Cell Lineage Determination Research Articles

StSNV: a comprehensive resource of SNVs in spatial transcriptome.

Single nucleotide variants (SNVs), as important componentsof genetic variation, affect gene expression, function and phenotype. Mining and summarizing the spatial distribution of SNVs in diseased and normal tissues for a better understanding of their characteristics and potential roles in cell-lineage determination, aging, or disease occurrence is significant. Herein, we have developed a comprehensive spatial mutation resource stSNV (http://bio-bigdata.hrbmu.edu.cn/stSNV/index.jsp), which provides an atlas of spatial SNVs in major diseased and normal tissues of human and mouse. stSNV documents 42202 spatial mutated genes involving 898908 SNVs called from 730067 spots within 450 slices from 19 diseased and 28 normal tissues. Importantly, potential characteristics of SNVs are explored and provided by analyzing the perturbation of the SNVs to gene expression, spatial communication,biological function, region-specific mutated genes, spatial mutant signatures, SNV-cell co-localizationand mutation core region. All these spatial mutation data and in-depth analyses have been integrated into a user-friendly interface, visualized through intuitive tables and various image formats. Flexible tools are developed to explore co-localization among clusters, genes, cell types and SNVs in the same slice. In summary, stSNV as a valuable resourcehelps to dissect intra-tissue genetic heterogeneity and lays the groundwork for understanding the SNVs' biological regulatory mechanisms.

OrganogenesisDB: A Comprehensive Database Exploring the Cell-Type Identities and Gene Expression Dynamics during Organogenesis.

Organogenesis,the phase ofembryonic developmentthat starts at the end ofgastrulationand continues untilbirth is the critical process for understanding cellular differentiation and maturation during organ development. The rapid development of single-cell transcriptomics technology has led to many novel discoveries in understanding organogenesis while also accumulating a large quantity of data. To fill this gap, OrganogenesisDB (http://organogenesisdb.com/), which is a comprehensive database dedicated to exploring cell-type identification and gene expression dynamics during organogenesis, is developed. OrganogenesisDB contains single-cell RNA sequencing data for more than 1.4 million cells from 49 published datasets spanning various developmental stages. Additionally, 3324 cell markers are manually curated for 1120 cell types across 9 human organs and 4 mouse organs. OrganogenesisDB leverages various analysis tools to assist users in annotating and understanding cell types at different developmental stages and helps in mining and presenting genes that exhibit specific patterns and play key regulatory roles during cell maturation and differentiation. This work provides a critical resource and useful tool for deciphering cell lineage determination and uncovering the mechanisms underlying organogenesis.

Abstract We091: Maternal hyperglycemia disrupts human cardiac cell lineage differentiation

Maternal diabetes is associated with a 4-fold increased risk of offspring developing congenital heart defects (CHDs). It remains unknown how maternal diabetes interferes with cardiac cell lineage determination and leads to malformations in the heart. In this study, we aim to elucidate the cellular mechanisms by which maternal hyperglycemia causes high risk of CHDs in newborns. Here we leverage an in vitro hyperglycemic model using human induced pluripotent stem cells (iPSCs), which could recapture cardiac differentiation and cell lineage commitment during embryonic heart development. We collected differentiating cells at D5 (cardiac mesoderm), D10 (cardiac progenitors), and D14 (early cardiomyocytes) during cardiac differentiation under normal and hyperglycemic conditions, and performed single-cell transcriptomic analysis. We found that hyperglycemia significantly impedes cardiac differentiation of human iPSCs as robust cardiac differentiation is rarely observed in multiple iPSC lines (n=10) under hyperglycemia. At the cellular level, hyperglycemia interferes with cardiac differentiation of human iPSCs in response to WNT signaling activation, which is manifested by reduced number of cardiac mesoderm (MESP1+ PDGFRA+) cells at D5 of differentiation. In contrast, neural differentiation is enhanced under hyperglycemia, with a high proportion of neural cell lineage (SOX2+ PAX6+). At D10, differentiated neural cell lineage dominates the cell population under hyperglycemia at the expense of cardiac progenitors and early cardiomyocytes. At D14, the prevalence of neural lineage persist whereas early cardiomyocytes only accounts for a small portion of cell population under hyperglycemia. Moreover, we treated D30 iPSC-derived cardiomyocytes (iPSC-CMs) with high glucose concentration (25 mM) for 7 days and found that iPSC-CMs show reduced mitochondria respiration and ATP production, but elevated apoptosis and reactive oxygen species (ROS) generation. Together, our data suggest that maternal hyperglycemia could interrupt human embryonic heart morphogenesis through overriding WNT-medicated cardiac differentiation and promoting neural cell lineage determination by default.

