Expression Of Lineage-specific Transcription Factors Research Articles

Landscape and Treatment Options of Shapeshifting Small Cell Lung Cancer.

Small cell lung cancer (SCLC) is a deadly neuroendocrine malignancy, notorious for its rapid tumor growth, early metastasis, and relatively "cold" immune environment. Only standard chemotherapies and a few immune checkpoint inhibitors have been approved for SCLC treatment, revealing an urgent need for novel therapeutic approaches. Moreover, SCLC has been recently recognized as a malignancy with high intratumoral and intertumoral heterogeneity, which explains the modest response rate in some patients and the early relapse. Molecular subtypes defined by the expression of lineage-specific transcription factors (ASCL1, NEUROD1, POU2F3, and, in some studies, YAP1) or immune-related genes display different degrees of neuroendocrine differentiation, immune cell infiltration, and response to treatment. Despite the complexity of this malignancy, a few biomarkers and targets have been identified and many promising drugs are currently undergoing clinical trials. In this review, we integrate the current progress on the genomic landscape of this shapeshifting malignancy, the characteristics and treatment vulnerabilities of each subtype, and promising drugs in clinical phases.

Abstract PO4-16-03: Ectopic expression of lineage-specific transcription factors in Breast Cancer

Abstract Background: Lineage-specific transcription factors (LTFs) are purported to define cellular lineages and their expression could be used to identify subtypes of cancer. In many cancers, tuft cell differentiation appears to have prognostic impact. However, such differentiation has not been documented in breast cancer. Methods: We sought to study the fidelity of LTFs (MYOG, POU2F3, FOXA1, FOXA2, FOXA3, FOXJ1, HNF4A, HNF4G, SPIB, SOX8, GRHL1, GRHL2, and GRHL3) for their presence and prognostic impact in breast cancer. Single cell RNA data from GSE 161529, which contains 421,761 single cells, was downloaded and analyzed for the expression and co-expression of LTFs. The data was first subdivided based on ER+ and TNBC status. Analysis was performed using cells from 10ER+ samples and 4 TNBC samples. Cell clustering using Louvain’s method was run using all genes. Cell type assignment was then performed using well-known markers for both tumor and lymphocyte marker from existing publications. Further analysis was restricted to keratin expressing cells and to study the presence and co-expression of LTFs in the epithelial cells. KM-Plotter was employed to determine the prognostic value of these LTFs. The analysis was further stratified by expression of estrogen receptor positive and negative status. The endpoint of relapse free survival was used, as it provided data on the largest number of patients. Results: Single-cell analysis pipeline successfully identified distinct populations of tumor epithelial cells accompanied by various lymphocytes, including B cells, CD8+ T cells, CD4+ T cells, dendritic cells, and macrophages, in both ER+ tumors and TNBC tumor samples. We further confirmed the expression of the LTFs in the epithelial (keratin+) compartment. Expression of all LTFs (except MYOG, and FOXA2) was scattered in the different epithelial clusters. Further analysis of single LTFs showed differentiational co-expression based on ER+ status. POU2F3+/ER+ cells also co-expressed FOXA1 and GRHL1 while POU2F3+/ER- cells were GRHL1+ and (weak) SPIB+. The analysis of LTFs using KM-Plotter tool revealed their expression documenting multiple cell lineages in breast cancer; this was noted in ER+, HER2+ cancers and TNBCs. The directionality of the outcome (good or bad prognosis) varied with the specific TFs and subtype of breast cancer. High expression of POU2F3, and FOXA1, and SPIB were associated with better relapse-free survival (RFS) in ER+ breast cancer. POU2F3, FOXA2, FOXA3 were associated with worse RFS, whereas SPIB was with prognosis in ER- breast cancers. Conclusions: We for the first time document the expression of LTFs in breast cancer and their prognostic relevance based on ER+ and ERneg status. Expression of POU2F3, a marker of tuft cells which are known to regulate immune response, was noted to be associated with improved outcomes in ER+ tumors. Further characterization of the relevance of LTFs and their co-expression with respect to therapeutic response is ongoing. Citation Format: Sunil Badve, Jian Hu, Yu Zhang, Yesim Gokmen-Polar. Ectopic expression of lineage-specific transcription factors in Breast Cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-16-03.

