Gioana Litscher’s research while affiliated with University of Zurich and other places

Group 1 innate lymphoid cells (ILCs) encompass NK cells and ILC1s, which have non-redundant roles in host protection against pathogens and cancer. Despite their circulating nature, NK cells can establish residency in selected tissues during ontogeny, forming a distinct functional subset. The mechanisms that initiate, maintain, and regulate the conversion of NK cells into tissue-resident NK (trNK) cells are currently not well understood. Here, we identify autocrine transforming growth factor-β (TGF-β) as a cell-autonomous driver for NK cell tissue residency across multiple glandular tissues during development. Cell-intrinsic production of TGF-β was continuously required for the maintenance of trNK cells and synergized with Hobit to enhance cytotoxic function. Whereas autocrine TGF-β was redundant in tumors, our study revealed that NK cell–derived TGF-β allowed the expansion of cytotoxic trNK cells during local infection with murine cytomegalovirus (MCMV) and contributed to viral control in the salivary gland. Collectively, our findings reveal tissue-specific regulation of trNK cell differentiation and function by autocrine TGF-β1, which is relevant for antiviral immunity.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) has a non-redundant role in the emergence and maintenance of alveolar macrophages (AMs). However, its role in developmental and steady-state myelopoiesis outside the lung is largely unexplored. Scanning through developing tissues using a Fate-map and reporter of GM-CSF mouse strain, we discovered that GM-CSF was produced by type 2 innate lymphoid cells (ILC2s) in the submandibular and sublingual salivary gland (SG) during postnatal development. GM-CSF producing ILC2s foster the development of a hitherto undescribed phagocyte subset, which we named adenophages. Detailed analysis focusing on phenotypic and transcriptional profiling revealed that adenophages display shared aspects of both, macrophages and dendritic cells (DCs). We found them to be homogenously distributed across the SG, but always in close proximity to GM-CSF producing ILC2s and myoepithelial cells. Importantly, adenophages were present throughout all analyzed exocrine glands such as lacrimal glands and mammary glands, and were also identified in human SG sections, indicating a conserved role in exocrine glands across species.

Metabolic-associated fatty liver disease (MAFLD) is a spectrum of clinical manifestations ranging from benign steatosis to cirrhosis. A key event in the pathophysiology of MAFLD is the development of nonalcoholic steatohepatitis (NASH), which can potentially lead to fibrosis and hepatocellular carcinoma, but the triggers of MAFLD-associated inflammation are not well understood. We have observed that lipid accumulation in hepatocytes induces expression of ligands specific to the activating immune receptor NKG2D. Tissue-resident innate-like T cells, most notably γδ T cells, are activated through NKG2D and secrete IL-17A. IL-17A licenses hepatocytes to produce chemokines that recruit proinflammatory cells into the liver, which causes NASH and fibrosis. NKG2D-deficient mice did not develop fibrosis in dietary models of NASH and had a decreased incidence of hepatic tumors. The frequency of IL-17A ⁺ γδ T cells in the blood of patients with MAFLD correlated directly with liver pathology. Our findings identify a key molecular mechanism through which stressed hepatocytes trigger inflammation in the context of MAFLD.

Group 1 innate lymphoid cells (ILCs) comprising circulating natural killer (cNK) cells and tissue-resident ILC1s are critical for host defense against pathogens and tumors. Despite a growing understanding of their role in homeostasis and disease, the ontogeny of group 1 ILCs remains largely unknown. Here, we used fate mapping and single-cell transcriptomics to comprehensively investigate the origin and turnover of murine group 1 ILCs. Whereas cNK cells are continuously replaced throughout life, we uncovered tissue-dependent development and turnover of ILC1s. A first wave of ILC1s emerges during embryogenesis in the liver and transiently colonizes fetal tissues. After birth, a second wave quickly replaces ILC1s in most tissues apart from the liver, where they layer with embryonic ILC1s, persist until adulthood, and undergo a specific developmental program. Whereas embryonically derived ILC1s give rise to a cytotoxic subset, the neonatal wave establishes the full spectrum of ILC1s. Our findings uncover key ontogenic features of murine group 1 ILCs and their association with cellular identities and functions.

