Donor regulatory T cells rapidly adapt to recipient tissues to control murine acute graft-versus-host disease.

The adoptive transfer of regulatory T cells is a promising strategy to prevent graft-versus-host disease after allogeneic bone marrow transplantation. Here, we use a major histocompatibility complex-mismatched mouse model to follow the fate of in vitro expanded donor regulatory T cells upon migration to target organs. Employing comprehensive gene expression and repertoire profiling, we show that they retain their suppressive function and plasticity after transfer. Upon entering non-lymphoid tissues, donor regulatory T cells acquire organ-specific gene expression profiles resembling tissue-resident cells and activate hallmark suppressive and cytotoxic pathways, most evidently in the colon, when co-transplanted with graft-versus-host disease-inducing conventional T cells. Dominant T cell receptor clonotypes overlap between organs and across recipients and their relative abundance correlates with protection efficacy. Thus, this study reveals donor regulatory T cell selection and adaptation mechanisms in target organs and highlights protective features of Treg to guide the development of improved graft-versus-host disease prevention strategies.

The effector program of human CD8 T cells supports tissue remodeling.

CD8 T lymphocytes are classically viewed as cytotoxic T cells. Whether human CD8 T cells can, in parallel, induce a tissue regeneration program is poorly understood. Here, antigen-specific assay systems revealed that human CD8 T cells not only mediated cytotoxicity but also promoted tissue remodeling. Activated CD8 T cells could produce the epidermal growth factor receptor (EGFR)-ligand amphiregulin (AREG) and sensitize epithelial cells for enhanced regeneration potential. Blocking the EGFR or the effector cytokines IFN-γ and TNF could inhibit tissue remodeling. This regenerative program enhanced tumor spheroid and stem cell-mediated organoid growth. Using single-cell gene expression analysis, we identified an AREG+, tissue-resident CD8 T cell population in skin and adipose tissue from patients undergoing abdominal wall or abdominoplasty surgery. These tissue-resident CD8 T cells showed a strong TCR clonal relation to blood PD1+TIGIT+ CD8 T cells with tissue remodeling abilities. These findings may help to understand the complex CD8 biology in tumors and could become relevant for the design of therapeutic T cell products.

Biosensors for inflammation as a strategy to engineer regulatory T cells for cell therapy.

Engineered regulatory T cell (Treg cell) therapy is a promising strategy to treat patients suffering from inflammatory diseases, autoimmunity, and transplant rejection. However, in many cases, disease-related antigens that can be targeted by Treg cells are not available. In this study, we introduce a class of synthetic biosensors, named artificial immune receptors (AIRs), for murine and human Treg cells. AIRs consist of three domains: (a) extracellular binding domain of a tumor necrosis factor (TNF)-receptor superfamily member, (b) intracellular costimulatory signaling domain of CD28, and (c) T cell receptor signaling domain of CD3-ζ chain. These AIR receptors equip Treg cells with an inflammation-sensing machinery and translate this environmental information into a CD3-ζ chain-dependent TCR-activation program. Different AIRs were generated, recognizing the inflammatory ligands of the TNF-receptor superfamily, including LIGHT, TNFα, and TNF-like ligand 1A (TL1A), leading to activation, differentiation, and proliferation of AIR-Treg cells. In a graft-versus-host disease model, Treg cells expressing lymphotoxin ß receptor-AIR, which can be activated by the ligand LIGHT, protect significantly better than control Treg cells. Expression and signaling of the corresponding human AIR in human Treg cells prove that this concept can be translated. Engineering Treg cells that target inflammatory ligands leading to TCR signaling and activation might be used as a Treg cell-based therapy approach for a broad range of inflammation-driven diseases.

Single-cell chromatin accessibility landscape identifies tissue repair program in human regulatory T cells.

Murine regulatory T (Treg) cells in tissues promote tissue homeostasis and regeneration. We sought to identify features that characterize human Treg cells with these functions in healthy tissues. Single-cell chromatin accessibility profiles of murine and human tissue Treg cells defined a conserved, microbiota-independent tissue-repair Treg signature with a prevailing footprint of the transcription factor BATF. This signature, combined with gene expression profiling and TCR fate mapping, identified a population of tissue-like Treg cells in human peripheral blood that expressed BATF, chemokine receptor CCR8 and HLA-DR. Human BATF+CCR8+ Treg cells from normal skin and adipose tissue shared features with nonlymphoid T follicular helper-like (Tfh-like) cells, and induction of a Tfh-like differentiation program in naive human Treg cells partially recapitulated tissue Treg regenerative characteristics, including wound healing potential. Human BATF+CCR8+ Treg cells from healthy tissue share features with tumor-resident Treg cells, highlighting the importance of understanding the context-specific functions of these cells.

