T cells are key players in our defence against infections and malignancies. When T cells differentiate or become activated, they undergo substantial alterations in gene expression. Even though RNA ...expression levels are now well documented throughout different stages of T cells, it is not well understood how mRNA expression translates into the protein landscape. By combining paired RNA sequencing and mass spectrometry data of primary human CD8
+
T cells, we report that mRNA expression is a poor proxy for the overall protein output, irrespective of the differentiation or activation status. Yet, gene class stratification revealed a function-specific correlation of mRNA with protein expression. This gene class-specific expression pattern associated with differences in gene characteristics such as sequence conservation and untranslated region (UTR) lengths. In addition, the presence of AU-rich elements in the 3’UTR associated with alterations in mRNA and protein abundance T cell activation dependent, gene class-specific manner. In conclusion, our study highlights the role of gene characteristics as a determinant for gene expression in T cells.
Effective T cell responses against infections and tumors require a swift and ample production of cytokines, chemokines, and cytotoxic molecules. The production of these effector molecules relies on ...rapid changes of gene expression, determined by cell-intrinsic signals and environmental cues. Here, we review our current understanding of gene-specific regulatory networks that define the magnitude and timing of cytokine production in CD8+ T cells. We discuss the dynamic features of post-transcriptional control during CD8+ T cell homeostasis and activation, and focus on the crosstalk between cell signaling and RNA-binding proteins. Elucidating gene-specific regulatory circuits may help in the future to rectify dysfunctional T cell responses.
The rapid remodeling of the T cell proteome upon activation depends on the integration of transcriptional and post-transcriptional control of gene expression.Post-transcriptional events are heterogeneous, dynamic, transcript specific, and cell type specific.RNA-binding proteins mediate post-transcriptional events and their activity is strictly regulated by signal transduction.Post-transcriptional control preserves the silent state of memory T cells, and tailors the magnitude and kinetics of cytokine production in effector T cells.Upon chronic insults, CD8+ T cells can fail to respond, partially because cytokine mRNA is not stabilized and translated.
Abstract
Hematopoietic stem cells differentiate into a broad range of specialized blood cells. This process is tightly regulated and depends on transcription factors, micro-RNAs, and long non-coding ...RNAs. Recently, also circular RNA (circRNA) were found to regulate cellular processes. Their expression pattern and their identity is however less well defined. Here, we provide the first comprehensive analysis of circRNA expression in human hematopoietic progenitors, and in differentiated lymphoid and myeloid cells. We here show that the expression of circRNA is cell-type specific, and increases upon maturation. CircRNA splicing variants can also be cell-type specific. Furthermore, nucleated hematopoietic cells contain circRNA that have higher expression levels than the corresponding linear RNA. Enucleated blood cells, i.e. platelets and erythrocytes, were suggested to use RNA to maintain their function, respond to environmental factors or to transmit signals to other cells via microvesicles. Here we show that platelets and erythrocytes contain the highest number of circRNA of all hematopoietic cells, and that the type and numbers of circRNA changes during maturation. This cell-type specific expression pattern of circRNA in hematopoietic cells suggests a hithero unappreciated role in differentiation and cellular function.
Clinical implementation of tumor organoids for personalized medicine requires that pure tumor organoids can be reliably established. Here, we present our experience with organoid cultures from >70 ...non-small cell lung cancer (NSCLC) samples. We systematically evaluate several methods to identify tumor purity of organoids established from intrapulmonary tumors. Eighty percent of organoids from intrapulmonary lesions have a normal copy number profile, suggesting overgrowth by normal airway organoids (AOs). This is further supported by the failure to detect mutations found in the original tumor in organoids. Histomorphology alone is insufficient to determine tumor purity, but when combined with p63 immunostaining, tumor and normal AOs can be distinguished. Taking into account overgrowth by normal AOs, the establishment rate of pure NSCLC organoids is 17%. Therefore, current methods are insufficient to establish pure NSCLC organoids from intrapulmonary lesions. We discourage their use unless steps are taken to prevent overgrowth by normal AOs.
