scholarly journals Spectral Analysis of ATP-Dependent Mechanical Vibrations in T Cells

2021 ◽  
Vol 9 ◽  
Author(s):  
Ishay Wohl ◽  
Eilon Sherman
Keyword(s):  
T Cells ◽  
Spectral Analysis ◽  
Plasma Membrane ◽  
Mean Square Displacement ◽  
Mechanical Work ◽  
Mechanical Vibrations ◽  
Effector Cells ◽  
Immune Synapse ◽  
Content Organization ◽  
And Function

Mechanical vibrations affect multiple cell properties, including its diffusivity, entropy, internal content organization, and thus—function. Here, we used Differential Interference Contrast (DIC), confocal, and Total Internal Reflection Fluorescence (TIRF) microscopies to study mechanical vibrations in live (Jurkat) T cells. Vibrations were measured via the motion of intracellular particles and plasma membrane. These vibrations depend on adenosine triphosphate (ATP) consumption and on Myosin II activity. We then used spectral analysis of these vibrations to distinguish the effects of thermal agitation, ATP-dependent mechanical work and cytoskeletal visco-elasticity. Parameters of spectral analyses could be related to mean square displacement (MSD) analyses with specific advantages in characterizing intracellular mechanical work. We identified two spectral ranges where mechanical work dominated vibrations of intracellular components: 0–3 Hz for intracellular particles and the plasma-membrane, and 100–150 Hz for the plasma-membrane. The 0–3 Hz vibrations of the cell membrane that we measured in an experimental model of immune synapse (IS) are expected to affect the IS formation and function in effector cells. It may also facilitate immunological escape of extensively vibrating malignant cells.

scholarly journals T-cells play the classics with a different spin

2014 ◽  
Vol 25 (11) ◽  
pp. 1699-1703 ◽  
Author(s):  
Michael L. Dustin
Keyword(s):  
T Cells ◽  
Plasma Membrane ◽  
Immune Cells ◽  
Cell Biology ◽  
Immunological Synapse ◽  
Cell Communication ◽  
Intraflagellar Transport ◽  
Cell Types ◽  
Immune Synapse ◽  
And Function

The immune system uses much of the classic machinery of cell biology, but in ways that put a different spin on organization and function. Striking recent examples include the demonstration of intraflagellar transport protein and hedgehog contributions to the immune synapse, even though immune cells lack a primary cilium that would be the typical setting for this machinery. In a second example, lymphocytes have their own subfamily of integrins, the β2 subfamily, and only integrins in this family form a stable adhesion ring using freely mobile ligands, a key feature of the immunological synapse. Finally, we showed recently that T-cells use endosomal sorting complexes required for transport (ESCRTs) at the plasma membrane to generate T-cell antigen receptor–enriched microvesicles. It is unusual for the ESCRT pathway to operate at the plasma membrane, but this may allow a novel form of cell–cell communication by providing a multivalent ligand for major histocompatibility complex–peptide complexes and perhaps other receptors on the partnering B-cell. Immune cells are thus an exciting system for novel cell biology even with classical pathways that have been studied extensively in other cell types.

scholarly journals S-acylation targets ORAI1 channels to lipid rafts for efficient Ca2+ signaling by T cell receptors at the immune synapse

2021 ◽  
Author(s):  
Amado Carreras-Sureda ◽  
Laurence Abrami ◽  
Ji-Hee Kim ◽  
Maud Frieden ◽  
Monica Didier ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Receptors ◽  
Hek 293 ◽  
Immune Synapse ◽  
Cell Receptors ◽  
Channel Assembly ◽  
And Function ◽  
Antigen Presenting ◽  
Reduced Mobility