Abstract We029: Common and Divergent Cellular Etiologies Underlying Hypoplastic Left Heart Syndrome and Hypoplastic Right Heart Syndrome

Single ventricle heart defects are the most severe forms of congenital heart defects and are classified based on the affected ventricles: hypoplastic left heart syndrome (HLHS) and hypoplastic right heart syndrome (HRHS). Cellular etiologies underlying differential left and right ventricular hypoplasia are unknown and remain intractable without an experimental model to probe common and divergent cellular etiologies between HLHS and HRHS. In this study, we leveraged patient-specific induced pluripotent stem cells (iPSCs) and single-cell transcriptomics to interrogate distinct etiologies of HLHS and HRHS. Both HLHS and HRHS iPSC-derived cardiomyocytes (iPSC-CMs) exhibit a reduced proliferation capacity compared to sex-matched family controls under both static and cyclic stretch conditions. Biological pathways related to cell cycle progression, DNA replication, and cell proliferation are downregulated in both HLHS and HRHS iPSC-CMs, suggesting an intrinsic cardiac proliferation deficiency. Single-cell transcriptomics indicate that differentiation of cardiac mesoderm towards second heart field (SHF) progenitors are compromised in both HLHS and HRHS. However, differentiation of epicardial progenitors is enhanced at the expense of SHF progenitors in HLHS, whereas first heart field (FHF) progenitors are prevalent in HRHS. Trajectory inference analysis uncovers distinct cell lineage determination pathways in FHF and SHF progenitors between HLHS and HRHS. Moreover, HLHS iPSC-CMs exhibit reduced mitochondrial activities whereas HRHS iPSC-CMs show enhanced mitochondrial respiration and ATP production compared to controls. In sum, these common and divergent cellular etiologies of HLHS and HRHS may underlie ventricular hypoplasia in the left side and right side of the heart.

Investigating InDels in YAP and TAZ genes and their impact on growth characteristics in goats

Abstract. Yes-associated protein (YAP) and a transcriptional co-activator with PDZ-binding motif (TAZ) genes are crucial for regulating the size of mammalian tissues and organs as well as for many biological processes such as bone formation, cell lineage determination, tissue regeneration, and cell proliferation. The purpose of this study was to characterize the YAP and TAZ gene polymorphisms in 266 Guanzhong Dairy Goats and 299 Shanbei White Cashmere Goats and to explore their potential relationship with growth characteristics such as body weight and body length. After genotyping and using PCR amplification and Sanger sequencing to find polymorphisms in the YAP and TAZ genes, five InDels loci were found in the goat YAP gene and three InDels loci in the TAZ gene. The findings of the association analysis demonstrated that the goats' body weight, height, cannon circumference, chest depth, chest breadth, and chest circumference were all substantially influenced by five InDels loci in the YAP gene (p<0.05). Goat body height, trunk breadth, trunk length, body length, and body weight were all substantially impacted by three InDels loci in the TAZ gene (p<0.05). In conclusion, eight InDels loci of goat YAP and TAZ were found in this study, and their impacts on goat phenotype were disclosed. These results might offer fresh avenues for boosting goat molecular breeding.

