Cell Fate Plasticity Research Articles

De-Differentiation of Corneal Epithelial Cells Into Functional Limbal Epithelial Stem Cells After the Ablation of Innate Stem Cells.

Regeneration after tissue injury is often associated with cell fate plasticity, which restores damaged or lost cells. Here, we examined the de-differentiation of corneal epithelial cells (CECs) into functional limbal epithelial stem cells (LESCs) after the ablation of innate stem cells. The regeneration of LESCs after the ablation of innate LESCs was identified by a set of markers: ApoE+/Cx43low/CK12-. CK14-CreERT2 or Slc1a3-CreERT mice were crossed with reporter mice to trace the fate of CECs. YAP-TEAD inhibitor verteporfin (VTP) and LATS inhibitor TRULI were used to examine the role of Hippo/YAP pathway in the de-differentiation of CECs. LESCs-ablation cornea showed to be functionally normal, including the maintenance of corneal transparency, prevention of conjunctivalization, and wound healing rate equivalent to that of normal cornea. ApoE+/Cx43low/CK12- LESCs regenerated at the limbus at 6 days after the ablation of innate stem cells, and maintained for at least 6 months. Corneal epithelial lineage tracing showed that CECs migrated back to the limbus after the ablation of innate stem cells, and de-differentiated into active and quiescent LESCs (aLESCs and qLESCs), which participated in corneal epithelial homeostasis and wound healing, respectively, like their innate counterparts. However, when the limbus niche was destroyed by NaOH (1 M, 5 seconds), CECs that occupied the limbus could not de-differentiate into ApoE+/Cx43low/CK12- LESCs and cornea developed into conjunctivalization. In addition, the protein level and activity of YAP increased at the early stage (1-2 days) after the ablation of limbal epithelium, and decreased when the de-differentiation occurred. The YAP-TEAD inhibitor VTP promoted the de-differentiation, whereas LATS inhibitor TRULI inhibited the de-differentiation of CECs. However, the persistent activation of YAP prevented the de-differentiation of CECs after an additional NaOH burn to the limbal stroma, and VTP could not rescue the capacity of CECs to de-differentiate into LESCs. These results reveal the de-differentiation of CECs into functional LESCs after the ablation of innate stem cells, and suggest potential role of Hippo/YAP pathway in the de-differentiation of CECs in vivo.

Eph-ephrin signaling couples endothelial cell sorting and arterial specification

Cell segregation allows the compartmentalization of cells with similar fates during morphogenesis, which can be enhanced by cell fate plasticity in response to local molecular and biomechanical cues. Endothelial tip cells in the growing retina, which lead vessel sprouts, give rise to arterial endothelial cells and thereby mediate arterial growth. Here, we have combined cell type-specific and inducible mouse genetics, flow experiments in vitro, single-cell RNA sequencing and biochemistry to show that the balance between ephrin-B2 and its receptor EphB4 is critical for arterial specification, cell sorting and arteriovenous patterning. At the molecular level, elevated ephrin-B2 function after loss of EphB4 enhances signaling responses by the Notch pathway, VEGF and the transcription factor Dach1, which is influenced by endothelial shear stress. Our findings reveal how Eph-ephrin interactions integrate cell segregation and arteriovenous specification in the vasculature, which has potential relevance for human vascular malformations caused by EPHB4 mutations.

Abstract 3270: SH3GLB1 is involved with hydrogen peroxide/akt signaling in glioblastoma multiforme cells

