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An Amendment to this paper has been published and can be accessed via a link at the top of the paper., Author(s): Saiyu Hang [sup.1], Donggi Paik [sup.1], Lina Yao [sup.2], Eunha Kim [sup.1], Jamma Trinath [sup.3], Jingping Lu [sup.4], Soyoung Ha [sup.1], Brandon N. Nelson [sup.5], Samantha P. Kelly [sup.5], [...]

Here we report a unique cellular reprogramming phenomenon, called stimulus-triggered acquisition of pluripotency (STAP), which requires neither nuclear transfer nor the introduction of transcription factors. In STAP, strong external stimuli such as a transient low-pH stressor reprogrammed mammalian somatic cells, resulting in the generation of pluripotent cells. Through real-time imaging of STAP cells derived from purified lymphocytes, as well as gene rearrangement analysis, we found that committed somatic cells give rise to STAP cells by reprogramming rather than selection. STAP cells showed a substantial decrease in DNA methylation in the regulatory regions of pluripotency marker genes. Blastocyst injection showed that STAP cells efficiently contribute to chimaeric embryos and to offspring via germline transmission. We also demonstrate the derivation of robustly expandable pluripotent cell lines from STAP cells. Thus, our findings indicate that epigenetic fate determination of mammalian cells can be markedly converted in a context-dependent manner by strong environmental cues. One of two papers describing a reprogramming phenomenon called stimulus-triggered acquisition of pluripotency (STAP) -- in STAP, lineage-committed adult somatic cells are reprogrammed to pluripotency by transient exposure to low-pH treatment, and extensive analysis of the molecular features and developmental potential of STAP cells indicates that they represent a unique state of pluripotency. A new way to induce pluripotency The fates of the somatic cells that form the bulk of the mammalian body are thought to be largely determined by the time the cellular differentiation processes of development have been completed. Reprogramming in response to environmental stress has been observed in plants but not so far in mammalian cells. Now two manuscripts by Haruko Obokata and colleagues describe an unexpected reprogramming phenomenon, which the authors call stimulus-triggered acquisition of pluripotency (STAP). In STAP, mouse somatic cells such as CD45.sup.+ haematopoietic cells are reprogrammed to pluripotency by transient exposure to low pH. Extensive analysis of the molecular features and developmental potential of STAP cells suggests that they represent a unique state of pluripotency -- and provide an alternative source of pluripotent cells to the use of transcription factors, as has become routine for induced pluripotent stem cell production., Author(s): Haruko Obokata [sup.1] [sup.2] [sup.3], Teruhiko Wakayama [sup.3] [sup.8], Yoshiki Sasai [sup.4], Koji Kojima [sup.1], Martin P. Vacanti [sup.1] [sup.5], Hitoshi Niwa [sup.6], Masayuki Yamato [sup.7], Charles A. Vacanti [...]

Introduction T follicular helper (Tfh) cells are a subset of specialized effector [CD4.sup.+] T cells mediating B cell selection, proliferation, and differentiation in germinal centers (GCs) (1). Aberrant activation of [...], T follicular helper (Tfh) cells are indispensable for the formation of germinal center (GC) reactions, whereas T follicular regulatory (Tfr) cells inhibit Tfh-mediated GC responses. Aberrant activation of Tfh cells contributes substantially to the pathogenesis of autoimmune diseases, such as systemic lupus erythematosus (SLE). Nonetheless, the molecular mechanisms mitigating excessive Tfh cell differentiation are not fully understood. Herein we demonstrate that the adenovirus E4 promoter-binding protein (E4BP4) mediates a feedback loop and acts as a transcriptional brake to inhibit Tfh cell differentiation. Furthermore, we show that such an immunological mechanism is compromised in patients with SLE. Establishing mice with either conditional knockout (cKO) or knockin (cKI) of the E4bp4 gene in T cells reveals that E4BP4 strongly inhibits Tfh cell differentiation. Mechanistically, E4BP4 regulates Bcl6 transcription by recruiting the repressive epigenetic modifiers HDAC1 and EZH2. E4BP4 phosphorylation site mutants have limited capability with regard to inhibiting Tfh cell differentiation. In SLE, we detected impaired phosphorylation of E4BP4, finding that this compromised transcription factor is positively correlated with disease activity. These findings unveiled molecular mechanisms by which E4BP4 restrains Tfh cell differentiation, whose compromised function is associated with uncontrolled autoimmune reactions in SLE.