Bcl-6 expression by CD4+ T cells determines concomitant immunity and host resistance across distinct parasitic infections

Cluster of differentiation (CD4+) T cells consist of multiple subtypes, defined by expression of lineage-specific transcription factors, that contribute to the control of infectious diseases by providing help to immune and nonimmune target cells. In the current study, we examined the role of B cell lymphoma (Bcl)-6, a transcriptional repressor and master regulator of T follicular helper cell differentiation, in T cell-mediated host defense against intestinal and systemic parasitic infections. We demonstrate that while Bcl-6 expression by CD4+ T cells is critical for antibody-mediated protective immunity against secondary infection with the nematode Heligmosoides polygyrus bakeri, it paradoxically compromises worm expulsion during primary infection by limiting the generation of interleukin-10 (IL-10)-producing Gata3+ T helper 2 cells. Enhanced worm expulsion in the absence of Bcl-6 expressing T cells was associated with amplified intestinal goblet cell differentiation and increased generation of alternatively activated macrophages, effects that were reversed by neutralization of IL-10 signals. An increase in IL-10 production by Bcl-6-deficient CD4+ T cells was also evident in the context of systemic Leishmania donovani infection, but in contrast to Heligmosoides polygyrus bakeri infection, compromised T helper 1-mediated liver macrophage activation and increased susceptibility to this distinct parasitic challenge. Collectively, our studies suggest that host defense pathways that protect against parasite superinfection and lethal systemic protozoal infections can be engaged at the cost of compromised primary resistance to well-tolerated helminths.

Robust protein-based engineering of hepatocyte-like cells from human mesenchymal stem cells.

Cells of interest can be prepared from somatic cells by forced expression of lineage-specific transcription factors, but it is required to establish a vector-free system for their clinical use. Here, we report a protein-based artificial transcription system for engineering hepatocyte-like cells from human umbilical cord-derived mesenchymal stem cells (MSCs). MSCs were treated for 5 days with 4 artificial transcription factors (4F), which targeted hepatocyte nuclear factor (HNF)1α, HNF3γ, HNF4α, and GATA-binding protein 4 (GATA4). Then, engineered MSCs (4F-Heps) were subjected to epigenetic analysis, biochemical analysis and flow cytometry analysis with antibodies to marker proteins of mature hepatocytes and hepatic progenitors such as delta-like homolog 1 (DLK1) and trophoblast cell surface antigen 2 (TROP2). Functional properties of the cells were also examined by injecting them to mice with lethal hepatic failure. Epigenetic analysis revealed that a 5-day treatment of 4F upregulated the expression of genes involved in hepatic differentiation, and repressed genes related to pluripotency of MSCs. Flow cytometry analysis detected that 4F-Heps were composed of small numbers of mature hepatocytes (at most 1%), bile duct cells (~19%) and hepatic progenitors (~50%). Interestingly, ~20% of 4F-Heps were positive for cytochrome P450 3A4, 80% of which were DLK1-positive. Injection of 4F-Heps significantly increased survival of mice with lethal hepatic failure, and transplanted 4F-Heps expanded to more than 50-fold of human albumin-positive cells in the mouse livers, well consistent with the observation that 4F-Heps contained DLK1-positive and/or TROP2-positive cells. Taken together with observations that 4F-Heps were not tumorigenic in immunocompromised mice for at least 2 years, we propose that this artificial transcription system is a versatile tool for cell therapy for hepatic failures.

Progress and challenges in directing the differentiation of human iPSCs into spinal motor neurons.