Abstract Glioblastoma (GBM), a malignant brain tumor, has poor survival outcomes due to recurrence or drug resistance. In our previous studies, it was found that superoxide dismutase 2 (SOD2) was associated with temozolomide (TMZ) resistance. Moreover, SOD2 can convert O2 •− to H2O2, and H2O2 redox signaling is vital for physiological reactions and promotes tumor progression. We used cell lines, patient primary cells and pharmacological inhibitors/activators to confirm the significance of H2O2 signaling. We used the cultured cells to verify roles of H2O2 signaling in the plasticity of cell fate and did animal experiments to identify the optimal treatment strategy. In the present study, SOD2 was found to be the upstream mediator, and combining SOD inhibitor and TMZ, the cells showed reduced SH3GLB1 and autophagy levels. We then found that SH3GLB1 was regulated by H2O2 through AKT signaling using experiments of regulating redox homeostasis. The change of intracellular H2O2 levels in mitochondria following the treatments is similar to that in cytosol, however, the cell fate of parental or resistant cells was quite diverse. Notably, CCCP, a mitochondrial membrane potential disrupter, increased endogenous H2O2 production, and TMZ-increased SH3GLB1 levels can be reversed by HgCl2, reflecting the aquaporin function. In animal models, a combination of TMZ and 4-hydroxynonenal reduced growth of the SH3GLB1-knockdown resistant cells, along with suppressed autophagy levels. Our results characterizing the mechanism of H2O2-SH3GLB1 axis signaling in acquired TMZ resistance of GBM cells will provide new therapeutic strategies against the disease in the future. Citation Format: Chia-Hung Chien, Kwang-Yu Chang. SH3GLB1 is involved with hydrogen peroxide/akt signaling in glioblastoma multiforme cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3270.

Abstract 17130: Metabolic and Epigenetic Coupled Regulation in Angiogenic Transdifferentiation and Vascular Recovery

Recent studies from our group and others have characterized the phenomenon of "angiogenic transdifferentiation” from fibroblasts to endothelial cells as a novel endogenous mechanism for microvasculature expansion and perfusion restoration in murine models of hindlimb ischemia and myocardial infarction. Metabolic regulation of epigenetic reprogramming has emerged as a key regulatory process controlling cell fate transition. To study the comprehensive role of metabolism in this healing process, we conducted liquid chromatography-mass spectrometry (LC-MS) based untargeted and targeted metabolomics during transdifferentiation. Our analysis revealed the upregulation of key metabolites, including UDP-GlcNAc (substrate for post-translational modification O-GlcNAcylation) and acetyl-CoA (substrate for histone acetylation). With our in vitro transdifferentiation protocol, we found that pharmacologic or genetic inhibition of OGT (O-GlcNAc transferase, the enzyme responsible for protein O-GlcNAcylation) or ACL (ATP citrate lyase, the enzyme converting citrate to acetyl-CoA) in human BJ fibroblasts impairs transdifferentiation. Employing a preclinical peripheral arterial disease (PAD) hindlimb ischemia model with fibroblast lineage tracing mice, we discovered that O-GlcNAcylation and ACL were accumulated in fibroblast lineage cells in ischemic limbs, and pharmacologic inhibition of OGT or ACL diminished the transdifferentiation cell population in the ischemic tissue. Furthermore, we identified fibroblast-specific OGT or ACL KO impaired vascular recovery post femoral artery ligation in the mice. Finally, we showed that metabolism-dependent post-translational modifications on histones (acetylation of H3K9, H3K27, H3K14) or histone chaperons (O-GlcNAcylation of HIRA, a histone H3.3 chaperone) mechanistically enhanced chromatin openness, thereby facilitating cell fate plasticity and vascular regeneration. In conclusion, our studies demonstrate the significant regulatory potential of metabolic factors in angiogenic transdifferentiation which presents an innovative avenue to enhance vascular recovery in ischemic vascular diseases such as PAD.

Thymic epithelial cell fate and potency in early organogenesis assessed by single cell transcriptional and functional analysis.