PATH-02. THE IMPACT OF EVOLVING DIAGNOSTIC METHODOLOGY ON THE CLASSIFICATION AND OUTCOME OF CENTRAL NERVOUS SYSTEM TUMOURS IN CHILDHOOD: A RETROSPECTIVE COHORT STUDY

Abstract BACKGROUND Central nervous system tumours are the second most common childhood tumours and the largest cause of childhood cancer death in Australia. Tumours arise from many cell types within the developing and mature CNS, leading to a wide spectrum of clinical behaviour despite overlapping histological appearances. Emerging molecular testing methods allow more precise determination of cell lineage as well as genetic and epigenetic changes driving tumour development. Particularly since 2016, molecular findings have been incorporated into international classification schema, potentially impacting the frequency of individual diagnoses and their observed prognosis. METHODS We performed a retrospective cohort study including patients with newly diagnosed CNS tumours at Royal Children’s Hospital, Melbourne between 2011 and 2020 (total 473). The first and last 100 eligible patients in the data set were reviewed for diagnosis, survival and recorded use of molecular testing. Reviewed patients were assigned to one of two cohorts, Cohort 1 2011-2013 (pre-2016 WHO revision) or Cohort 2 2018-2020 (post-2016 WHO revision). RESULTS The frequency of diagnoses on biopsy rather than imaging alone was stable (81% Cohort 1 vs 83% Cohort 2). Molecular testing utilization increased markedly over time (10 molecular investigations in Cohort 1 vs 348 in Cohort 2). The incidence of astrocytic and oligodendroglial diagnoses was stable. The incidence of embryonal and neuronal/ mixed glial neuronal diagnoses increased in Cohort 2, and the incidence of ependymal diagnoses decreased (p=0.0122). There were no event free or overall survival differences between the whole of Cohorts 1 and 2, with numbers of individual diagnoses too small for meaningful comparisons over time. CONCLUSIONS Access to molecular testing has impacted on the pattern of reported diagnoses of central nervous system tumours in children. Although outcomes have not changed, it is hoped that over time, improved molecular understanding will lead to more effective therapy selection.

O13 Benchmarking a human pluripotent stem cell-derived hair-bearing skin organoid model to prenatal skin

Abstract Introduction and aims Molecular regulation of mammalian skin development has been largely derived from murine studies. The lack of human studies stems primarily from the difficulties in prenatal human skin access. There is an important need to study human prenatal skin owing to the notable differences between human and mouse skin and the paucity of mechanistic understanding of congenital skin disorders. Methods We assembled the first comprehensive multiomic reference atlas of prenatal human skin (21 samples from 7 to 16 postconception weeks), combining single-cell (235 201 cells) and spatial transcriptomic data. We followed 84 skin organoids (SkOs) using single-cell analysis over a 5-month culture period and characterized the spatial organization of the skin in mature SkOs. Prenatal skin, SkO and any relevant pre-existing single-cell RNA-Seq datasets were integrated into the analysis. Cell–cell interactions and cell-lineage determination were explored using CellPhoneDB and trajectory inferences. We evaluated the differentiation trajectory alignment between prenatal skin and SkO using the Genes2Genes analysis. Results Our findings reveal predicted factors regulating the epithelial–mesenchymal crosstalk during human hair follicle formation; progressive expression of scar-promoting genes in later gestation, potentially contributing to scarless healing properties of human prenatal skin observed during the early stages of gestation and expression of genes causing genetic skin disorders in the same cell types in prenatal human skin and SkO. We also found that SkOs morphologically recapitulate human skin development from early (periderm formation and shedding) to late (cornification and hair follicle formation) stages. We also demonstrated that SkO and prenatal skin mesenchyme differentiation into hair follicle dermal condensate and dermal papilla is highly conserved at the cellular and molecular level. Conclusions SkOs faithfully recapitulate human prenatal skin development and are a valuable model for mechanistic studies on skin development including hair follicle biology and congenital skin disorders.

The regulation and differentiation of regulatory T cells and their dysfunction in autoimmune diseases.

