The fact that the identity of the cells that initiate metastasis in most human cancers is unknown hampers the development of antimetastatic therapies. Here we describe a subpopulation of CD44
cells ...in human oral carcinomas that do not overexpress mesenchymal genes, are slow-cycling, express high levels of the fatty acid receptor CD36 and lipid metabolism genes, and are unique in their ability to initiate metastasis. Palmitic acid or a high-fat diet specifically boosts the metastatic potential of CD36
metastasis-initiating cells in a CD36-dependent manner. The use of neutralizing antibodies to block CD36 causes almost complete inhibition of metastasis in immunodeficient or immunocompetent orthotopic mouse models of human oral cancer, with no side effects. Clinically, the presence of CD36
metastasis-initiating cells correlates with a poor prognosis for numerous types of carcinomas, and inhibition of CD36 also impairs metastasis, at least in human melanoma- and breast cancer-derived tumours. Together, our results indicate that metastasis-initiating cells particularly rely on dietary lipids to promote metastasis.
The human genome encodes hundreds of transfer RNA (tRNA) genes but their individual contribution to the tRNA pool is not fully understood. Deep sequencing of tRNA transcripts (tRNA-Seq) can estimate ...tRNA abundance at single gene resolution, but tRNA structures and posttranscriptional modifications impair these analyses. Here we present a bioinformatics strategy to investigate differential tRNA gene expression and use it to compare tRNA-Seq datasets from cultured human cells and human brain. We find that sequencing caveats affect quantitation of only a subset of human tRNA genes. Unexpectedly, we detect several cases where the differences in tRNA expression among samples do not involve variations at the level of isoacceptor tRNA sets (tRNAs charged with the same amino acid but using different anticodons), but rather among tRNA genes within the same isodecoder set (tRNAs having the same anticodon sequence). Because isodecoder tRNAs are functionally equal in terms of genetic translation, their differential expression may be related to noncanonical tRNA functions. We show that several instances of differential tRNA gene expression result in changes in the abundance of tRNA-derived fragments (tRFs) but not of mature tRNAs. Examples of differentially expressed tRFs include PIWI-associated RNAs, tRFs present in tissue samples but not in cells cultured in vitro, and somatic tissue-specific tRFs. Our data support that differential expression of tRNA genes regulate noncanonical tRNA functions performed by tRFs.
Most patients with colorectal cancer die as a result of the disease spreading to other organs. However, no prevalent mutations have been associated with metastatic colorectal cancers. Instead, ...particular features of the tumour microenvironment, such as lack of T-cell infiltration, low type 1 T-helper cell (T
1) activity and reduced immune cytotoxicity or increased TGFβ levels predict adverse outcomes in patients with colorectal cancer. Here we analyse the interplay between genetic alterations and the tumour microenvironment by crossing mice bearing conditional alleles of four main colorectal cancer mutations in intestinal stem cells. Quadruple-mutant mice developed metastatic intestinal tumours that display key hallmarks of human microsatellite-stable colorectal cancers, including low mutational burden, T-cell exclusion and TGFβ-activated stroma. Inhibition of the PD-1-PD-L1 immune checkpoint provoked a limited response in this model system. By contrast, inhibition of TGFβ unleashed a potent and enduring cytotoxic T-cell response against tumour cells that prevented metastasis. In mice with progressive liver metastatic disease, blockade of TGFβ signalling rendered tumours susceptible to anti-PD-1-PD-L1 therapy. Our data show that increased TGFβ in the tumour microenvironment represents a primary mechanism of immune evasion that promotes T-cell exclusion and blocks acquisition of the T
1-effector phenotype. Immunotherapies directed against TGFβ signalling may therefore have broad applications in treating patients with advanced colorectal cancer.
The expression of the pluripotency factors OCT4, SOX2, KLF4, and MYC (OSKM) can convert somatic differentiated cells into pluripotent stem cells in a process known as reprogramming. Notably, partial ...and reversible reprogramming does not change cell identity but can reverse markers of aging in cells, improve the capacity of aged mice to repair tissue injuries, and extend longevity in progeroid mice. However, little is known about the mechanisms involved. Here, we have studied changes in the DNA methylome, transcriptome, and metabolome in naturally aged mice subject to a single period of transient OSKM expression. We found that this is sufficient to reverse DNA methylation changes that occur upon aging in the pancreas, liver, spleen, and blood. Similarly, we observed reversion of transcriptional changes, especially regarding biological processes known to change during aging. Finally, some serum metabolites and biomarkers altered with aging were also restored to young levels upon transient reprogramming. These observations indicate that a single period of OSKM expression can drive epigenetic, transcriptomic, and metabolomic changes toward a younger configuration in multiple tissues and in the serum.