Introduction Arsenic-resistance protein 2 [ARS2, also known as SRRT (Serrate RNA effector molecule)], is a stable component of the nuclear cap-binding complex and is required for the processing and transport [...], During a developmental period that extends postnatally in the mouse, proliferating multipotent retinal progenitor cells produce one of 7 major cell types (rod, cone, bipolar, horizontal, amacrine, ganglion, and Muller glial cells) as they exit the cell cycle in consecutive waves. Cell production in the retina is tightly regulated by intrinsic, extrinsic, spatial, and temporal cues, and is coupled to the timing of cell cycle exit. Arsenic-resistance protein 2 (ARS2, also known as SRRT) is a component of the nuclear cap-binding complex involved in RNA Polymerase II transcription, and is required for cell cycle progression. We show that postnatal retinal progenitor cells (RPCs) require ARS2 for proper progression through S phase, and ARS2 disruption leads to early exit from the cell cycle. Furthermore, we observe an increase in the proportion of cells expressing a rod photoreceptor marker, and a loss of Muller glia marker expression, indicating a role for ARS2 in regulating cell fate specification or differentiation. Knockdown of Flice Associated Huge protein (FLASH), which interacts with ARS2 and is required for cell cycle progression and 3'-end processing of replication-dependent histone transcripts, phenocopies ARS2 knockdown. These data implicate ARS2--FLASH-mediated histone mRNA processing in regulating RPC cell cycle kinetics and neuroglial cell fate specification during postnatal retinal development. Key words: Ars2, retina, retinal progenitor cells, cell cycle, photoreceptor cells, Muller cells. Durant une periode developpementale qui s'etend a la periode postnatale chez la souris, les cellules souches retiniennes multipotentes en proliferation donnent naissance a un des sept types principaux de cellules (batonnets, cones, cellules bipolaires, cellules horizontales, cellules amacrines, cellules ganglionnaires et cellules gliales de Muller) au fur et a mesure de leur sortie du cycle cellulaire par vagues consecutives. La production de cellules dans la retine est etroitement regulee par des signaux intrinseques, extrinseques, spatiaux et temporels et elle est couplee au moment du deroulement de la sortie du cycle cellulaire. ARS2 (aussi connue comme SRRT) est une composante du complexe nucleaire de liaison de la coiffe impliquee dans la transcription par l'ARN polymerase II, et elle est requise a la progression du cycle cellulaire. Les auteurs montrent que les cellules souches retiniennes postnatales requierent ARS2 afin de progresser adequatement a travers la phase S, et que le dereglement d'ARS2 donne lieu a une sortie hative du cycle cellulaire. De plus, ils observent une augmentation de la proportion de cellules qui expriment un marqueur des batonnets, et une perte d'expression d'un marqueur des cellules gliales de Muller, indiquant qu'ARS2 pourrait jouer un role dans la regulation de la specification de la destinee cellulaire ou la differenciation. Le knockdown de FLASH, qui interagit avec ARS2 et qui est necessaire a la progression du cycle cellulaire et a la maturation de l'extremite 3' des transcrits d'histones dependants de la replication, est une phenocopie du knockdown d'ARS2. Ces donnees impliquent la maturation de l'ARNm des histones mediee par ARS2--FLASH dans la regulation de la cinetique du cycle cellulaire des cellules souches retiniennes et dans la specification de la destinee cellulaire neurogliale durant le developpement retinien postnatal. [Traduit par la Redaction] Mots-cles : Ars2, retine, cellules souches retiniennes, cycle cellulaire, photorecepteurs, cellules de Muller.

Introduction T cell development in the thymus is a multistep process. Early thymic progenitor cells (TPCs) differentiate into T cell receptor-expressing (TCR-expressing) [CD4.sup.+][CD8.sup.+] double-positive (DP) thymocytes in the cortex and [...], Histone H3K27 demethylase JMJD3 plays a critical role in gene expression and T cell differentiation. However, the role and mechanisms of JMJD3 in T cell trafficking remain poorly understood. Here, we show that JMJD3 deficiency in [CD4.sup.+] T cells resulted in an accumulation of T cells in the thymus and reduction of T cell number in the secondary lymphoid organs. We identified PDLIM4 as a significantly downregulated target gene in JMJD3- deficient [CD4.sup.+] T cells by gene profiling and ChIP-Seq analyses. We further showed that PDLIM4 functioned as an adaptor protein to interact with sphingosine-1 phosphate receptor 1 (S1P1) and filamentous actin (F-actin), thus serving as a key regulator of T cell trafficking. Mechanistically, JMJD3 bound to the promoter and gene-body regions of the Pdlim4 gene and regulated its expression by interacting with zinc finger transcription factor KLF2. Our findings have identified Pdlim4 as a JMJD3 target gene that affects T cell trafficking by cooperating with S1P1 and have provided insights into the molecular mechanisms by which JMJD3 regulates genes involved in T cell trafficking.