The discovery of FOXP3+ regulatory T (Treg)cells as a distinct cell lineage with a central role in regulating immune responses provided a deeper understanding of self-tolerance. The transcription factor FOXP3 serves a key role in Treg cell lineage determination and maintenance, but is not sufficient to enable the full potential of Treg cell suppression, indicating that other factors orchestrate the fine-tuning of Treg cell function. Moreover, FOXP3-independent mechanisms have recently been shown to contribute to Treg cell dysfunction. FOXP3 mutations in humans cause lethal fulminant systemic autoinflammation (IPEX syndrome). However, it remains unclear to what degree Treg cell dysfunction is contributing to the pathophysiology of common autoimmune diseases. In this Review, we discuss the origins of Treg cells in the periphery and the multilayered mechanisms by which Treg cells are induced, as well as the FOXP3-dependent and FOXP3-independent cellular programmes that maintain the suppressive function of Treg cells in humans and mice. Further, we examine evidence for Treg cell dysfunction in the context of common autoimmune diseases such as multiple sclerosis, inflammatory bowel disease, systemic lupus erythematosus and rheumatoid arthritis.

TET2 mutation as prototypic clonal hematopoiesis lesion

Loss of function TET2 mutation (TET2MT) is one of the most frequently observed lesions in clonal hematopoiesis (CH). TET2 a member TET-dioxygenase family of enzymes that along with TET1 and TET3, progressively oxidize 5-methyl cytosine (mC) resulting in regulated demethylation of promoter, enhancer and silencer elements of the genome. This process is critical for efficient transcription that determine cell lineage fate, proliferation and survival and the maintenance of the genomic fidelity with aging of the organism. Partial or complete loss-of-function TET2 mutations create regional and contextual DNA hypermethylation leading to gene silencing or activation that result in skewed myeloid differentiation and clonal expansion. In addition to myeloid skewing, loss of TET2 creates differentiation block and provides proliferative advantage to hematopoietic stem and progenitor cells (HSPCs). TET2MT is a prototypical lesion in CH, since the mutant clones dominate during stress hematopoiesis and often associates with evolution of myeloid malignancies. TET2MT clones has unique privilege to create and persist in pro-inflammatory milieu. Despite extensive knowledge regarding biochemical mechanisms underlying distorted myeloid differentiation, and enhanced self-replication of TET2MT HSPC, the mechanistic link of various pathogenesis associated with TET2 loss in CHIP is less understood. Here we review the recent development in TET2 biology and its probable mechanistic link in CH with aging and inflammation. We also explored the therapeutic strategies of targeting TET2MT associated CHIP and the utility of targeting TET2 in normal hematopoiesis and somatic cell reprograming. We explore the biochemical mechanisms and candidate therapies that emerged in last decade of research.

Differentiating Between Epstein-Barr Virus-positive Lymphoid Neoplasm Relapse and Post-transplant Lymphoproliferative Disorder After Sex-mismatched Hematopoietic Stem Cell Transplantation.

After allogeneic hematopoietic stem cell transplantation (HSCT), accurate differentiation between donor-derived post-transplant lymphoproliferative disorder (PTLD) and relapse of recipient-derived lymphoproliferative disorder (LPD) is crucial for determining treatment. Conventional diagnostic approaches for PTLD include histopathological examination, flow cytometry, and chimerism analysis of bulk tumor tissue. However, these methods are inconclusive in cases in which the primary disease is an Epstein-Barr virus (EBV)-positive LPD and is of the same lineage as that of the post-HSCT LPD tumor cells. Particularly, in cases where the number of tumor cells in the tissue is low, it is difficult to determine the origin of tumor cells. In this study, we developed a new method to simultaneously detect signals using sex chromosome fluorescence in situ hybridization, immunofluorescence staining, and EBV-encoded small RNA in situ hybridization on a single section of formalin-fixed paraffin-embedded histopathological specimen. The utility of the method was validated using specimens from 6 cases of EBV-positive LPD after sex-mismatched HSCT that were previously difficult to diagnose, including Hodgkin lymphoma-like PTLD that developed after HSCT for Hodgkin lymphoma and recurrence of chronic active EBV infection. This method successfully preserved the histologic structure after staining and allowed accurate determination of tumor cell origin and lineage at the single-cell level, providing a definitive diagnosis in all cases. This method provides a powerful tool for the diagnosis of LPDs after sex-mismatched HSCT.

Participation of the TBP-associatedfactors (TAFs) in cell differentiation.