Motor neuron (MN) diseases, including amyotrophic lateral sclerosis, progressive bulbar palsy, primary lateral sclerosis and spinal muscular atrophy, cause progressive paralysis and, in many cases, death. A better understanding of the molecular mechanisms of pathogenesis is urgently needed to identify more effective therapies. However, studying MNs has been extremely difficult because they are inaccessible in the spinal cord. Induced pluripotent stem cells (iPSCs) can generate a theoretically limitless number of MNs from a specific patient, making them powerful tools for studying MN diseases. However, to reach their potential, iPSCs need to be directed to efficiently differentiate into functional MNs. Here, we review the reported differentiation protocols for spinal MNs, including induction with small molecules, expression of lineage-specific transcription factors, 2-dimensional and 3-dimensional cultures, as well as the implementation of microfluidics devices and co-cultures with other cell types, including skeletal muscle. We will summarize the advantages and disadvantages of each strategy. In addition, we will provide insights into how to address some of the remaining challenges, including reproducibly obtaining mature and aged MNs.

Generation of a Pure Culture of Neuron-like Cells with a Glutamatergic Phenotype from Mouse Astrocytes.

Astrocyte-to-neuron reprogramming is a promising therapeutic approach for treatment of neurodegenerative diseases. The use of small molecules as an alternative to the virus-mediated ectopic expression of lineage-specific transcription factors negates the tumorigenic risk associated with viral genetic manipulation and uncontrolled differentiation of stem cells. However, because previously developed methods for small-molecule reprogramming of astrocytes to neurons are multistep, complex, and lengthy, their applications in biomedicine, including clinical treatment, are limited. Therefore, our objective in this study was to develop a novel chemical-based approach to the cellular reprogramming of astrocytes into neurons with high efficiency and low complexity. To accomplish that, we used C8-D1a, a mouse astrocyte cell line, to assess the role of small molecules in reprogramming protocols that otherwise suffer from inconsistencies caused by variations in donor of the primary cell. We developed a new protocol by which a chemical mixture formulated with Y26732, DAPT, RepSox, CHIR99021, ruxolitinib, and SAG rapidly and efficiently induced the neural reprogramming of astrocytes in four days, with a conversion efficiency of 82 ± 6%. Upon exposure to the maturation medium, those reprogrammed cells acquired a glutaminergic phenotype over the next eleven days. We also demonstrated the neuronal functionality of the induced cells by confirming KCL-induced calcium flux.

Oncohistone mutations enhance chromatin remodeling and alter cell fates.

Whole genome sequencing data mining efforts have revealed numerous histone mutations in a wide range of cancer types. These occur in all four core histones in both the tail and globular domains and remain largely uncharacterized. Here we used two high-throughput approaches, a DNA-barcoded mononucleosome library and a humanized yeast library, to profile the biochemical and cellular effects of these mutations. We identified cancer-associated mutations in the histone globular domains that enhance fundamental chromatin remodeling processes, histone exchange and nucleosome sliding, and are lethal in yeast. In mammalian cells, these mutations upregulate cancer-associated gene pathways and inhibit cellular differentiation by altering expression of lineage-specific transcription factors. This work represents a comprehensive functional analysis of the histone mutational landscape in human cancers and leads to a model in which histone mutations that perturb nucleosome remodeling may contribute to disease development and/or progression.