During development, cortical (c) and medullary (m) thymic epithelial cells (TEC) arise from the third pharyngeal pouch endoderm. Current models suggest that within the thymic primordium most TEC exist in a bipotent/common thymic epithelial progenitor cell (TEPC) state able to generate both cTEC and mTEC, at least until embryonic day 12.5 (E12.5) in the mouse. This view, however, is challenged by recent transcriptomics and genetic evidence. We therefore set out to investigate the fate and potency of TEC in the early thymus. Here using single cell (sc) RNAseq we identify a candidate mTEC progenitor population at E12.5, consistent with recent reports. Via lineage-tracing we demonstrate this population as mTEC fate-restricted, validating our bioinformatics prediction. Using potency analyses we also establish that most E11.5 and E12.5 progenitor TEC are cTEC-fated. Finally we show that overnight culture causes most if not all E12.5 cTEC-fated TEPC to acquire functional bipotency, and provide a likely molecular mechanism for this changed differentiation potential. Collectively, our data overturn the widely held view that a common TEPC predominates in the E12.5 thymus, showing instead that sublineage-primed progenitors are present from the earliest stages of thymus organogenesis but that these early fetal TEPC exhibit cell-fate plasticity in response to extrinsic factors. Our data provide a significant advance in the understanding of fetal thymic epithelial development and thus have implications for thymus-related clinical research, in particular research focussed on generating TEC from pluripotent stem cells.

Perfect duet: Dual recombinases improve genetic resolution.

As a powerful genetic tool, site-specific recombinases (SSRs) have been widely used in genomic manipulation to elucidate cell fate plasticity in vivo, advancing research in stem cell and regeneration medicine. However, the low resolution of conventional single-recombinase-mediated lineage tracing strategies, which rely heavily on the specificity of one marker gene, has led to controversial conclusions in many scientific questions. Therefore, different SSRs systems are combined to improve the accuracy of lineage tracing. Here we review the recent advances in dual-recombinase-mediated genetic approaches, including the development of novel genetic recombination technologies and their applications in cell differentiation, proliferation, and genetic manipulation. In comparison with the single-recombinase system, we also discuss the advantages of dual-genetic strategies in solving scientific issues as well as their technical limitations.

Increasing cell culture density during a developmental window prevents fated rod precursors derailment toward hybrid rod-glia cells

In proliferating multipotent retinal progenitors, transcription factors dynamics set the fate of postmitotic daughter cells, but postmitotic cell fate plasticity driven by extrinsic factors remains controversial. Transcriptome analysis reveals the concurrent expression by postmitotic rod precursors of genes critical for the Müller glia cell fate, which are rarely generated from terminally-dividing progenitors as a pair with rod precursors. By combining gene expression and functional characterisation in single cultured rod precursors, we identified a time-restricted window where increasing cell culture density switches off the expression of genes critical for Müller glial cells. Intriguingly, rod precursors in low cell culture density maintain the expression of genes of rod and glial cell fate and develop a mixed rod/Muller glial cells electrophysiological fingerprint, revealing rods derailment toward a hybrid rod-glial phenotype. The notion of cell culture density as an extrinsic factor critical for preventing rod-fated cells diversion toward a hybrid cell state may explain the occurrence of hybrid rod/MG cells in the adult retina and provide a strategy to improve engraftment yield in regenerative approaches to retinal degenerative disease by stabilising the fate of grafted rod precursors.

Use of a dual genetic system to decipher exocrine cell fate conversions in the adult pancreas

Unraveling cell fate plasticity during tissue homeostasis and repair can reveal actionable insights for stem cell biology and regenerative medicine. In the pancreas, it remains controversial whether lineage transdifferentiation among the exocrine cells occur under pathophysiological conditions. Here, to address this question, we used a dual recombinase-mediated genetic system that enables simultaneous tracing of pancreatic acinar and ductal cells using two distinct genetic reporters, avoiding the “ectopic” labeling by Cre-loxP recombination system. We found that acinar-to-ductal transdifferentiation occurs after pancreatic duct ligation or during caerulein-induced pancreatitis, but not during homeostasis or after partial pancreatectomy. On the other hand, pancreatic ductal cells contribute to new acinar cells after significant acinar cell loss. By genetic tracing of cell proliferation, we also quantify the cell proliferation dynamics and deduce the turnover rate of pancreatic exocrine lineages during homeostasis. Together, these results suggest that the lineage transdifferentiation happens between acinar cells and ductal cells in the pancreatic exocrine glands under specific conditions.