Introduction The members of the annexin family of calcium- and phospholipid-binding proteins participate in different cellular processes (Gerke and Moss 2002; Rescher and Gerke 2004; Monastyrskaya et al. 2009). They [...], The members of the annexin family of calcium- and phospholipid-binding proteins participate in different cellular processes. Annexin A2 binds to S100A10, forming a functional heterotetrameric protein that has been involved in many cellular functions, such as exocytosis, endocytosis, cell junction formation, and actin cytoskeleton dynamics. Herein, we studied annexin A2 cellular movements and looked for its partners during epithelial cell differentiation. By using immunofluorescence, mass spectrometry (MS), and western blot analyses after S100A10 affinity column separation, we identified several annexin A2--S100A10 partner candidates. The association of putative annexin A2--S100A10 partner candidates obtained by MS after column affinity was validated by immunofluorescence and sucrose density gradient separation. The results show that three proteins are clearly associated with annexin A2: E-cadherin, actin, and caveolin 1. Overall, the data show that annexin A2 can associate with molecular complexes containing actin, caveolin 1, and flotillin 2 before epithelial differentiation and with complexes containing E-cadherin, actin, and caveolin 1, but not flotillin 2 after cell differentiation. The results indicate that actin, caveolin 1, and E-cadherin are the principal protein partners of annexin A2 in epithelial cells and that the serine phosphorylation of the N-terminal domain does not play an essential role during epithelial cell differentiation. Key words: actin, annexin A2, caveolin, epithelial cell differentiation, E-cadherin. Les membres de la famille des annexines, des protéines liant le calcium et les phospholipides, participent à différents processus cellulaires. L&apos;annexine A2 se lie à S100A10, formant une protéine hétérotétramérique fonctionnelle qui a été impliquée dans plusieurs fonctions cellulaires telles que l&apos;exocytose, l&apos;endocytose, la formation des jonctions cellulaires et la dynamique du cytosquelette. Les auteurs ont étudié ici les mouvements cellulaires de l&apos;annexine A2 et recherché ses partenaires durant la différenciation des cellules épithéliales. À l&apos;aide de l&apos;immunofluorescence, de la spectrométrie de masse et des analyses de transferts << western >> après séparation sur une colonne d&apos;affinité S100A10, ils ont identifié plusieurs candidats partenaires de l&apos;annexine A2--S100A10. L&apos;association de candidats partenaires présumés de l&apos;annexine A2--S100A10 obtenus par SM après la colonne d&apos;affinité a été validée par immunofluorescence et séparation sur un gradient de densité de sucrose. Les résultats montrent que trois protéines sont clairement associées à l&apos;annexine A2 : la E-cadhérine, l&apos;actine et la cavéoline 1. Globalement, les données montrent que l&apos;annexine A2 peut s&apos;associer à des complexes moléculaires comprenant l&apos;actine, la cavéoline et la flotilline 2 avant la différenciation épithéliale et à des complexes comprenant la E-cadhérine, l&apos;actine et la cavéoline 1, mais pas la flotilline 2, après la différenciation cellulaire. Les résultats indiquent que l&apos;actine, la cavéoline et la E-cadhérine constituent les principaux partenaires protéiques de l&apos;annexine A2 dans les cellules épithéliales et la phosphorylation des sérines du domaine N-terminal ne joue pas un rôle essentiel durant la différenciation des cellules épithéliales. [Traduit par la Rédaction] Mots-clés: actine, annexine A2, cavéoline, différenciation des cellules épithéliales, E-cadhérine.