AbstractEfficient immune responses require Ca2+ fluxes across ORAI1 channels during engagement of T cell receptors (TCR) at the immune synapse (IS) between T cells and antigen presenting cells. Here, we show that ZDHHC20-mediated S-acylation of the ORAI1 channel at residue Cys143 is required for TCR assembly and signaling at the IS. Cys143 mutations reduced ORAI1 currents and store-operated Ca2+ entry in HEK-293 cells and nearly abrogated long-lasting Ca2+ elevations, NFATC1 translocation, and IL-2 secretion evoked by TCR engagement in Jurkat T cells. The acylation-deficient channel had reduced mobility in lipids, accumulated in cholesterol-poor domains, formed tiny clusters, failed to reach the IS and unexpectedly also prevented TCR recruitment to the IS. Our results establish S-acylation as a critical regulator of ORAI1 channel assembly and function at the IS and reveal that local Ca2+ fluxes are required for TCR recruitment to the synapse.

scholarly journals Augmented TCR-mediated signaling in infant T cells enables robust responses to respiratory virus infection

2021 ◽  
Author(s):  
Donna Farber ◽  
Puspa Thapa ◽  
Rebecca Guyer ◽  
Alexander Yang ◽  
Christopher Parks ◽  
...  
Keyword(s):  
T Cells ◽  
Immune Responses ◽  
Influenza Infection ◽  
Life Stage ◽  
Cell Receptor ◽  
Human Infant ◽  
Transfer Model ◽  
Effector Cells ◽  
Immune Synapse

Abstract Infants require coordinated immune responses to prevent succumbing to multiple infectious challenges, particularly in the respiratory tract. The mechanisms by which infant T cells are functionally adapted for these responses are not well understood. Here, we demonstrate using an in vivo co-transfer model that infant T cells generate greater numbers of lung-homing effector cells to influenza infection compared to adult T cells in the same host, due to augmented TCF-1 downregulation and T cell receptor (TCR)-mediated signaling. Importantly, infant T cells show increased sensitivity to low antigen doses, originating at the interface between T cells and antigen-bearing accessory cells–through actin-mediated mobilization of signaling molecules to the immune synapse. This enhanced signaling was also observed in human infant versus adult T cells. Our findings provide a mechanism for how infants control pathogen load and dissemination, important for designing developmentally-appropriate strategies for promoting immune responses at this vulnerable life stage.

scholarly journals Regulatory T cell expansion by a highly CD25-dependent IL-2 mutein arrests ongoing autoimmunity

2019 ◽  
Author(s):  
Liliane Khoryati ◽  
Minh Nguyet Pham ◽  
McKenna Sherve ◽  
Swarnima Kumari ◽  
Kevin Cook ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Dose Range ◽  
Interleukin 2 ◽  
Autoimmune Response ◽  
Wild Type ◽  
Effector Cells ◽  
Inflammatory Conditions ◽  
Receptor Component ◽  
And Function

AbstractInterleukin-2 (IL-2) controls the homeostasis and function of regulatory T cells (Tregs) and defects in the IL-2 pathway contribute to multiple autoimmune diseases. Although recombinant IL-2 therapy has been efficacious in certain inflammatory conditions, the capacity for IL-2 to also activate inflammatory effector responses highlights the need for IL-2-based therapeutics with improved Treg-specificity. From a panel of rationally designed IL-2 variants, we identified IL-2 muteins with reduced potency and enhanced Treg-selectivity due to increased dependence on the IL-2-receptor component CD25. As an Fc-fused homodimer, the optimal Fc.IL-2 mutein induced selective Treg enrichment and reduced agonism of effector cells across a wide dose range. Furthermore, despite being a weaker agonist, overall Treg growth was greater and more sustained due to reduced receptor-mediated clearance of the Fc.IL-2 mutein compared to Fc-fused wild-type IL-2. Preferential Treg enrichment was also observed in the presence of activated pathogenic T cells in the autoimmune target organ, despite a loss of Treg-selectivity in an IL-2R-proximal response. These features allowed for extended resolution of spontaneous autoimmunity using infrequent dosing schedules. Thus, IL-2 muteins enable efficient, flexible, and targeted control of the autoimmune response.One Sentence SummaryA CD25-dependent IL-2 mutein selectively expands regulatory T cells and provides potent and targeted control of autoimmunity.

scholarly journals S-acylation by ZDHHC20 targets ORAI1 channels to lipid rafts for efficient Ca2+ signaling by Jurkat T cell receptors at the immune synapse