A single cycle of transient OSKM activation in naturally aged mice is able to partially reverse age‐associated changes in several tissues. Specifically, we could capture reversion of alterations occurring with aging at the level of DNA methylation, transcription, as well as, serum metabolome. These changes were stable for a period of up to four weeks after OSKM cessation.
The analysis of stem cell hierarchies in human cancers has been hampered by the impossibility of identifying or tracking tumor cell populations in an intact environment. To overcome this limitation, ...we devised a strategy based on editing the genomes of patient‐derived tumor organoids using CRISPR/Cas9 technology to integrate reporter cassettes at desired marker genes. As proof of concept, we engineered human colorectal cancer (CRC) organoids that carry EGFP and lineage‐tracing cassettes knocked in the LGR5 locus. Analysis of LGR5‐EGFP+ cells isolated from organoid‐derived xenografts demonstrated that these cells express a gene program similar to that of normal intestinal stem cells and that they propagate the disease to recipient mice very efficiently. Lineage‐tracing experiments showed that LGR5+ CRC cells self‐renew and generate progeny over long time periods that undergo differentiation toward mucosecreting‐ and absorptive‐like phenotypes. These genetic experiments confirm that human CRCs adopt a hierarchical organization reminiscent of that of the normal colonic epithelium. The strategy described herein may have broad applications to study cell heterogeneity in human tumors.
Synopsis
To overcome technical and conceptual limitations in the analysis of tumor cell heterogeneity, the genomes of human cancer organoids were edited to integrate reporter and lineage tracing cassettes that enable tracking selected tumor cell populations in vivo.
CRISPR/Cas9‐mediated genome editing facilitated the integration of cassettes at marker genes in colorectal cancer (CRC) patient‐derived organoids (PDOs).
A GFP reporter integrated at the LGR5 locus labeled a population of ISC‐like tumor cells in human CRCs.
The LGR5‐GFP+ cell population was enriched in tumor‐initiating cells.
Lineage tracing analysis using CreERT2 knock‐in PDOs demonstrated that in intact tumors LGR5+ CRC cells are multipotent, long‐lived and produce cell clones that scale proportional to tumor growth.
Double edited LGR5‐GFP/Ki67‐RFP PDOs revealed the existence of proliferating and quiescent LGR5+ CRC cells.
To overcome technical and conceptual limitations in the analysis of tumor cell heterogeneity, the genomes of human cancer organoids were edited to integrate reporter and lineage tracing cassettes that enable tracking selected tumor cell populations in vivo.
The generation of multiciliated cells (MCCs) is required for the proper function of many tissues, including the respiratory tract, brain, and germline. Defects in MCC development have been ...demonstrated to cause a subclass of mucociliary clearance disorders termed reduced generation of multiple motile cilia (RGMC). To date, only two genes, Multicilin (MCIDAS) and cyclin O (CCNO) have been identified in this disorder in humans. Here, we describe mice lacking GEMC1 (GMNC), a protein with a similar domain organization as Multicilin that has been implicated in DNA replication control. We have found that GEMC1‐deficient mice are growth impaired, develop hydrocephaly with a high penetrance, and are infertile, due to defects in the formation of MCCs in the brain, respiratory tract, and germline. Our data demonstrate that GEMC1 is a critical regulator of MCC differentiation and a candidate gene for human RGMC or related disorders.
Synopsis
In addition to a role in DNA replicaion, Geminin‐like coiled‐coil containing protein 1 (GEMC1) interacts with E2F4/5‐DP1 and Multicilin to control transcriptional programs required for multiciliated cell (MCC) differentiation in mammals.
GEMC1 is required for normal growth and fertility in mice.
Multiciliated cells and sperm require GEMC1.
GEMC1 interacts with E2F4/5‐DP1 and Multicilin.
Multiciliated cells transcriptional programs are activated by GEMC1.
GEMC1 is a candidate gene for human ciliopathies.
In addition to a role in DNA replicaion, Geminin‐like coiled‐coil containing protein 1 (GEMC1) interacts with E2F4/5‐DP1 and Multicilin to control transcriptional programs required for multiciliated cell (MCC) differentiation in mammals.