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•Frequent overgrowth of lung cancer organoids by normal airway organoids•Copy number profiling and IHC can be used to distinguish tumor and normal organoids•Low establishment rate of pure lung cancer organoids limits their clinical utility
Using a combination of genetic analyses and immunohistochemistry, Dijkstra et al. find that the majority of organoids derived from intrapulmonary lung cancers are overgrown by normal airway organoids. This limits the overall establishment rate of pure lung cancer organoids to 17%, restricting their utility for personalized medicine.
T cells are central players of the adaptive immune system by protecting us from recurring infections and by killing malignant cells. Protective T cell responses rely on the concerted production of ...effector molecules such as cytolytic mediators, granzymes, and perforins, as well as pro‐inflammatory cytokines and chemokines. Once activated, T cells drastically change their gene expression and rapidly respond to insults by producing ample amounts of effector molecules. In the absence of antigen, T cells remain in a quiescent state and survey our body for possible pathogenic insults. Resting T cells are, however, not inert, but continuously regulate their protein production to survive and to be prepared for possible re‐infections. Here, we review our current knowledge on the regulation of gene expression in activated and quiescent T cells. We specifically focus on post‐transcriptional mechanisms that define the protein output and that allow dormant cells to undergo active signaling and selective translation, keeping them poised for activation. Finally, we discuss which signals drive T cell survival and their preparedness to respond to insults and which mechanisms are involved in these processes.
T cells continuously receive signals for protein production. Although quiescent T cells produce selected proteins needed for homeostasis, activated T cells undergo substantial proteome remodeling when activated through the TCR. Here, we describe how post‐transcriptional mechanisms define the protein production in T cells, with a focus on RNA‐binding proteins (RBPs).
Vagal nerve efferent activation has been shown to ameliorate the course of many inflammatory disease states. This neuro-modulatory effect has been suggested to rest on acetylcholine receptor (AChR) ...activation on tissue macrophages or dendritic cells (DCs). In more recent studies, vagal anti-inflammatory activity was shown involve adrenergic, splenic, pathways. Here we provide evidence that the adrenergic, rather than cholinergic, receptor activation on bone marrow derived DCs results in enhanced endocytosis uptake, enhanced IL-10 production but a decreased IL-6, IL-12p70 and IL-23 production. In antigen specific T cell stimulation assays, adrenergic β2 receptor activation on bone marrow DCs led to an enhanced potential to induce Foxp3 positive suppressive Treg cells. These effects were independent of IL10-R activation, TGFβ release, or retinoic acid (RA) secretion. Hence, adrenergic receptor β2 activation modulates DC function resulting in skewing towards anti-inflammatory T cell phenotypes.
Cancer immunotherapies have shown substantial clinical activity for a subset of patients with epithelial cancers. Still, technological platforms to study cancer T-cell interactions for individual ...patients and understand determinants of responsiveness are presently lacking. Here, we establish and validate a platform to induce and analyze tumor-specific T cell responses to epithelial cancers in a personalized manner. We demonstrate that co-cultures of autologous tumor organoids and peripheral blood lymphocytes can be used to enrich tumor-reactive T cells from peripheral blood of patients with mismatch repair-deficient colorectal cancer and non-small-cell lung cancer. Furthermore, we demonstrate that these T cells can be used to assess the efficiency of killing of matched tumor organoids. This platform provides an unbiased strategy for the isolation of tumor-reactive T cells and provides a means by which to assess the sensitivity of tumor cells to T cell-mediated attack at the level of the individual patient.
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•Induction of tumor-reactive T cells by co-culture of PBMCs and tumor organoids•Induced T cell populations do not recognize healthy organoids or tissue•This platform can be used to assess the efficiency of T-cell-mediated tumor killing
A modified patient-derived tumor organoids system allows the expansion of tumor-specific T cells from blood for personalized analysis of their anti-cancer properties.