Author(s): Yu Shi [sup.1] , Weina Gao [sup.2] [sup.3] , Nikki K. Lytle [sup.4] [sup.5] , Peiwu Huang [sup.2] [sup.3] [sup.6] , Xiao Yuan [sup.2] [sup.3] , Amanda M. Dann [...], Pancreatic ductal adenocarcinoma (PDAC) has a dismal prognosis largely owing to inefficient diagnosis and tenacious drug resistance. Activation of pancreatic stellate cells (PSCs) and consequent development of dense stroma are prominent features accounting for this aggressive biology.sup.1,2. The reciprocal interplay between PSCs and pancreatic cancer cells (PCCs) not only enhances tumour progression and metastasis but also sustains their own activation, facilitating a vicious cycle to exacerbate tumorigenesis and drug resistance.sup.3-7. Furthermore, PSC activation occurs very early during PDAC tumorigenesis.sup.8-10, and activated PSCs comprise a substantial fraction of the tumour mass, providing a rich source of readily detectable factors. Therefore, we hypothesized that the communication between PSCs and PCCs could be an exploitable target to develop effective strategies for PDAC therapy and diagnosis. Here, starting with a systematic proteomic investigation of secreted disease mediators and underlying molecular mechanisms, we reveal that leukaemia inhibitory factor (LIF) is a key paracrine factor from activated PSCs acting on cancer cells. Both pharmacologic LIF blockade and genetic Lifr deletion markedly slow tumour progression and augment the efficacy of chemotherapy to prolong survival of PDAC mouse models, mainly by modulating cancer cell differentiation and epithelial-mesenchymal transition status. Moreover, in both mouse models and human PDAC, aberrant production of LIF in the pancreas is restricted to pathological conditions and correlates with PDAC pathogenesis, and changes in the levels of circulating LIF correlate well with tumour response to therapy. Collectively, these findings reveal a function of LIF in PDAC tumorigenesis, and suggest its translational potential as an attractive therapeutic target and circulating marker. Our studies underscore how a better understanding of cell-cell communication within the tumour microenvironment can suggest novel strategies for cancer therapy. A systematic proteomic investigation of disease mediators secreted by pancreatic stellate cells identifies leukaemia inhibitory factor (LIF) as a key factor that acts on cancer cells, promoting tumour progression and chemoresistance.

Author(s): Judith Feucht [sup.1] , Jie Sun [sup.1] [sup.2] , Justin Eyquem [sup.1] , Yu-Jui Ho [sup.3] , Zeguo Zhao [sup.1] , Josef Leibold [sup.3] , Anton Dobrin [sup.1] , [...], Chimeric antigen receptors (CARs) are synthetic receptors that target and reprogram T cells to acquire augmented antitumor properties.sup.1. CD19-specific CARs that comprise CD28 and CD3[zeta] signaling motifs.sup.2 have induced remarkable responses in patients with refractory leukemia.sup.3-5 and lymphoma.sup.6 and were recently approved by the US Food and Drug Administration.sup.7. These CARs program highly performing effector functions that mediate potent tumor elimination.sup.4,8 despite the limited persistence they confer on T cells.sup.3-6,8. Extending their functional persistence without compromising their potency should improve current CAR therapies. Strong T cell activation drives exhaustion.sup.9,10, which may be accentuated by the redundancy of CD28 and CD3[zeta] signaling.sup.11,12 as well as the spatiotemporal constraints imparted by the structure of second-generation CARs.sup.2. Thus, we hypothesized that calibrating the activation potential of CD28-based CARs would differentially reprogram T cell function and differentiation. Here, we show that CARs encoding a single immunoreceptor tyrosine-based activation motif direct T cells to different fates by balancing effector and memory programs, thereby yielding CAR designs with enhanced therapeutic profiles. A novel chimeric antigen receptor (CAR) T cell design improves therapeutic efficacy by balancing effector and memory profiles.

Author(s): Weiqi Zhang [sup.1] [sup.2] [sup.3] [sup.4] , Haifeng Wan [sup.2] [sup.3] [sup.4] , Guihai Feng [sup.2] [sup.3] [sup.4] , Jing Qu [sup.2] [sup.3] [sup.4] , Jiaqiang Wang [sup.2] [sup.3] [...], SIRT6 acts as a longevity protein in rodents.sup.1,2. However, its biological function in primates remains largely unknown. Here we generate a SIRT6-null cynomolgus monkey (Macaca fascicularis) model using a CRISPR-Cas9-based approach. SIRT6-deficient monkeys die hours after birth and exhibit severe prenatal developmental retardation. SIRT6 loss delays neuronal differentiation by transcriptionally activating the long non-coding RNA H19 (a developmental repressor), and we were able to recapitulate this process in a human neural progenitor cell differentiation system. SIRT6 deficiency results in histone hyperacetylation at the imprinting control region of H19, CTCF recruitment and upregulation of H19. Our results suggest that SIRT6 is involved in regulating development in non-human primates, and may provide mechanistic insight into human perinatal lethality syndrome.A cynomolgus monkey model deficient in SIRT6 protein exhibits severe retardation in prenatal development, in which neuronal differentiation is delayed by activation of the H19 long non-coding RNA.