Modeling and targeting of erythroleukemia by hematopoietic genome editing

Acute erythroid leukemia (AEL) is characterized by a distinct morphology, mutational spectrum, lack of preclinical models, and poor prognosis. Here, using multiplexed genome editing of mouse hematopoietic stem and progenitor cells and transplant assays, we developed preclinical models of AEL and non-erythroid acute leukemia and describe the central role of mutational cooperativity in determining leukemia lineage. Different combination of mutations in Trp53, Bcor, Dnmt3a, Rb1, and Nfix resulted in the development of leukemia with an erythroid phenotype, accompanied by the acquisition of alterations in signaling and transcription factor genes that recapitulate human AEL by cross-species genomic analysis. Clonal expansion during tumor evolution was driven by mutational cooccurrence, with clones harboring a higher number of founder and secondary lesions (eg, mutations in signaling genes) showing greater evolutionary fitness. Mouse and human AEL exhibited deregulation of genes regulating erythroid development, notably Gata1, Klf1, and Nfe2, driven by the interaction of mutations of the epigenetic modifiers Dnmt3a and Tet2 that perturbed methylation and thus expression of lineage-specific transcription factors. The established mouse leukemias were used as a platform for drug screening. Drug sensitivity was associated with the leukemia genotype, with the poly (ADP-ribose) polymerase inhibitor talazoparib and the demethylating agent decitabine efficacious in Trp53/Bcor–mutant AEL, CDK7/9 inhibitors in Trp53/Bcor/Dnmt3a–mutant AEL, and gemcitabine and bromodomain inhibitors in NUP98-KDM5A leukemia. In conclusion, combinatorial genome editing has shown the interplay of founding and secondary genetic alterations in phenotype and clonal evolution, epigenetic regulation of lineage-specific transcription factors, and therapeutic tractability in erythroid leukemogenesis.

Hematological Disorders in Human Patients with Son Mutations

Precise regulation of transcription and RNA splicing is critical for controlling hematopoietic cell fate determination and lineage differentiation. Alteration of expression of lineage-specific transcription factors and several core spliceosome components in hematopoietic malignancies highlight the significance of abnormal transcription and RNA splicing as disease-causing factors. Our group previously demonstrated that SON, a large nuclear speckle protein possessing dual abilities to bind both DNA and RNA, functions as a splicing factor as well as a transcriptional repressor. We recently identified heterozygous loss-of-function mutations in the SON gene from children with intellectual disability and developmental delay often with a broad spectrum of other congenital anomalies. The disorder caused by SON haploinsufficiency has been designated as ZTTK syndrome (Zhu-Tokita-Takenouchi-Kim syndrome; OMIM #617140). The majority of the mutations found in these patients are frameshift or nonsense mutations which cause degradation of the mutation-bearing transcript. While the most prominent features of these patients are brain malformations and musculoskeletal abnormalities, we identified various hematological disorders from children with ZTTK syndrome. Notable symptoms include bone marrow failure, severe anemia, immunoglobulin deficiency, thalassemia, polycythemia, polycythemia vera, stroke due to blood clots, and leukocytopenia. Apart from the ZTTK syndrome, SON is known to be upregulated in acute myeloid leukemia (AML) patients and is correlated to altered hematopoietic differentiation. To investigate how altered SON expression affects hematopoiesis, we generated a mouse line with the Son gene deleted specifically in the hematopoietic lineage. Homozygous deletion of Son in hematopoietic lineage led to embryonic lethality, indicating that SON expression in blood cells is indispensable during development. Mice with heterozygous deletion of Son in the hematopoietic lineage were viable and born without notable defects or sign of diseases. However, there is a significant decrease in bone marrow cellularity in the mice with heterozygous deletion of Son. Furthermore, Son haploinsufficiency decreased the size of the lineage negative (Lin-) cell population and short-term hematopoietic stem cell (ST-HSC) population with a concurrent increase in megakaryocyte/erythrocyte lineage-biased multipotent progenitors (MPP2) within hematopoietic stem/progenitor cells. These findings suggest that the level of Son expression potentially affects stem cell maintenance and MPP lineage bias, and the distortion of the subpopulation balance within hematopoietic stem/progenitors is possibly linked to multiple hematological disorders. Our ongoing analyses of hematopoiesis and gene expression changes using this mouse model will expand our knowledge about the role of SON in several hematological disorders and benefit clinical practice for ZTTK syndrome patients. Disclosures No relevant conflicts of interest to declare.

Single-cell analyses identify distinct and intermediate states of zebrafish pancreatic islet development.