Towards Defining Principles of Cell Fate Plasticity

Towards Defining Principles of Cell Fate Plasticity

Monitoring of cell-cell communication and contact history in mammals.

Monitoring of cell-cell communication in multicellular organisms is fundamental to understanding diverse biological processes such as embryogenesis and tumorigenesis. To track cell-cell contacts in vivo, we developed an intercellular genetic technology to monitor cell-cell contact and to trace cell contact histories by permanently marking contacts between cells. In mice, we engineered an artificial Notch ligand into one cell (the sender cell) and an artificial receptor into another cell (the receiver cell). Contact between the sender and receiver cells triggered a synthetic Notch signaling that activated downstream transcriptional programs in the receiver cell, thereby transiently or permanently labeling it. In vivo cell-cell contact was observed during development, tissue homeostasis, and tumor growth. This technology may be useful for studying dynamic in vivo cell-cell contacts and cell fate plasticity.

Early developmental plasticity enables the induction of an intermediate extraembryonic cell state

Two fundamental elements of pre-implantation embryogenesis are cells' intrinsic self-organization program and their developmental plasticity, which allows embryos to compensate for alterations in cell position and number; yet, these elements are still poorly understood. To be able to decipher these features, we established culture conditions that enable the two fates of blastocysts' extraembryonic lineages-the primitive endoderm and the trophectoderm-to coexist. This plasticity emerges following the mechanisms of the first lineage segregation in the mouse embryo, and it manifests as an extended potential for extraembryonic chimerism during the pre-implantation embryogenesis. Moreover, this shared state enables robust assembly into higher-order blastocyst-like structures, thus combining both the cell fate plasticity and self-organization features of the early extraembryonic lineages.

A guiding role of the Arabidopsis circadian clock in cell differentiation revealed by time-series single-cell RNA sequencing.

Circadian rhythms and progression of cell differentiation are closely coupled in multicellular organisms. However, whether establishment of circadian rhythms regulates cell differentiation or vice versa has not been elucidated due to technical limitations. Here, we exploit high cell fate plasticity of plant cells to perform single-cell RNA sequencing during the entire process of cell differentiation. By analyzing reconstructed actual time series of the differentiation processes at single-cell resolution using a method we developed (PeakMatch), we find that the expression profile of clock genes is changed prior to cell differentiation, including induction of the clock gene LUX ARRYTHMO (LUX). ChIP sequencing analysis reveals that LUX induction in early differentiating cells directly targets genes involved in cell-cycle progression to regulate cell differentiation. Taken together, these results not only reveal a guiding role of the plant circadian clock in cell differentiation but also provide an approach for time-series analysis at single-cell resolution.

Inference of cell state transitions and cell fate plasticity from single-cell with MARGARET.

Despite recent advances in inferring cellular dynamics using single-cell RNA-seq data, existing trajectory inference (TI) methods face difficulty in accurately reconstructing the cell-state manifold and cell-fate plasticity for complex topologies. Here, we present MARGARET (https://github.com/Zafar-Lab/Margaret) for inferring single-cell trajectory and fate mapping for diverse dynamic cellular processes. MARGARET reconstructs complex trajectory topologies using a deep unsupervised metric learning and a graph-partitioning approach based on a novel connectivity measure, automatically detects terminal cell states, and generalizes the quantification of fate plasticity for complex topologies. On a diverse benchmark consisting of synthetic and real datasets, MARGARET outperformed state-of-the-art methods in recovering global topology and cell pseudotime ordering. For human hematopoiesis, MARGARET accurately identified all major lineages and associated gene expression trends and helped identify transitional progenitors associated with key branching events. For embryoid body differentiation, MARGARET identified novel transitional populations that were validated by bulk sequencing and functionally characterized different precursor populations in the mesoderm lineage. For colon differentiation, MARGARET characterized the lineage for BEST4/OTOP2 cells and the heterogeneity in goblet cell lineage in the colon under normal and inflamed ulcerative colitis conditions. Finally, we demonstrated that MARGARET can scale to large scRNA-seq datasets consisting of ∼ millions of cells.