CD8(+) T cells can respond to unrelated infections in an Ag-independent manner. This rapid innate-like immune response allows Ag-experienced T cells to alert other immune cell types to pathogenic ...intruders. In this study, we show that murine CD8(+) T cells can sense TLR2 and TLR7 ligands, resulting in rapid production of IFN-γ but not of TNF-α and IL-2. Importantly, Ag-experienced T cells activated by TLR ligands produce sufficient IFN-γ to augment the activation of macrophages. In contrast to Ag-specific reactivation, TLR-dependent production of IFN-γ by CD8(+) T cells relies exclusively on newly synthesized transcripts without inducing mRNA stability. Furthermore, transcription of IFN-γ upon TLR triggering depends on the activation of PI3K and serine-threonine kinase Akt, and protein synthesis relies on the activation of the mechanistic target of rapamycin. We next investigated which energy source drives the TLR-induced production of IFN-γ. Although Ag-specific cytokine production requires a glycolytic switch for optimal cytokine release, glucose availability does not alter the rate of IFN-γ production upon TLR-mediated activation. Rather, mitochondrial respiration provides sufficient energy for TLR-induced IFN-γ production. To our knowledge, this is the first report describing that TLR-mediated bystander activation elicits a helper phenotype of CD8(+) T cells. It induces a short boost of IFN-γ production that leads to a significant but limited activation of Ag-experienced CD8(+) T cells. This activation suffices to prime macrophages but keeps T cell responses limited to unrelated infections.
CD29 Enriches for Cytotoxic Human CD4+ T Cells Nicolet, Benoît P.; Guislain, Aurelie; Wolkers, Monika C.
The Journal of immunology (1950),
12/2021, Volume:
207, Issue:
12
Journal Article
Peer reviewed
Open access
Abstract
CD4+ T cells are key contributors in the induction of adaptive immune responses against pathogens. Even though CD4+ T cells are primarily classified as noncytotoxic helper T cells, it has ...become appreciated that a subset of CD4+ T cells is cytotoxic. However, tools to identify these cytotoxic CD4+ T cells are lacking. We recently showed that CD29 (integrin β1, ITGB1) expression on human CD8+ T cells enriches for the most potent cytotoxic T cells. In this study, we questioned whether CD29 expression also associates with cytotoxic CD4+ T cells. We show that human peripheral blood–derived CD29hiCD4+ T cells display a cytotoxic gene expression profile, which closely resembles that of CD29hi cytotoxic CD8+ T cells. This CD29hi cytotoxic phenotype was observed ex vivo and was maintained in in vitro cultures. CD29 expression enriched for CD4+ T cells, which effectively produced the proinflammatory cytokines IFN-γ, IL-2, and TNF-α, and cytotoxic molecules. Lastly, CD29-expressing CD4+ T cells transduced with a MART1-specific TCR showed target cell killing in vitro. In conclusion, we demonstrate in this study that CD29 can be employed to enrich for cytotoxic human CD4+ T cells.
B cells and T cells are key players in the defence against infections and malignancies. To exert their function, B cells and T cells differentiate into effector and memory cells. Tight regulation of ...these differentiation processes is key to prevent their malfunction, which can result in life-threatening disease. Lymphocyte differentiation relies on the appropriate timing and dosage of regulatory molecules, and post-transcriptional gene regulation (PTR) is a key player herein. PTR includes the regulation through RNA-binding proteins (RBPs), which control the fate of RNA and its translation into proteins. To date, a comprehensive overview of the RBP expression throughout lymphocyte differentiation is lacking. Using transcriptome and proteome analyses, we here catalogued the RBP expression for human B cells and T cells. We observed that even though the overall RBP expression is conserved, the relative RBP expression is distinct between B cells and T cells. Differentiation into effector and memory cells alters the RBP expression, resulting into preferential expression of different classes of RBPs. For instance, whereas naive T cells express high levels of translation-regulating RBPs, effector T cells preferentially express RBPs that modulate mRNA stability. Lastly, we found that cytotoxic CD8
and CD4
T cells express a common RBP repertoire. Combined, our study reveals a cell type-specific and differentiation-dependent RBP expression landscape in human lymphocytes, which will help unravel the role of RBPs in lymphocyte function.
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