Pancreatic endocrine islets are vital for glucose homeostasis. However, the islet developmental trajectory and its regulatory network are not well understood. To define the features of these specification and differentiation processes, we isolated individual islet cells from TgBAC(neurod1:EGFP) transgenic zebrafish and analyzed islet developmental dynamics across four different embryonic stages using a single-cell RNA-seq strategy. We identified proliferative endocrine progenitors, which could be further categorized by different cell cycle phases with the G1/S subpopulation displaying a distinct differentiation potential. We identified endocrine precursors, a heterogeneous intermediate-state population consisting of lineage-primed alpha, beta and delta cells that were characterized by the expression of lineage-specific transcription factors and relatively low expression of terminally differentiation markers. The terminally differentiated alpha, beta, and delta cells displayed stage-dependent differentiation states, which were related to their functional maturation. Our data unveiled distinct states, events and molecular features during the islet developmental transition, and provided resources to comprehensively understand the lineage hierarchy of islet development at the single-cell level.

Direct Reprograming to Regenerate Myocardium and Repair Its Pacemaker and Conduction System.

The regenerative medicine field has been revolutionized by the direct conversion of one cell type to another by ectopic expression of lineage-specific transcription factors. The direct reprogramming of fibroblasts to induced cardiac myocytes (iCMs) by core cardiac transcription factors (Gata4, Mef2c, Tbx5) both in vitro and in vivo has paved the way in cardiac regeneration and repair. Several independent research groups have successfully reported the direct reprogramming of fibroblasts in injured myocardium to cardiac myocytes employing a variety of approaches that rely on transcription factors, small molecules, and micro RNAs (miRNAs). Recently, this technology has been considered for local repair of the pacemaker and the cardiac conduction system. To address this, we will first discuss the direct reprograming advancements in the setting of working myocardium regeneration, and then elaborate on how this technology can be applied to repair the cardiac pacemaker and the conduction system.

Exogenous TPRX1 homeoprotein modulates the gene expression of lineage-specific transcription factors in human embryonal carcinoma cells

ABSTRACTWe reported previously that EGAM1 homeoproteins (EGAM1 and EGAM1N), transcribed from the Crxos gene as splicing variants, are expressed in preimplantation mouse embryos and mouse embryonic stem (ES) cells. Exogenous expression of these proteins affects the maintenance of an undifferentiated state and the progression of differentiation in mouse ES cells. Human tetrapeptide-repeat homeobox 1 (TPRX1), a member of the eutherian totipotent cell homeobox (ETCHbox) genes, is an ortholog of Crxos. However, the roles of TPRX1 in the differentiation of human pluripotent cells are still unknown. Because the TPRX1 transcripts were undetectable in an undifferentiated state and during the progression of differentiation in wild-type human embryonal carcinoma NT2/D1 cells, it would be advantageous to clarify the relationship between the exogenous expression of TPRX1 and the induction of genes encoding lineage-specific transcription factors in pluripotent cells. The expression of GATA6 and FOXA2, crucial transcription factors for the formation of the primitive endoderm, was upregulated, whereas that of CDX2, a crucial transcription factor for the formation of the trophectoderm, was downregulated by enforced expression of TPRX1. Overall, we suggest that TPRX1 is capable of modulating the expression of lineage-specific transcription factors in pluripotent cells derived from humans.

Comparative genomic analysis of embryonic, lineage-converted, and stem cell-derived motor neurons

Advances in stem cell science allow the production of different cell types in vitro either through the recapitulation of developmental processes, often termed 'directed differentiation', or the forced expression of lineage-specific transcription factors. Although cells produced by both approaches are increasingly used in translational applications, their quantitative similarity to their primary counterparts remains largely unresolved. To investigate the similarity between in vitro-derived and primary cell types, we harvested and purified mouse spinal motor neurons and compared them with motor neurons produced by transcription factor-mediated lineage conversion of fibroblasts or directed differentiation of pluripotent stem cells. To enable unbiased analysis of these motor neuron types and their cells of origin, we then subjected them to whole transcriptome and DNA methylome analysis by RNA sequencing (RNA-seq) and reduced representation bisulfite sequencing (RRBS). Despite major differences in methodology, lineage conversion and directed differentiation both produce cells that closely approximate the primary motor neuron state. However, we identify differences in Fas signaling, the Hox code and synaptic gene expression between lineage-converted and directed differentiation motor neurons that affect their utility in translational studies.