Differential gene expression analysis identified determinants of cell fate plasticity during radiation-induced regeneration in Drosophila.

Ionizing radiation (IR) is used to treat half of all cancer patients because of its ability to kill cells. IR, however, can induce stem cell-like properties in non-stem cancer cells, potentiating tumor regrowth and reduced therapeutic success. We identified previously a subpopulation of cells in Drosophila larval wing discs that exhibit IR-induced stem cell-like properties. These cells reside in the future wing hinge, are resistant to IR-induced apoptosis, and are capable of translocating, changing fate, and participating in regenerating the pouch that suffers more IR-induced apoptosis. We used here a combination of lineage tracing, FACS-sorting of cells that change fate, genome-wide RNAseq, and functional testing of 42 genes, to identify two key changes that are required cell-autonomously for IR-induced hinge-to-pouch fate change: (1) repression of hinge determinants Wg (Drosophila Wnt1) and conserved zinc-finger transcription factor Zfh2 and (2) upregulation of three ribosome biogenesis factors. Additional data indicate a role for Myc, a transcriptional activator of ribosome biogenesis genes, in the process. These results provide a molecular understanding of IR-induced cell fate plasticity that may be leveraged to improve radiation therapy.

Yap-Sox9 signaling determines hepatocyte plasticity and lineage-specific hepatocarcinogenesis

Primary liver tumors comprise distinct subtypes. A subset of intrahepatic cholangiocarcinoma (iCCA) can arise from cell fate reprogramming of mature hepatocytes in mouse models. However, the underpinning of cell fate plasticity during hepatocarcinogenesis is still poorly understood, hampering therapeutic development for primary liver cancer. As YAP activation induces liver tumor formation and cell fate plasticity, we investigated the role of Sox9, a transcription factor downstream of Yap activation that is expressed in biliary epithelial cells (BECs), in Yap-induced cell fate plasticity during hepatocarcinogenesis. To evaluate the function of Sox9 in YAP-induced hepatocarcinogenesis invivo, we used several genetic mouse models of inducible hepatocyte-specific YAP activation with simultaneous Sox9 removal. Cell fate reprogramming was determined by lineage tracing and immunohistochemistry. The molecular mechanism underlying Yap and Sox9 function in hepatocyte plasticity was investigated by transcription and transcriptomic analyses of mouse and human liver tumors. Sox9, a marker of liver progenitor cells (LPCs) and BECs, is differentially required in YAP-induced stepwise hepatocyte programming. While Sox9 has a limited role in hepatocyte dedifferentiation to LPCs, it is required for BEC differentiation from LPCs. YAP activation in Sox9-deficient hepatocytes resulted in more aggressive HCC with enhanced Yap activity at the expense of iCCA-like tumors. Furthermore, we showed that 20% of primary human liver tumors were associated with a YAP activation signature, and tumor plasticity is highly correlated with YAP activation and SOX9 expression. Our data demonstrated that Yap-Sox9 signaling determines hepatocyte plasticity and tumor heterogeneity in hepatocarcinogenesis in both mouse and human liver tumors. We identified Sox9 as a critical transcription factor required for Yap-induced hepatocyte cell fate reprogramming during hepatocarcinogenesis. Sox9, a marker of liver progenitor cells and bile duct lining cells, is a downstream target of YAP protein activation. Herein, we found that YAP activation in hepatocytes leads to a transition from mature hepatocytes to liver progenitor cells and then to bile duct lining cells. Sox9 is required in the second step during mouse hepatocarcinogenesis. We also found that human YAP and SOX9 may play similar roles in liver cancers.