Genetic mutations of oncogenes lead to altered protein functions, resulting in increased cellular proliferation, differentiation blockade, and, ultimately, neoplastic transformation. With progression, tumors accumulate mutations and mutational load is generally [...], Effector T cell responses directed toward cancer neoantigens mediate tumor regression following checkpoint blockade or adoptive T cell immunotherapy, but are generally "private", thus requiring considerable effort for their identification. In this issue of the JCI, Malekzadeh et al. show that a fraction of patients with epithelial cancers mount antigen-specific T cell responses to "hot spot" p53 mutations that in some cases are shared among patients. This work suggests that other genes frequently mutated in human cancer can be immunogenic, thus offering a rapid way to screen for cancer neoantigens that can be targeted by subsequent T cell-based therapies.

Author(s): Stephanie J. Ellis [sup.1] , Nicholas C. Gomez [sup.1] , John Levorse [sup.1] , Aaron F. Mertz [sup.1] , Yejing Ge [sup.1] , Elaine Fuchs [sup.1] Author Affiliations: (1) [...], Cell competition--the sensing and elimination of less fit &apos;loser&apos; cells by neighbouring &apos;winner&apos; cells--was first described in Drosophila. Although cell competition has been proposed as a selection mechanism to optimize tissue and organ development, its evolutionary generality remains unclear. Here, by using live imaging, lineage tracing, single-cell transcriptomics and genetics, we identify two cell competition mechanisms that sequentially shape and maintain the architecture of stratified tissue during skin development in mice. In the single-layered epithelium of the early embryonic epidermis, winner progenitors kill and subsequently clear neighbouring loser cells by engulfment. Later, as the tissue begins to stratify, the basal layer instead expels losers through upward flux of differentiating progeny. This cell competition switch is physiologically relevant: when it is perturbed, so too is barrier formation. Our findings show that cell competition is a selective force that optimizes vertebrate tissue function, and illuminate how a tissue dynamically adjusts cell competition strategies to preserve fitness as its architectural complexity increases during morphogenesis. Cell competition in the developing mouse epithelium involves apoptosis and engulfment when the epithelium has only one layer, but switches to involve asymmetric cell division and differentiation of &apos;loser&apos; cells as the epithelium becomes stratified.

Author(s): Federico Gaiti [sup.1] [sup.2] , Ronan Chaligne [sup.1] [sup.2] , Hongcang Gu [sup.3] , Ryan M. Brand [sup.1] [sup.2] , Steven Kothen-Hill [sup.1] [sup.2] , Rafael C. Schulman [sup.1] [...], Genetic and epigenetic intra-tumoral heterogeneity cooperate to shape the evolutionary course of cancer.sup.1. Chronic lymphocytic leukaemia (CLL) is a highly informative model for cancer evolution as it undergoes substantial genetic diversification and evolution after therapy.sup.2,3. The CLL epigenome is also an important disease-defining feature.sup.4,5, and growing populations of cells in CLL diversify by stochastic changes in DNA methylation known as epimutations.sup.6. However, previous studies using bulk sequencing methods to analyse the patterns of DNA methylation were unable to determine whether epimutations affect CLL populations homogeneously. Here, to measure the epimutation rate at single-cell resolution, we applied multiplexed single-cell reduced-representation bisulfite sequencing to B cells from healthy donors and patients with CLL. We observed that the common clonal origin of CLL results in a consistently increased epimutation rate, with low variability in the cell-to-cell epimutation rate. By contrast, variable epimutation rates across healthy B cells reflect diverse evolutionary ages across the trajectory of B cell differentiation, consistent with epimutations serving as a molecular clock. Heritable epimutation information allowed us to reconstruct lineages at high-resolution with single-cell data, and to apply this directly to patient samples. The CLL lineage tree shape revealed earlier branching and longer branch lengths than in normal B cells, reflecting rapid drift after the initial malignant transformation and a greater proliferative history. Integration of single-cell bisulfite sequencing analysis with single-cell transcriptomes and genotyping confirmed that genetic subclones mapped to distinct clades, as inferred solely on the basis of epimutation information. Finally, to examine potential lineage biases during therapy, we profiled serial samples during ibrutinib-associated lymphocytosis, and identified clades of cells that were preferentially expelled from the lymph node after treatment, marked by distinct transcriptional profiles. The single-cell integration of genetic, epigenetic and transcriptional information thus charts the lineage history of CLL and its evolution with therapy. A single-cell approach is used to follow the heritable stochastic changes to DNA methylation that occur in primary chronic lymphocytic leukaemia and healthy B cells, allowing the tracing of cell lineage histories and evolution during treatment with ibrutinib.