Nuclear CRX and FOXJ1 Expression Differentiates Non-Germ Cell Pineal Region Tumors and Supports the Ependymal Differentiation of Papillary Tumor of the Pineal Region.

Papillary tumor of the pineal region (PTPR) is a neuroepithelial neoplasm first described in 2003. Despite the anatomic association of PTPR with the pineal gland, the features of these tumors resemble those of the ependymal circumventricular subcommissural organ (SCO) of the posterior third ventricle. Given the presumed distinct derivation of PTPR and pineal parenchymal tumors, we hypothesized that expression of lineage-specific transcription factors could distinguish these tumors and provide additional insight into the differentiation of PTPR. A broad series of pineal region samples was reviewed, including 7 benign pineal glands, 4 pineal cysts, 13 pineocytomas, 28 pineal parenchymal tumors of intermediate differentiation, 11 pineoblastomas, and 18 PTPR. All samples were evaluated by immunohistochemistry for expression of CRX, a master transcriptional regulator of photoreceptor differentiation expressed in pineal gland and retina and/or FOXJ1, a master transcriptional regulator of ciliogenesis expressed in normal ependymal cells and ependymal neoplasms. Diffuse nuclear CRX expression is present in 100% of pineal samples. FOXJ1 is negative in all pineal samples. CRX staining is present in 53% of PTPR, though expression is nearly always limited to rare cells. Diffuse nuclear FOXJ1 expression is present in 100% of PTPR. Fetal human SCO diffusely expressed FOXJ1 but was negative for CRX. Immunohistochemistry for FOXJ1 and CRX differentiates non-germ cell pineal region tumors with high sensitivity and specificity, including pineal parenchymal tumors and PTPR. Our findings support the hypothesis that PTPR have ependymal differentiation and are phenotypically more similar to SCO than pineal gland.

Mixed Gangliocytoma-Pituitary Adenoma: Insights on the Pathogenesis of a Rare Sellar Tumor.

Gangliocytomas originating in the sellar region are rare; most are tumors composed of gangliocytic and pituitary adenomatous elements, forming the so-called mixed gangliocytoma-pituitary adenoma. The majority of mixed gangliocytoma adenomas are associated with endocrinopathies, mainly acromegaly and less often Cushing disease and hyperprolactinemia. In the present study, 10 cases of mixed gangliocytoma and somatotroph adenomas were evaluated for patterns of cellular differentiation and expression of lineage-specific transcription factors. The tumors were characterized by immunohistochemistry for pituitary hormones, cytokeratins, Pit-1, and the neuronal markers NeuN, neurofilaments (NFP), and MAP2. Double-labeling immunohistochemistry for Pit-1/GH, Pit-1/NFP, Pit-1/MAP2, and NeuN/GH was performed in 9/10 tumors. Our data demonstrate that both adenomatous and ganglionic cells express the acidophilic lineage transcription factor Pit-1. Although mixed gangliocytomas and somatotroph adenomas show histologically distinct cellular populations, there is at least a small population of cells that coexpress the Pit-1 transcription factor and neuronal-associated cytoskeletal proteins favoring the theory of transdifferentiation of neuroendocrine cells into neuronal elements of these mixed tumors.