The understanding of the mechanisms that regulate gene expression to establish differentiation programs and determine cell lineages, is one of the major challenges in Developmental Biology. Besides the participation of tissue-specific transcription factors and epigenetic processes, the role of general transcription factors has been ignored. Only in recent years, there have been scarce studies that address this issue. Here, we review the studies on the biological activity of some TATA-box binding protein (TBP)-associatedfactors (TAFs)during the proliferation of stem/progenitor cells and their involvement in cell differentiation. Particularly, the accumulated evidence suggests that TAF4, TAF4b, TAF7L, TAF8, TAF9, and TAF10, among others, participate in nervous system development, adipogenesis, myogenesis, and epidermal differentiation; while TAF1, TAF7, TAF15 may be involved in the regulation of stem cell proliferative abilities and cell cycle progression. On the other hand, evidence suggests that TBP variants such as TBPL1 and TBPL2 might be regulating some developmental processes such as germ cell maturation and differentiation, myogenesis, or ventral specification during development. Our analysis shows that it is necessary to study in greater depth the biological function of these factors and its participation in the assembly of specific transcription complexes that contribute to the differential gene expression that gives rise to the great diversity of cell types existing in an organism. The understanding of TAFs'regulation might lead to the development of new therapies for patients which suffer from mutations, alterations, and dysregulation of these essential elements of the transcriptional machinery.

Differential Effect of Transferrin Lobe Iron Occupancy on a Murine Model of Inflammation

Introduction: Iron status is both affected by and a determinant of the inflammatory response. Several lines of evidence support a role for transferrin as a signaling as well as an iron delivery molecule. We previously demonstrated that mice with transferrin mutations that prevent iron binding to the C-lobe of transferrin (C-blocked) have increased red cell production relative to erythropoietin and increased hepcidin expression relative to iron status compared to mice with transferrin mutations that prevent iron binding to the N-lobe of transferrin (N-blocked). Based on these observations, we hypothesize that two forms of transferrin would differentially modulate the hematopoietic and iron distribution changes that occur with inflammation. We explored this hypothesis by comparing the effects of lipopolysaccharide (LPS) administered either acutely or in repeated doses to wild-type, N-lobe blocked, or C-lobe blocked transferrin mutant mice. Methods: For a model of acute inflammation, 14 day old wild-type (wt), N-blocked and C-blocked Tf mutant mice were treated with 1mg/kg LPS or carrier i.p. and sacrificed 6 hours later. Repeated LPS dosing, seven daily i.p. injections of 0.3 mg/kg LPS or carrier beginning at seven days of age, served as a model of chronic inflammation. Results: While wild-type and C-blocked mice had the expected hypoferremia in response to acute LPS administration, there was no effect on serum iron in the N-blocked mice. These mice moreover demonstrated an unanticipated increase in liver iron concentrations with LPS. All strains manifested expected increases in liver hepcidin expression; however, these were attenuated in the N-blocked mice relative to the C-blocked. Moreover, N-blocked mice have higher serum C reactive protein concentrations compared to C-blocked and wt mice following acute LPS administration. The elevated Hamp1 expression was not sustained in the chronic model. Although serum iron returned to baseline in wt and C-blocked mice, it remained elevated in N-blocked mice. Hemoglobin concentrations were decreased in all strains after repeated endotoxin dosing; however, C-blocked mice demonstrated higher neutrophil to lymphocyte ratios than wt or N-blocked mice, and increased circulating RBC count relative to WBCs. Conclusions: We conclude that the specificity of transferrin lobe iron occupancy is an important modulator of the inflammatory response and speculate that this modulation includes an effect on hematopoietic cell lineage determination.

Ganglioside GD3 regulates neural stem cell quiescence and controls postnatal neurogenesis.