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Amado Carreras-Sureda ◽  
Laurence Abrami ◽  
Kim Ji-Hee ◽  
Wen-An Wang ◽  
Christopher Henry ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Receptors ◽  
Hek 293 ◽  
Immune Synapse ◽  
Cell Receptors ◽  
Channel Trafficking ◽  
293 Cells ◽  
And Function ◽  
Antigen Presenting

Efficient immune responses require Ca2+ fluxes across ORAI1 channels during engagement of T cell receptors (TCR) at the immune synapse (IS) between T cells and antigen presenting cells. Here, we show that ZDHHC20-mediated S-acylation of the ORAI1 channel at residue Cys143 promotes TCR recruitment and signaling at the IS. Cys143 mutations reduced ORAI1 currents and store-operated Ca2+ entry in HEK-293 cells and nearly abrogated long-lasting Ca2+ elevations, NFATC1 translocation, and IL-2 secretion evoked by TCR engagement in Jurkat T cells. The acylation-deficient channel remained in cholesterol-poor domains upon enforced ZDHHC20 expression and was recruited less efficiently to the IS along with actin and TCR. Our results establish S-acylation as a critical regulator of ORAI1 channel trafficking and function at the IS and reveal that ORAI1 S-acylation enhances TCR recruitment to the synapse.

scholarly journals An IL-2 mutein engineered to promote expansion of regulatory T cells arrests ongoing autoimmunity in mice

2020 ◽  
Vol 5 (50) ◽  
pp. eaba5264 ◽  
Author(s):  
Liliane Khoryati ◽  
Minh Nguyet Pham ◽  
McKenna Sherve ◽  
Swarnima Kumari ◽  
Kevin Cook ◽  
...  
Keyword(s):  
T Cells ◽  
Dose Range ◽  
Interleukin 2 ◽  
Nod Mice ◽  
Effector Cells ◽  
Inflammatory Conditions ◽  
Cell Selectivity ◽  
Receptor Component ◽  
Cell Enrichment ◽  
And Function

Interleukin-2 (IL-2) controls the homeostasis and function of regulatory T (Treg) cells, and defects in the IL-2 pathway contribute to multiple autoimmune diseases. Although recombinant IL-2 therapy has been efficacious in certain inflammatory conditions, the capacity for IL-2 to also activate inflammatory effector responses highlights the need for IL-2–based therapeutics with improved Treg cell specificity. From a panel of rationally designed murine IL-2 variants, we identified IL-2 muteins with reduced potency and enhanced Treg cell selectivity due to increased dependence on the IL-2 receptor component CD25. As an Fc-fused homodimer, the optimal Fc.IL-2 mutein induced selective Treg cell enrichment and reduced agonism of effector cells across a wide dose range. Furthermore, despite being a weaker agonist, overall Treg cell growth was greater and more sustained due to reduced receptor-mediated clearance of the Fc.IL-2 mutein compared with Fc-fused wild-type IL-2. Preferential Treg cell enrichment was also observed in the presence of activated pathogenic T cells in the pancreas of nonobese diabetic (NOD) mice, despite a loss of Treg cell selectivity in an IL-2R proximal response. These properties facilitated potent and extended resolution of NOD diabetes with infrequent dosing schedules.

scholarly journals Role of mitotic diffusion barriers in regulating the asymmetric division of activated CD8 T cells

2021 ◽  
Author(s):  
Hulya Emurla ◽  
Yves Barral ◽  
Annette Oxenius
Keyword(s):  
T Cells ◽  
Plasma Membrane ◽  
Cd8 T Cells ◽  
Diffusion Barrier ◽  
Lateral Diffusion ◽  
Memory Cells ◽  
Effector Cells ◽  
Membrane Asymmetry