Baf60b-mediated ATM-p53 activation blocks cell identity conversion by sensing chromatin opening

Lineage conversion by expression of lineage-specific transcription factors is a process of epigenetic remodeling that has low efficiency. The mechanism by which a cell resists lineage conversion is largely unknown. Using hepatic-specific transcription factors Foxa3, Hnf1α and Gata4 (3TF) to induce hepatic conversion in mouse fibroblasts, we showed that 3TF induced strong activation of the ATM-p53 pathway, which led to proliferation arrest and cell death, and it further prevented hepatic conversion. Notably, ATM activation, independent of DNA damage, responded to chromatin opening during hepatic conversion. By characterizing the early molecular events during hepatic conversion, we found that Baf60b, a member of the SWI/SNF chromatin remodeling complex, links chromatin opening to ATM activation by facilitating ATM recruitment to the open chromatin regions of a panel of hepatic gene loci. These findings shed light on cellular responses to lineage conversion by revealing a function of the ATM-p53 pathway in sensing chromatin opening.

DNMT3A and TET2 Cooperate and Compete to Maintain HSC DNA Methylation

Abstract Combined loss of DNMT3A and TET2 in HSCs promotes lineage-specific transcription factor expression.

Chemical conversion of mouse fibroblasts into functional dopaminergic neurons

Ectopic expression of lineage-specific transcription factors facilitates the conversion of mammalian somatic cells into dopaminergic (DA) neurons, which is a promising strategy for cell therapy of Parkinson's disease (PD). However, this approach still has some drawbacks limiting its clinical application due to the potential risks of integrating vectors into the host genome. Therefore, it is critical to seek a more desired approach to generate DA neurons derived from mammalian somatic cells. Here, we report that mouse embryonic fibroblasts (MEFs) can be efficiently converted into DA neurons by using small molecules along with specific growth factors. These neuron-like cells generate DA neuronal morphology, and acquire immunocytochemical and calcium imaging special for neuronal electrophysiological profile. More importantly, these converted cells can secrete dopamine, indicating that they are functionally similar to DA neurons. Taken together, our study might provide a promising cell source for treating PD by using chemical approach without introduction of exogenous transcription factors.

Engineering cell fate: Spotlight on cell-activation and signaling-directed lineage conversion.

Direct reprogramming of specialized cells into other cell types has revolutionized the fields of stem cell, differentiation, and regenerative medicine. Direct reprogramming technology can convert various differentiated cell types to other fates by the forced expression of lineage-specific transcription factors. In addition to this approach, transdifferentiation can be induced in somatic cells by a method named cell-activation and signaling-directed (CASD) lineage conversion, which uses pluripotency reprogramming factors in combination with specific differentiation signals. This approach is capable of generating tissue-specific progenitors in addition to functional mature cells through a challenging transitory (pluripotent or non-pluripotent) state. Interestingly, the CASD lineage conversion has been accomplished by using small molecules and growth factors in a chemical-only paradigm. This approach will have a substantial positive impact on the field and bring reprogramming technology into the translational pipeline. From the regenerative medicine perspective, in future, new therapies might be designed based on the patient's own cells that are directly reprogrammed into the desired cell types in vitro and in situ. This review provides an overview of direct reprogramming to different lineages and describes what is known about the cell-activation and signaling-directed transdifferentiation.

DNMT3A and TET2 compete and cooperate to repress lineage-specific transcription factors in hematopoietic stem cells.

Mutations in the epigenetic modifiers DNMT3A and TET2 non-randomly co-occur in lymphoma and leukemia despite their epistasis in the methylation-hydroxymethylation pathway. Using Dnmt3a and Tet2 double knock-out (DKO) mice in which malignancy development is accelerated, we show that the DKO methylome reflects regions of independent, competitive and cooperative activity. Expression of lineage-specific transcription factors, including the erythroid regulator Klf1 is upregulated in DKO HSCs. DNMT3A and TET2 both repress Klf1 suggesting a model of cooperative inhibition by the epigenetic modifiers. These data demonstrate a dual role for TET2 in promoting and inhibiting HSC differentiation, loss of which, along with DNMT3A, obstructs differentiation leading to transformation.