Author(s): Zixu Liu [sup.1] , Katrina B. Mar [sup.1] , Natasha W. Hanners [sup.2] , Sofya S. Perelman [sup.1] , Mohammed Kanchwala [sup.3] , Chao Xing [sup.3] [sup.4] [sup.5] , [...], The non-canonical NF-[kappa]B signalling cascade is essential for lymphoid organogenesis, B cell maturation, osteoclast differentiation, and inflammation in mammals.sup.1,2; dysfunction of this system is associated with human diseases, including immunological disorders and cancer.sup.3-6. Although expression of NF-[kappa]B-inducing kinase (NIK, also known as MAP3K14) is the rate-limiting step in non-canonical NF-[kappa]B pathway activation.sup.2,7, the mechanisms by which transcriptional responses are regulated remain largely unknown. Here we show that the sine oculis homeobox (SIX) homologue family transcription factors SIX1 and SIX2 are integral components of the non-canonical NF-[kappa]B signalling cascade. The developmentally silenced SIX proteins are reactivated in differentiated macrophages by NIK-mediated suppression of the ubiquitin proteasome pathway. Consequently, SIX1 and SIX2 target a subset of inflammatory gene promoters and directly inhibit the trans-activation function of the transcription factors RELA and RELB in a negative feedback circuit. In support of a physiologically pivotal role for SIX proteins in host immunity, a human SIX1 transgene suppressed inflammation and promoted the recovery of mice from endotoxic shock. In addition, SIX1 and SIX2 protected RAS/P53-driven non-small-cell lung carcinomas from inflammatory cell death induced by SMAC-mimetic chemotherapeutic agents (small-molecule activators of the non-canonical NF-[kappa]B pathway). Our findings identify a NIK-SIX signalling axis that fine-tunes inflammatory gene expression programs under both physiological and pathological conditions. Non-canonical NF-[kappa]B is regulated by a negative feedback loop involving NIK-mediated activation of SIX1 and SIX2.

Kidney fibrosis is associated with a reduction in the functional renal parenchyma, leading to compromised kidney functions and eventual organ death (1). There is no effective treatment for renal fibrosis [...], Kidney fibrosis is marked by an epithelial-to-mesenchymal transition (EMT) of tubular epithelial cells (TECs). Here we find that, during renal fibrosis, TECs acquire a partial EMT program during which they remain associated with their basement membrane and express markers of both epithelial and mesenchymal cells. The functional consequence of the EMT program during fibrotic injury is an arrest in the G2 phase of the cell cycle and lower expression of several solute and solvent transporters in TECs. We also found that transgenic expression of either Twist1 (encoding twist family bHLH transcription factor 1, known as Twist) or Snail (encoding snail family zinc finger 1, known as Snail) expression is sufficient to promote prolonged TGF-β1-induced G2 arrest of TECs, limiting the cells' potential for repair and regeneration. In mouse models of experimentally induced renal fibrosis, conditional deletion of Twist1 or Snail in proximal TECs resulted in inhibition of the EMT program and the maintenance of TEC integrity, while also restoring cell proliferation, dedifferentiation-associated repair and regeneration of the kidney parenchyma and attenuating interstitial fibrosis. Thus, inhibition of the EMT program in TECs during chronic renal injury represents a potential anti-fibrosis therapy.