Notch-mediated re-specification of neuronal identity during central nervous system development

Neuronal identity has long been thought of as immutable, so that once a cell acquires a specific fate, it is maintained for life.1 Studies using the overexpression of potent transcription factors to experimentally reprogram neuronal fate in the mouse neocortex2,3 and retina4,5 have challenged this notion by revealing that post-mitotic neurons can switch their identity. Whether fate reprogramming is part of normal development in the central nervous system (CNS) is unclear. While there are some reports of physiological cell fate reprogramming in invertebrates,6,7 and in the vertebrate peripheral nervous system,8 endogenous fate reprogramming in the vertebrate CNS has not been documented. Here, we demonstrate spontaneous fate re-specification in an interneuron lineage in the zebrafish retina. We show that the visual system homeobox 1 (vsx1)-expressing lineage, which has been associated exclusively with excitatory bipolar cell (BC) interneurons,9-12 also generates inhibitory amacrine cells (ACs). We identify a role for Notch signaling in conferring plasticity to nascent vsx1 BCs, allowing suitable transcription factor programs to re-specify them to an AC fate. Overstimulating Notch signaling enhances this physiological phenotype so that both daughters of a vsx1 progenitor differentiate into ACs and partially differentiated vsx1 BCs can be converted into ACs. Furthermore, this physiological re-specification can be mimicked to allow experimental induction of an entirely distinct fate, that of retinal projection neurons, from the vsx1 lineage. Our observations reveal unanticipated plasticity of cell fate during retinal development.

Advanced Technologies to Target Cardiac Cell Fate Plasticity for Heart Regeneration.

The adult human heart can only adapt to heart diseases by starting a myocardial remodeling process to compensate for the loss of functional cardiomyocytes, which ultimately develop into heart failure. In recent decades, the evolution of new strategies to regenerate the injured myocardium based on cellular reprogramming represents a revolutionary new paradigm for cardiac repair by targeting some key signaling molecules governing cardiac cell fate plasticity. While the indirect reprogramming routes require an in vitro engineered 3D tissue to be transplanted in vivo, the direct cardiac reprogramming would allow the administration of reprogramming factors directly in situ, thus holding great potential as in vivo treatment for clinical applications. In this framework, cellular reprogramming in partnership with nanotechnologies and bioengineering will offer new perspectives in the field of cardiovascular research for disease modeling, drug screening, and tissue engineering applications. In this review, we will summarize the recent progress in developing innovative therapeutic strategies based on manipulating cardiac cell fate plasticity in combination with bioengineering and nanotechnology-based approaches for targeting the failing heart.

Myeloid Progenitors of Defined Features Provide Permissiveness to Leukemic Transformation

