the changes in the distribution of ordered domains. The distribution of the TCR in resting cells and its colocalization with actin filaments is cell cycle dependent. We conclude that actin filament attachment to the plasma membrane, which is regulated via PI(4,5)P2, plays a crucial role in the formation of ordered domains.

Place, publisher, year, edition, pages
Stockholm: The Wenner-Gren Institute, Stockholm University, 2011. 56 p.
Keyword
Membrane Organization, Lipid rafts, Actin, Laurdan, di-4-ANEPPDHQ, Cholesterol, T cell signaling, Colocalization, Generalized Polarization
National Category
Cell Biology
Research subject
Cellbiology
Identifiers
urn:nbn:se:su:diva-62279 (URN)978-91-7447-365-0 (ISBN)
Public defence
2011-10-14, E306, Arrheniuslaboratorierna, Svante Arrhenius väg 20 C, Stockholm, 10:00 (English)
Opponent
Supervisors
Note
At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 2: Submitted. Paper 4: Manuscript. Available from: 2011-09-22 Created: 2011-09-13 Last updated: 2011-09-16Bibliographically approved

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