To establish whether post-mitotic ciliated cells send a conventional feedback signal to regulate the replication of their parent stem and progenitor cells, we genetically ablated ciliated cells using FOXJ1-creER; LSL-DTA [...], Stem cells integrate inputs from multiple sources. Stem cell niches provide signals that promote stem cell maintenance (1, 2), while differentiated daughter cells are known to provide feedback signals to regulate stem cell replication and differentiation (3-6). Recently, stem cells have been shown to regulate themselves using an autocrine mechanism (7). The existence of a 'stem cell niche' was first postulated by Schofield in 1978 to define local environments necessary for the maintenance of haematopoietic stem cells (1). Since then, an increasing body of work has focused on defining stem cell niches (1-6). Yet little is known about how progenitor cell and differentiated cell numbers and proportions are maintained. In the airway epithelium, basal cells function as stem/progenitor cells that can both self-renew and produce differentiated secretory cells and ciliated cells (8, 9). Secretory cells also act as transit-amplifying cells that eventually differentiate into post-mitotic ciliated cells (9, 10). Here we describe a mode of cell regulation in which adult mammalian stem/progenitor cells relay a forward signal to their own progeny. Surprisingly, this forward signal is shown to be necessary for daughter cell maintenance. Using a combination of cell ablation, lineage tracing and signalling pathway modulation, we show that airway basal stem/progenitor cells continuously supply a Notch ligand to their daughter secretory cells. Without these forward signals, the secretory progenitor cell pool fails to be maintained and secretory cells execute a terminal differentiation program and convert into ciliated cells. Thus, a parent stem/progenitor cell can serve as a functional daughter cell niche.

We have previously shown that four PLT transcription factors with graded distribution (PLT1, PLT2, PLT3 and BBM (also known as PLT4)) are necessary for stem cell maintenance and cell division [...], During plant growth, dividing cells in meristems must coordinate transitions from division to expansion and differentiation, thus generating three distinct developmental zones: the meristem, elongation zone and differentiation zone (1). Simultaneously, plants display tropisms, rapid adjustments of their direction of growth to adapt to environmental conditions. It is unclear how stable zonation is maintained during transient adjustments in growth direction. In Arabidopsis roots, many aspects of zonation are controlled by the phytohormone auxin and auxin-induced PLETHORA (PLT) transcription factors, both of which display a graded distribution with a maximum near the root tip (2-12). In addition, auxin is also pivotal for tropic responses (13,14). Here, using an iterative experimental and computational approach, we show how an interplay between auxin and PLTs controls zonation and gravitropism. We find that the PLT gradient is not a direct, proportionate readout of the auxin gradient. Rather, prolonged high auxin levels generate a narrow PLT transcription domain from which a gradient of PLT protein is subsequently generated through slow growth dilution and cell-to-cell movement. The resulting PLT levels define the location of developmental zones. In addition to slowly promoting PLT transcription, auxin also rapidly influences division, expansion and differentiation rates. We demonstrate how this specific regulatory design in which auxin cooperates with PLTs through different mechanisms and on different timescales enables both the fast tropic environmental responses and stable zonation dynamics necessary for coordinated cell differentiation.

Author(s): Peter Savas [sup.1] [sup.2] , Balaji Virassamy [sup.1] , Chengzhong Ye [sup.3] [sup.4] [sup.5] , Agus Salim [sup.3] [sup.6] , Christopher P. Mintoff [sup.1] , Franco Caramia [sup.1] , [...], The quantity of tumor-infiltrating lymphocytes (TILs) in breast cancer (BC) is a robust prognostic factor for improved patient survival, particularly in triple-negative and HER2-overexpressing BC subtypes.sup.1. Although T cells are the predominant TIL population.sup.2, the relationship between quantitative and qualitative differences in T cell subpopulations and patient prognosis remains unknown. We performed single-cell RNA sequencing (scRNA-seq) of 6,311 T cells isolated from human BCs and show that significant heterogeneity exists in the infiltrating T cell population. We demonstrate that BCs with a high number of TILs contained CD8.sup.+ T cells with features of tissue-resident memory T (T.sub.RM) cell differentiation and that these CD8.sup.+ T.sub.RM cells expressed high levels of immune checkpoint molecules and effector proteins. A CD8.sup.+ T.sub.RM gene signature developed from the scRNA-seq data was significantly associated with improved patient survival in early-stage triple-negative breast cancer (TNBC) and provided better prognostication than CD8 expression alone. Our data suggest that CD8.sup.+ T.sub.RM cells contribute to BC immunosurveillance and are the key targets of modulation by immune checkpoint inhibition. Further understanding of the development, maintenance and regulation of T.sub.RM cells will be crucial for successful immunotherapeutic development in BC. Extensive, high-dimensional characterization of T cells in breast cancer reveals activated T.sub.RM population and a gene signature associated with improved prognosis.