Aging exerts profound and paradoxical effects on the immune system, at once impairing proliferation, cytotoxicity and phagocytosis, and inducing chronic inflammation. Previous studies have focused on individual tissues or cell types, while a comprehensive multisystem study of tissue-resident and circulating immune populations during aging is lacking. Here we reveal an atlas of age-related changes in the abundance and phenotype of immune cell populations across 12 mouse tissues. Using cytometry-based high parametric analysis of 37 mass-cytometry and 55 spectral flow-cytometry parameters, mapping samples from young and aged animals revealed conserved and tissue-type-specific patterns of both immune atrophy and expansion. We uncovered clear phenotypic changes in both lymphoid and myeloid lineages in aged mice, and in particular a contraction in natural killer cells and plasmacytoid dendritic cells. These changes correlated with a skewing towards myelopoiesis at the expense of early lymphocyte genesis in aged mice. Taken together, this atlas represents a comprehensive, systematic and thorough resource of the age-dependent alterations of the mammalian immune system in lymphoid, barrier and solid tissues.

Formation of cytoplasmic RNA-protein structures called stress granules (SGs) is a highly conserved cellular response to stress. Abnormal metabolism of SGs may contribute to the pathogenesis of (neuro)degenerative diseases such as amyotrophic lateral sclerosis (ALS). Many SG proteins are affected by mutations causative of these conditions, including fused in sarcoma (FUS). Mutant FUS variants have high affinity to SGs and also spontaneously form de novo cytoplasmic RNA granules. Mutant FUS-containing assemblies (mFAs), often called “pathological SGs”, are proposed to play a role in ALS-FUS pathogenesis. However, structural differences between mFAs and physiological SGs remain largely unknown therefore it is unclear whether mFAs can functionally substitute for SGs and how they affect cellular stress responses. Here we used affinity purification to isolate mFAs and physiological SGs and compare their protein composition. We found that proteins within mFAs form significantly more physical interactions than those in SGs however mFAs fail to recruit many factors involved in signal transduction. Furthermore, we found that proteasome subunits and certain nucleocytoplasmic transport factors are depleted from mFAs, whereas translation elongation, mRNA surveillance and splicing factors as well as mitochondrial proteins are enriched in mFAs, as compared to SGs. Validation experiments for a mFA-specific protein, hnRNPA3, confirmed its RNA-dependent interaction with FUS and its sequestration into FUS inclusions in cultured cells and a FUS transgenic mouse model. Silencing of the Drosophila hnRNPA3 ortholog was deleterious and potentiated human FUS toxicity in the retina of transgenic flies. In conclusion, we show that SG-like structures formed by mutant FUS are structurally distinct from SGs, prone to persistence, likely cannot functionally replace SGs, and affect a spectrum of cellular pathways in stressed cells. Results of our study support a pathogenic role for cytoplasmic FUS assemblies in ALS-FUS.

Significance Innate lymphoid cells hold great promise for the treatment of metastases. Development of effective therapies based on these versatile immune cells, however, is hampered by our limited knowledge of their behavior in the metastatic niche. Here, we describe that defense against liver metastasis requires collaboration between two innate lymphocyte subsets, conventional NK cells (cNKs) and tissue-resident type I innate lymphoid cells (trILC1s). We show that different cancers generate their own particular metastatic niche–inducing specific changes in cNKs and trILC1s. Furthermore, we uncover specific cNK subsets that can be manipulated to improve their antimetastatic potential. Our work contributes to understanding how cancer-specific factors and hepatic innate lymphocytes exert mutual influence and how this can be exploited for therapeutic purposes.