SummaryUpon their activation, naïve CD8 T cells divide and differentiate into short-lived effector cells, relevant for exerting immune control, and long-lived memory cells, relevant for long-term immunity. The proportion of memory cells generated depends highly on the context of activation and whether the activated cell divides symmetrically or asymmetrically. However, how T cells control the extent of their asymmetry during their first division in response to contextual signals is not known. Using fluorescence loss in photo-bleaching (FLIP) experiments, we show that the metabolic and plasma membrane asymmetry of mitotic T cells depend on the regulated assembly of a lateral diffusion barrier in their endoplasmic reticulum (ER-) membrane. In asymmetrically dividing T cells, the degrees of asymmetry correlated tightly to barrier strength, whereas symmetrically dividing T cells did not establish such a barrier. Direct positive or negative interference with barrier assembly enhanced or abrogated metabolic and plasma membrane asymmetry, respectively, indicating that barrier strength is a direct and decisive determinant of mitotic asymmetry. Thus, together our data identify diffusion barrier-mediated compartmentalization as a mechanism for how asymmetric T cell regulate their long-term response as a function of the activatory context.

scholarly journals Fluctuations in the Homogeneity of Cell Medium Distinguish Benign from Malignant Lymphocytes in a Cellular Model of Acute T Cells Leukemia

2020 ◽  
Vol 10 (24) ◽  
pp. 8894
Author(s):  
Ishay Wohl ◽  
Oren Yakovian ◽  
Eilon Sherman
Keyword(s):  
T Cells ◽  
Spectral Analysis ◽  
Mitochondrial Dysfunction ◽  
Information Entropy ◽  
Atp Depletion ◽  
Mechanical Work ◽  
Jurkat Cells ◽  
Image Correlation ◽  
Label Free ◽  
Cell Functions

Intracellular mechanical work facilitates multiple cell functions, such as material transport, cell motility, etc., and is indicative of the cell’s physiological condition. Still, the characterization of intracellular mechanical work and resultant dynamics remain hard to determine in intact label-free cells. For that, we imaged live T cells via bright-field microscopy and studied fluctuations in the homogeneity of their intracellular medium. Specifically, we characterized medium homogeneity and dynamics by using the information entropy of its related intensity gray levels (termed Gray Level Information Entropy (GLIE)) and spectral analysis of GLIE fluctuations, respectively. First, we provide simple examples of particle motion, to demonstrate the utility of our approach. Using this approach, we could further study and distinguish mitochondrial dysfunction and ATP depletion state in live Jurkat cells. The relation of our results to intracellular dynamics was confirmed by comparison to image correlation spectroscopy (ICS) results in the same cells. Importantly, GLIE fluctuations combined with spectral analysis enabled differentiation of malignant Jurkat cells from benign lymphocytes with 86% accuracy for single cells and 95% for populations of 10 cells each. Our approach can serve for label-free live-cell study and diagnostics of important pathophysiological conditions, such as mitochondrial dysfunction and malignancy.

scholarly journals Prevalence and Regulation of Effector CD8+ T Cell Subsets in B-Cell NHL

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 4134-4134
Author(s):  
Hongyan Wu ◽  
Zhi-Zhang Yang ◽  
Hyo Jin Kim ◽  
Shahrzad Jalali ◽  
Tammy Price-Troska ◽  
...  
Keyword(s):  
T Cells ◽  
B Cell ◽  
Cd8 T Cells ◽  
Research Funding ◽  
Granzyme B ◽  
Median Percentage ◽  
Effector Cells ◽  
Ifn Γ ◽  
And Function ◽  
Lower Expression