Highly proliferative Lgr5+ stem cells maintain the intestinal epithelium and are thought to be largely homogeneous. Although quiescent intestinal stem cell (ISC) populations have been described, the ...identity and features of such a population remain controversial. Here we report unanticipated heterogeneity within the Lgr5+ ISC pool. We found that expression of the RNA-binding protein Mex3a labels a slowly cycling subpopulation of Lgr5+ ISCs that contribute to all intestinal lineages with distinct kinetics. Single-cell transcriptome profiling revealed that Lgr5+ cells adopt two discrete states, one of which is defined by a Mex3a expression program and relatively low levels of proliferation genes. During homeostasis, Mex3a+ cells continually shift into the rapidly dividing, self-renewing ISC pool. Chemotherapy and radiation preferentially target rapidly dividing Lgr5+ cells but spare the Mex3a-high/Lgr5+ population, helping to promote regeneration of the intestinal epithelium following toxic insults. Thus, Mex3a defines a reserve-like ISC population within the Lgr5+ compartment.
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•Mex3a is expressed in a subset of Lgr5+ cells that proliferate slowly•Mex3a+ cells are multipotent and can generate all intestinal lineages•Mex3a+ cells can convert to fast-dividing ISCs to maintain homeostasis•Mex3a+ cells regenerate the epithelium after chemotherapeutic insult
Lgr5+ intestinal stem cells are considered to be a homogeneous and rapidly proliferating population. Barriga et al. show that the RNA binding protein Mex3a defines a subset of slowly proliferating Lgr5+ cells that contribute to all intestinal lineages with slow kinetics, are resistant to chemotherapy, and support intestinal regeneration.
Cellular senescence is a stress response that activates innate immune cells, but little is known about its interplay with the adaptive immune system. Here, we show that senescent cells combine ...several features that render them highly efficient in activating dendritic cells (DC) and antigen-specific CD8 T cells. This includes the release of alarmins, activation of IFN signaling, enhanced MHC class I machinery, and presentation of senescence-associated self-peptides that can activate CD8 T cells. In the context of cancer, immunization with senescent cancer cells elicits strong antitumor protection mediated by DCs and CD8 T cells. Interestingly, this protection is superior to immunization with cancer cells undergoing immunogenic cell death. Finally, the induction of senescence in human primary cancer cells also augments their ability to activate autologous antigen-specific tumor-infiltrating CD8 lymphocytes. Our study indicates that senescent cancer cells can be exploited to develop efficient and protective CD8-dependent antitumor immune responses.
Our study shows that senescent cells are endowed with a high immunogenic potential-superior to the gold standard of immunogenic cell death. We harness these properties of senescent cells to trigger efficient and protective CD8-dependent antitumor immune responses. See related article by Chen et al., p. 432. This article is highlighted in the In This Issue feature, p. 247.
Recent molecular classifications of colorectal cancer (CRC) based on global gene expression profiles have defined subtypes displaying resistance to therapy and poor prognosis. Upon evaluation of ...these classification systems, we discovered that their predictive power arises from genes expressed by stromal cells rather than epithelial tumor cells. Bioinformatic and immunohistochemical analyses identify stromal markers that associate robustly with disease relapse across the various classifications. Functional studies indicate that cancer-associated fibroblasts (CAFs) increase the frequency of tumor-initiating cells, an effect that is dramatically enhanced by transforming growth factor (TGF)-β signaling. Likewise, we find that all poor-prognosis CRC subtypes share a gene program induced by TGF-β in tumor stromal cells. Using patient-derived tumor organoids and xenografts, we show that the use of TGF-β signaling inhibitors to block the cross-talk between cancer cells and the microenvironment halts disease progression.
The specification of tissue identity during embryonic development requires precise spatio-temporal coordination of gene expression. Many transcription factors required for the development of organs ...have been identified and their expression patterns are known; however, the mechanisms through which they coordinate gene expression in time remain poorly understood. Here, we show that hormone-induced transcription factor Blimp-1 participates in the temporal coordination of tubulogenesis in
by regulating the expression of many genes involved in tube maturation. In particular, we demonstrate that Blimp-1 regulates the expression of genes involved in chitin deposition and F-actin organization. We show that Blimp-1 is involved in the temporal control of lumen maturation by regulating the beginning of chitin deposition. We also report that Blimp-1 represses a variety of genes involved in tracheal maturation. Finally, we reveal that the kinase Btk29A serves as a link between Blimp-1 transcriptional repression and apical extracellular matrix organization.
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