Formation of cytoplasmic RNA-protein structures called stress granules (SGs) is a highly conserved cellular response to stress. Abnormal metabolism of SGs may contribute to the pathogenesis of (neuro)degenerative diseases such as amyotrophic lateral sclerosis (ALS). Many SG proteins are affected by mutations causative of these conditions, including fused in sarcoma (FUS). Mutant FUS variants have high affinity to SGs and also spontaneously form de novo cytoplasmic RNA granules. Mutant FUS-containing assemblies (mFAs), often called "pathological SGs", are proposed to play a role in ALS-FUS pathogenesis. However, global structural differences between mFAs and physiological SGs remain largely unknown, therefore it is unclear whether and how mFAs may affect cellular stress responses. Here we used affinity purification to characterise the protein and RNA composition of normal SGs and mFAs purified from stressed cells. Comparison of the SG and mFA proteomes revealed that proteasome subunits and certain nucleocytoplasmic transport factors are depleted from mFAs, whereas translation elongation, mRNA surveillance and splicing factors as well as mitochondrial proteins are enriched in mFAs, as compared to SGs. Validation experiments for a hit from our analysis, a splicing factor hnRNPA3, confirmed its RNA-dependent sequestration into mFAs in cells and into pathological FUS inclusions in a FUS transgenic mouse model. Furthermore, silencing of the Drosophila hnRNPA3 ortholog dramatically enhanced FUS toxicity in transgenic flies. Comparative transcriptomic analysis of SGs and mFAs revealed that mFAs recruit a significantly less diverse spectrum of RNAs, including reduced recruitment of transcripts encoding proteins involved in protein translation, DNA damage response, and apoptotic signalling. However mFAs abnormally sequester certain mRNAs encoding proteins involved in stress signalling cascades. Overall, our study establishes molecular differences between physiological SGs and mFAs and identifies the spectrum of proteins, RNAs and respective cellular pathways affected by mFAs in stressed cells. In conclusion, we show that mFAs are compositionally distinct from SGs and that they cannot fully substitute for SG functions while gaining novel, potentially toxic functions in cellular stress response. Results of our study support a pathogenic role for stress-induced cytoplasmic FUS assemblies in ALS-FUS.

The liver is a major metastatic target organ, and little is known about the role of immunity in controlling hepatic metastases. Here, we discovered that the concerted and non-redundant action of two innate lymphocyte subpopulations, conventional NK cells (cNKs) and tissue-resident type I Innate Lymphoid Cells (trILC1s), is essential for anti-metastatic defense. Using different preclinical models for liver metastasis, we found that trILC1 control metastatic seeding, whereas cNKs restrain outgrowth. The antimetastatic activity of cNKs is regulated in a tumor type-specific fashion. Thereby, individual cancer cell lines orchestrate the emergence of cNK subsets with unique phenotypic and functional traits. Understanding cancer-cell- as well as innate-cell-intrinsic factors will allow the exploitation of hepatic innate cells for development of novel cancer therapies. Significance Innate lymphoid cells hold great promise for the treatment of metastases. Development of effective therapies based on these versatile immune cells, however, is hampered by our limited knowledge of their behavior in the metastatic niche. Here, we describe that defense against liver metastasis requires the collaboration between two innate lymphocyte subsets, conventional NK cells (cNKs) and tissue-resident type I innate lymphoid cells (trILC1s). We show that different cancers generate their own particular metastatic niche inducing specific changes in cNKs and trILC1s. Further, we uncover specific cNK subsets that can be manipulated to improve their anti-metastatic potential. Our work contributes to understanding how cancer-specific factors and hepatic innate lymphocytes exert mutual influence and how this can be exploited for therapeutic purposes. Highlights cNKs and trILC1s collaborate to control hepatic metastasis trILC1s restrict seeding and cNKs control outgrowth of cancer cells in the liver Individual cancer cell lines orchestrate a distinct metastatic niche The metastatic niche dictates the phenotype and function of cNKs