Abstract Effector CD8+ T cells play a crucial role in anti-tumor immunity. Two major subsets of effector CD8+ T cells have been identified based on expression of KLRG1 and CD127: KLRG1+CD127- short-lived effector cells (SLECs) and KLRG1-CD127+ memory precursor effector cells (MPECs). In B-cell non-Hodgkin lymphoma (NHL), the frequency and function of SLECs and MPECs are unknown. Furthermore, the underlying mechanism by which these two CD8+ subsets were regulated and differentiate in B-cell NHL remains poorly understood. The goal of the present study is therefore to phenotypically and functionally characterize SLECs and MPECs in B-cell NHL. Using patient biopsy specimens, we observed that the median percentage of SLECs was greater than MPECs and was 39.9% (range: 22.7%~ 54.7%) and 18% (range: 7.66%~31.6%), respectively. In controls, the median percentage of SLECs was lower than that of MPECs and was 13.71% (range: 5.26%~29.2%) and 51.63% (range: 22.5%~70.1%) in PBMC and 12.14% (range: 4.01%~26%) and 51.62% (range: 29.2%~70.3%) in tonsil, respectively. Using mass cytometry (CyTOF), we observed that SLECs have higher expression levels of TIGIT and PD-1 and lower expression of CCR7, CD26 and CD27 when compared to MPECs. SLECs were functionally superior to MPECs as the numbers of cytokine (IFN-γ and TNF-α) - and granule (granzyme B and perforin)-producing cells were higher in SLECs than MPECs. However, SLECs had a lower proliferative capacity and a higher apoptosis rate when compared to MPECs. We observed that a reciprocal differentiation pathway exists between SLECs and MPECs. Activation and cytokine stimulation (IL-2 and IL-15) promoted the development of SLECs and decreased the number of MPECs. This effect was reversed when cells were treated with neutralizing antibodies to block IL-2 or IL-15 signaling. We also found that these two subsets had a distinct transcription factor profile as SLECs had higher expression of Eomes and T-bet, and lower expression of Tcf-1than MPECs. IL-2 and IL-15 enhanced the expression of T-bet and decreased the expression of Tcf-1. Taken together, our results show that SLECs are more prevalent than MPECs in B-cell NHL when compared to normal control tissue. These two effector cell subsets have distinct phenotypical and function profiles and their development is controlled by cytokines that may be dysregulated in lymphoma. Despite the fact that SLECs produce more granzyme B and IFN-γ, they are less proliferative and more susceptible to apoptosis. This may compromise an effective anti-tumor immune response in B-cell NHL. Disclosures Ansell: Celldex: Research Funding; Takeda: Research Funding; Merck & Co: Research Funding; Affimed: Research Funding; Bristol-Myers Squibb: Research Funding; Trillium: Research Funding; Regeneron: Research Funding; LAM Therapeutics: Research Funding; Pfizer: Research Funding; Seattle Genetics: Research Funding.

scholarly journals Reduced Numbers and Impaired Function of Regulatory T Cells in Peripheral Blood of Ischemic Stroke Patients

2016 ◽  
Vol 2016 ◽  
pp. 1-9 ◽  
Author(s):  
Johanna Ruhnau ◽  
Juliane Schulze ◽  
Bettina von Sarnowski ◽  
Marie Heinrich ◽  
Sönke Langner ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Peripheral Blood ◽  
Experimental Stroke ◽  
Stroke Patients ◽  
Effector Cells ◽  
T Effector Cells ◽  
Treg Function ◽  
Age Dependent ◽  
And Function

Background and Purpose. Regulatory T cells (Tregs) have been suggested to modulate stroke-induced immune responses. However, analyses of Tregs in patients and in experimental stroke have yielded contradictory findings. We performed the current study to assess the regulation and function of Tregs in peripheral blood of stroke patients. Age dependent expression of CD39 on Tregs was quantified in mice and men. Methods. Total FoxP3+ Tregs and CD39+FoxP3+ Tregs were quantified by flow cytometry in controls and stroke patients on admission and on days 1, 3, 5, and 7 thereafter. Treg function was assessed by quantifying the inhibition of activation-induced expression of CD69 and CD154 on T effector cells (Teffs). Results. Total Tregs accounted for 5.0% of CD4+ T cells in controls and <2.8% in stroke patients on admission. They remained below control values until day 7. CD39+ Tregs were most strongly reduced in stroke patients. On day 3 the Treg-mediated inhibition of CD154 upregulation on CD4+ Teff was impaired in stroke patients. CD39 expression on Treg increased with age in peripheral blood of mice and men. Conclusion. We demonstrate a loss of active FoxP3+CD39+ Tregs from stroke patient’s peripheral blood. The suppressive Treg function of remaining Tregs is impaired after stroke.

Sign in / Sign up

Export Citation Format

Share Document