The conventional view of oncogenesis posits that driver mutations, when present in appropriate cell types, initiate malignant transformation. At the apex of the differentiation hierarchy, stem cells, such as hematopoietic stem cells, have been suggested to be the cell-of-origin for a variety of hematopoietic malignancies. However, since the mutation bearing hematopoietic stem cells can differentiate to give rise to downstream progeny, it suggests the possibility that some of the downstream cells be the relevant cell-of-origin permitting malignant transformation. Given that cancer cell-of-origin has been a product of logic deduction evading rigorous experimentation, whether and how their unique cellular properties, if any, contribute to transformation remain undetermined.To experimentally define the key properties of the cell-of-origin permissive to oncogenic transformation, we constructed a doxycycline (Dox) inducible MLL-AF9 knock-in mouse model, which allows precise temporal control of oncogene expression in desired hematopoietic cells, in vitro and in vivo . Whole body induction by Dox led to 100% lethality due to acute myeloid leukemia (AML) within 6-8 weeks, while the un-induced littermates remained healthy, confirming the inducibility and potency of the MLL-AF9 oncogene. To minimize the differences contributed by cellular differentiation stages and to focus solely on the contributions by specific cellular properties, we narrowed our analysis on a phenotypically homogeneous population, the granulocyte-macrophage progenitors (GMPs), which have been shown to be critically important in supporting leukemic development (Ugale et al ., 2014, Cell Reports; Ye et al ., 2015, Cell Stem Cell). We found that although MLL-AF9 expression can be induced in all GMPs, only a subset (~25%) of them is permissive to transformation. These results prompted us to investigate the underlying cellular and molecular properties that enable oncogenic transformation.Inspired by our earlier work demonstrating that the cell fate plasticity of GMPs is related to their rapid cell cycle behavior (Guo et al ., 2014, Cell), we tested whether the rapid proliferation of GMPs underlies their permissiveness to oncogenic transformation. If so, transformation potential should be enriched in the faster-proliferating GMP subsets. To test this possibility, we designed single cell assays to investigate the transformation efficiencies of GMPs with regard to their cell cycle speeds. Specifically, the intrinsic cell cycle speed was measured within 24 hours of Dox addition, and transformation status of hundreds of individual GMPs was determined by methylcellulose colony formation assay. Within the first 24 hours of Dox addition, the cell cycle speed remained unchanged, reflecting the intrinsic cell cycle kinetics, rather than a proliferative response to oncogene expression. Our results show that the faster a GMP divides, the more likely it transforms in response to MLL-AF9 expression. Strikingly, most GMPs that can divide more than 3 times within 24 hours are transformed. These data indicate the fastest cycling GMPs provide the oncogene with a permissive cellular context to exert its function. Furthermore, prospectively isolated faster cycling GMPs enriched for transformed colony formation in vitro, and induced earlier lethal AML in vivo . Importantly, transient deceleration of GMPs for 48 hours by low-dose CDK4/6 inhibitor, Palbociclib, significantly reduced transformation efficiency by ~ 60%. Taken together, these results demonstrate that MLL-AF9 initiated transformation is dependent on specific cellular properties which are associated with a rapid cell cycle. Genomic analyses of the transformation-permissive GMPs, using non-permissive GMPs as controls, are being performed to define the permissiveness in molecular details, on both transcriptional and epigenetic levels. DisclosuresNo relevant conflicts of interest to declare.

Context-aware synthetic biology by controller design: Engineering the mammalian cell.

The rise of systems biology has ushered a new paradigm: the view of the cell as a system that processes environmental inputs to drive phenotypic outputs. Synthetic biology provides a complementary approach, allowing us to program cell behavior through the addition of synthetic genetic devices into the cellular processor. These devices, and the complex genetic circuits they compose, are engineered using a design-prototype-test cycle, allowing for predictable device performance to be achieved in a context-dependent manner. Within mammalian cells, context effects impact device performance at multiple scales, including the genetic, cellular, and extracellular levels. In order for synthetic genetic devices to achieve predictable behaviors, approaches to overcome context dependence are necessary. Here, we describe control systems approaches for achieving context-aware devices that are robust to context effects. We then consider cell fate programing as a case study to explore the potential impact of context-aware devices for regenerative medicine applications.

Regeneration Potential of Jellyfish: Cellular Mechanisms and Molecular Insights.

Medusozoans, the Cnidarian subphylum, have multiple life stages including sessile polyps and free-swimming medusae or jellyfish, which are typically bell-shaped gelatinous zooplanktons that exhibit diverse morphologies. Despite having a relatively complex body structure with well-developed muscles and nervous systems, the adult medusa stage maintains a high regenerative ability that enables organ regeneration as well as whole body reconstitution from the part of the body. This remarkable regeneration potential of jellyfish has long been acknowledged in different species; however, recent studies have begun dissecting the exact processes underpinning regeneration events. In this article, we introduce the current understanding of regeneration mechanisms in medusae, particularly focusing on cellular behaviors during regeneration such as wound healing, blastema formation by stem/progenitor cells or cell fate plasticity, and the organism-level patterning that restores radial symmetry. We also discuss putative molecular mechanisms involved in regeneration processes and introduce a variety of novel model jellyfish species in the effort to understand common principles and diverse mechanisms underlying the regeneration of complex organs and the entire body.