Author(s): Joseph A. Fraietta [sup.1] [sup.2] [sup.3] [sup.4] , Christopher L. Nobles [sup.5] , Morgan A. Sammons [sup.6] [sup.10] , Stefan Lundh [sup.1] [sup.2] , Shannon A. Carty [sup.2] [sup.11] [...], Cancer immunotherapy based on genetically redirecting T cells has been used successfully to treat B cell malignancies.sup.1-3. In this strategy, the T cell genome is modified by integration of viral vectors or transposons encoding chimaeric antigen receptors (CARs) that direct tumour cell killing. However, this approach is often limited by the extent of expansion and persistence of CAR T cells.sup.4,5. Here we report mechanistic insights from studies of a patient with chronic lymphocytic leukaemia treated with CAR T cells targeting the CD19 protein. Following infusion of CAR T cells, anti-tumour activity was evident in the peripheral blood, lymph nodes and bone marrow; this activity was accompanied by complete remission. Unexpectedly, at the peak of the response, 94% of CAR T cells originated from a single clone in which lentiviral vector-mediated insertion of the CAR transgene disrupted the methylcytosine dioxygenase TET2 gene. Further analysis revealed a hypomorphic mutation in this patient's second TET2 allele. TET2-disrupted CAR T cells exhibited an epigenetic profile consistent with altered T cell differentiation and, at the peak of expansion, displayed a central memory phenotype. Experimental knockdown of TET2 recapitulated the potency-enhancing effect of TET2 dysfunction in this patient's CAR T cells. These findings suggest that the progeny of a single CAR T cell induced leukaemia remission and that TET2 modification may be useful for improving immunotherapies.Genetically engineered T cells that induced remission in a patient with chronic lymphocytic leukaemia were found to have disruption of the TET2 gene, which caused T cell changes that potentiated their anti-tumour effects.

We recently discovered an intriguing phenomenon of cellular fate conversion: somatic cells regain pluripotency after experiencing sublethal stimuli such as a low-pH exposure (1). When splenic [CD45.sup.+] lymphocytes are exposed [...], We recently discovered an unexpected phenomenon of somatic cell reprogramming into pluripotent cells by exposure to sublethal stimuli, which we call stimulus-triggered acquisition of pluripotency (STAP) (1). This reprogramming does not require nuclear transfer (2,3) or genetic manipulation (4). Here we report that reprogrammed STAP cells, unlike embryonic stem (ES) cells, can contribute to both embryonic and placental tissues, as seen in a blastocyst injection assay. Mouse STAP cells lose the ability to contribute to the placenta as well as trophoblast marker expression on converting into ES-like stem cells by treatment with adrenocorticotropic hormone (ACTH) and leukaemia inhibitory factor (LIF). In contrast, when cultured with Fgf4, STAP cells give rise to proliferative stem cells with enhanced trophoblastic characteristics. Notably, unlike conventional trophoblast stem cells, the Fgf4-induced stem cells from STAP cells contribute to both embryonic and placental tissues in vivo and transform into ES-like cells when cultured with LIF-containing medium. Taken together, the developmental potential of STAP cells, shown by chimaera formation and in vitro cell conversion, indicates that they represent a unique state of pluripotency.

Humans generate 10 (11) erythrocytes every day, a process regulated by multiple hormones affecting several types of progenitors. Apoptosis, proliferation and terminal differentiation of CFU-E erythroid progenitors are mainly controlled [...], Stem cells and progenitors in many lineages undergo self-renewing divisions, but the extracellular and intracellular proteins that regulate this process are largely unknown. Glucocorticoids stimulate red blood cell formation by promoting self-renewal of early burst-forming unit-erythroid (BFU-E) progenitors (1-4). Here we show that the RNA-binding protein ZFP36L2 is a transcriptional target of the glucocorticoid receptor (GR) in BFU-Es and is required for BFU-E self-renewal. ZFP36L2 is normally downregulated during erythroid differentiation from the BFU-E stage, but its expression is maintained by all tested GR agonists that stimulate BFU-E self-renewal, and the GR binds to several potential enhancer regions of ZFP36L2. Knockdown of ZFP36L2 in cultured BFU-E cells did not affect the rate of cell division but disrupted glucocorticoid-induced BFU-E self-renewal, and knockdown of ZFP36L2 in transplanted erythroid progenitors prevented expansion of erythroid lineage progenitors normally seen following induction of anaemia by phenylhydrazine treatment. ZFP36L2 preferentially binds to messenger RNAs that are induced or maintained at high expression levels during terminal erythroid differentiation and negatively regulates their expression levels. ZFP36L2 therefore functions as part of a molecular switch promoting BFU-E self-renewal and a subsequent increase in the total numbers of colony-forming unit-erythroid (CFU-E) progenitors and erythroid cells that are generated.