The long non-coding RNA LINC00941 and SPRR5 are novel regulators of human epidermal homeostasis.

Several long non-coding RNAs (lncRNAs) act as regulators of cellular homeostasis; however, few of these molecules were functionally characterized in a mature human tissue environment. Here, we report that the lncRNA LINC00941 is a crucial regulator of human epidermal homeostasis. LINC00941 is enriched in progenitor keratinocytes and acts as a repressor of keratinocyte differentiation. Furthermore, LINC00941 represses SPRR5, a previously uncharacterized molecule, which functions as an essential positive regulator of keratinocyte differentiation. Interestingly, 54.8% of genes repressed in SPRR5-deficient epidermal tissue are induced in LINC00941-depleted organotypic epidermis, suggesting a common mode of action for both molecules.

Zygotic Genome Activation Occurs Shortly after Fertilization in Maize.

The formation of a zygote via the fusion of an egg and sperm cell and its subsequent asymmetric division herald the start of the plant's life cycle. Zygotic genome activation (ZGA) is thought to occur gradually, with the initial steps of zygote and embryo development being primarily maternally controlled, and subsequent steps being governed by the zygotic genome. Here, using maize (Zea mays) as a model plant system, we determined the timing of zygote development and generated RNA-seq transcriptome profiles of gametes, zygotes, and apical and basal daughter cells. ZGA occurs shortly after fertilization and involves ∼10% of the genome being activated in a highly dynamic pattern. In particular, genes encoding transcriptional regulators of various families are activated shortly after fertilization. Further analyses suggested that chromatin assembly is strongly modified after fertilization, that the egg cell is primed to activate the translational machinery, and that hormones likely play a minor role in the initial steps of early embryo development in maize. Our findings provide important insights into gamete and zygote activity in plants, and our RNA-seq transcriptome profiles represent a comprehensive, unique RNA-seq data set that can be used by the research community.

Phosphorylation of Argonaute proteins affects mRNA binding and is essential for microRNA-guided gene silencing in vivo.

Argonaute proteins associate with microRNAs and are key components of gene silencing pathways. With such a pivotal role, these proteins represent ideal targets for regulatory post-translational modifications. Using quantitative mass spectrometry, we find that a C-terminal serine/threonine cluster is phosphorylated at five different residues in human and Caenorhabditis elegans In human, hyper-phosphorylation does not affect microRNA binding, localization, or cleavage activity of Ago2. However, mRNA binding is strongly affected. Strikingly, on Ago2 mutants that cannot bind microRNAs or mRNAs, the cluster remains unphosphorylated indicating a role at late stages of gene silencing. In C. elegans, the phosphorylation of the conserved cluster of ALG-1 is essential for microRNA function in vivo Furthermore, a single point mutation within the cluster is sufficient to phenocopy the loss of its complete phosphorylation. Interestingly, this mutant retains its capacity to produce and bind microRNAs and represses expression when artificially tethered to an mRNA Altogether, our data suggest that the phosphorylation state of the serine/threonine cluster is important for Argonaute-mRNA interactions.

Nucleosome maps of the human cytomegalovirus genome reveal a temporal switch in chromatin organization linked to a major IE protein.

Human CMV (hCMV) establishes lifelong infections in most of us, causing developmental defects in human embryos and life-threatening disease in immunocompromised individuals. During productive infection, the viral >230,000-bp dsDNA genome is expressed widely and in a temporal cascade. The hCMV genome does not carry histones when encapsidated but has been proposed to form nucleosomes after release into the host cell nucleus. Here, we present hCMV genome-wide nucleosome occupancy and nascent transcript maps during infection of permissive human primary cells. We show that nucleosomes occupy nuclear viral DNA in a nonrandom and highly predictable fashion. At early times of infection, nucleosomes associate with the hCMV genome largely according to their intrinsic DNA sequence preferences, indicating that initial nucleosome formation is genetically encoded in the virus. However, as infection proceeds to the late phase, nucleosomes redistribute extensively to establish patterns mostly determined by nongenetic factors. We propose that these factors include key regulators of viral gene expression encoded at the hCMV major immediate-early (IE) locus. Indeed, mutant virus genomes deficient for IE1 expression exhibit globally increased nucleosome loads and reduced nucleosome dynamics compared with WT genomes. The temporal nucleosome occupancy differences between IE1-deficient and WT viruses correlate inversely with changes in the pattern of viral nascent and total transcript accumulation. These results provide a framework of spatial and temporal nucleosome organization across the genome of a major human pathogen and suggest that an hCMV major IE protein governs overall viral chromatin structure and function.