Biology subjects

Paillon, N.

Publications and source records attributed to Paillon, N..

3 recordsLinked to original sources

bioRxiv2025

The MARK2 kinase acts as a gatekeeper of CD28-dependent co-stimulation in T cells

Naive T cell activation requires not only antigen recognition through the TCR but also a co-stimulatory signal, mainly provided by CD28. Here, using a T cell-specific conditional knockout (cKO) model, we identify the microtubule-affinity kinase 2 (MARK2) as a key intracellular checkpoint that limits CD28-mediated co-stimulation. In vivo, MARK2 deficiency promotes the development of central memory T cells, enhances basal glycolysis activity in naive CD8 T cells, and leads to the development of systemic autoimmunity in aged mice. In MARK2-deficient CD8 T cells, TCR engagement alone drives sustained proliferation, cytokine production, and glycolysis, processes that normally require CD28 co-stimulation. Single-cell transcriptomic analysis reveals that MARK2 regulates the expression of genes involved in CD28 signaling and metabolic switch. We show that MARK2 restrains the PI3K-AKT-mTORC1 pathway by limiting CD28-driven transcriptional and metabolic programs. Mechanistically, we demonstrate that MARK2 phosphorylates CREB regulated transcription coactivator 2 (CRTC2) and suppresses CREB-mediated transcription and mTOR activation, whereas CD28 engagement lifts this inhibition. Together, our results redefine the role of CD28 that not only amplifies TCR signaling but also relieves a MARK2-dependent inhibitory signal. This work provides new insights into T cell activation, metabolism and immune tolerance with potential implications for immunotherapeutic strategies in cancer and autoimmunity.

immunology↗

bioRxiv2023

Label-free single-cell live imaging reveals fast metabolic switch in T lymphocytes

T cell activation induces a metabolic switch generating energy required for proliferation, survival, and fueling their functions. Thus, it is essential to monitor metabolism associated to subcellular functional and structural changes. We used non-invasive label-free two-photon fluorescence lifetime microscopy (2P-FLIM) to map the spatial and temporal dynamics of the metabolic NADH co-enzyme during T lymphocyte activation. 2P-FLIM measurements of the protein-bound and free NADH ratios provides a readout of the redox state (NAD+/ NADH) of the cells, and thus of their OXPHOS and glycolysis rates. Using this method, we followed the dynamics of fraction of bound NADH (fb NADH) in live single cells. Comparing fb NADH between resting and activated T cells, we show that T cell activation induces a rapid switch toward glycolysis. The switch takes only 10 minutes and remains stable for at least one hour. Three-dimensional (3D) analysis revealed that the intracellular distribution of fb NADH is symmetrically distributed in resting cells, whereas increases at the contact zone in activated cells. Finally, we show that fb NADH negatively correlates with spreading of activated T cells, suggesting a link between actin remodeling and metabolic changes. This study shows that 2P-FLIM measurement of fb NADH is well suited to follow a fast metabolic switch in 3D, in single T lymphocytes with subcellular resolution.

immunology↗

bioRxiv2019

Simultaneous tracking of division and differentiation from individual hematopoietic stem and progenitor cells reveals within-family homogeneity despite population heterogeneity

The advent of high throughput single cell methods such as scRNA-seq has uncovered substantial heterogeneity in the pool of hematopoietic stem and progenitor cells (HSPCs). A significant issue is how to reconcile those findings with the standard model of hematopoietic development, and a fundamental question is how much instruction is inherited by offspring from their ancestors. To address this, we further developed a high-throughput method that enables simultaneously determination of common ancestor, generation, and differentiation status of a large collection of single cells. Data from it revealed that while there is substantial population-level heterogeneity, cells that derived from a common ancestor were highly concordant in their division progression and share similar differentiation outcomes, revealing significant familial effects on both division and differentiation. Although each family diversifies to some extent, the overall collection of cell types observed in a population is largely composed of homogeneous families from heterogeneous ancestors. Heterogeneity between families could be explained, in part, by differences in ancestral expression of cell-surface markers that are used for phenotypic HSPC identification: CD48, SCA-1, c-kit and Flt3. These data call for a revision of the fundamental model of haematopoiesis from a single tree to an ensemble of trees from distinct ancestors where common ancestor effect must be considered. As HSPCs are cultured in the clinic before bone marrow transplantation, our results suggest that the broad range of engraftment and proliferation capacities of HSPCs could be consequences of the heterogeneity in their engrafted families, and altered culture conditions might reduce heterogeneity between families, possibly improving transplantation outcomes.

immunology↗