The postnatal neural stem cell (NSC) pool hosts quiescent and activated radial glia-like NSCs contributing to neurogenesis throughout adulthood. However, the underlying regulatory mechanism during the transition from quiescent NSCs to activated NSCs in the postnatal NSC niche is not fully understood. Lipid metabolism and lipid composition play important roles in regulating NSC fate determination. Biological lipid membranes define the individual cellular shape and help maintain cellular organization and are highly heterogeneous in structure and there exist diverse microdomains (also known as lipid rafts), which are enriched with sugar molecules, such as glycosphingolipids. An often overlooked but key aspect is that the functional activities of proteins and genes are highly dependent on their molecular environments. We previously reported that ganglioside GD3 is the predominant species in NSCs and that the reduced postnatal NSC pools are observed in global GD3-synthase knockout (GD3S-KO) mouse brains. The specific roles of GD3 in determining the stage and cell-lineage determination of NSCs remain unclear, since global GD3S-KO mice cannot distinguish if GD3 regulates postnatal neurogenesis or developmental impacts. Here, we show that inducible GD3 deletion in postnatal radial glia-like NSCs promotes NSC activation, resulting in the loss of the long-term maintenance of the adult NSC pools. The reduced neurogenesis in the subventricular zone (SVZ) and the dentate gyrus (DG) of GD3S-conditional-knockout mice led to the impaired olfactory and memory functions. Thus, our results provide convincing evidence that postnatal GD3 maintains the quiescent state of radial glia-like NSCs in the adult NSC niche.

A functional crosstalk between the H3K9 methylation writers and their reader HP1 in safeguarding embryonic stem cell identity.

Histone H3 lysine 9 (H3K9) methylation, as a hallmark of heterochromatin, has a central role in cell lineage and fate determination. Although evidence of a cooperation between H3K9 methylation writers and their readers has started to emerge, their actual interplay remains elusive. Here, we show that loss of H3K9 methylation readers, the Hp1 family, causes reduced expression of H3K9 methyltransferases, and that this subsequently leads to the exit of embryonic stem cells (ESCs) from pluripotency and a reciprocal gain of lineage-specific characteristics. Importantly, the phenotypes of Hp1-null ESCs can be rescued by ectopic expression of Setdb1, Nanog, and Oct4. Furthermore, Setdb1 ablation results in loss of ESC identity, which is accompanied by a reduction in the expression of Hp1 genes. Together, our data support a model in which the safeguarding of ESC identity involves the cooperation between the H3K9 methylation writers and their readers.

Abstract P1029: Abnormal Progenitor Cell Differentiation And Cardiomyocyte Proliferation In Hypoplastic Right Heart Syndrome

Pulmonary atresia with intact ventricular septum (PA-IVS) is a rare (4-8 per 100,000 live births) type of hypoplastic right heart syndrome (HRHS) where the right-sided structures in the heart are malformed. In PA-IVS, the pulmonary valve that acts to regulate the unidirectional flow of blood from the right ventricle (RV) to the lungs does not open, resulting in no connection between the RV and the pulmonary arteries. After surgical or catheter-based intervention, PA-IVS patients have clinical outcomes that range from single-ventricle palliation to a biventricular repair. Mechanisms underlying the spectrum of RV hypoplasia in PA-IVS are difficult to fully ascribe to an atretic pulmonary valve. There are no reliable animal models available for studying disease mechanisms of PA-IVS. In this study, we leverage patient-specific induced pluripotent stem cells (iPSCs) and single-cell RNA sequencing to elucidate cellular etiologies of ventricular hypoplasia in PA-IVS. PA-IVS iPSC-derived cardiomyocytes are less proliferative compared to controls under both static and cyclic stretch, suggesting that genetic factors may contribute to a spectrum of ventricular hypoplasia in PA-IVS. Single-cell transcriptomic analysis reveals that cell lineage determination towards second heart progenitors is suppressed, with enhanced differentiation into first heart field and epicardial lineages during cardiac differentiation. Additionally, biological pathways associated with cell proliferation and cell cycle progression are downregulated whereas mitochondrial activities are elevated in early cardiomyocytes originated from PA-IVS patients. In conclusion, abnormal cell lineage differentiation and cardiomyocyte proliferation may underlie the cellular and developmental etiologies of ventricular hypoplasia in HRHS.

Co-Expression of Multiple PAX Genes in Renal Cell Carcinoma (RCC) and Correlation of High PAX Expression with Favorable Clinical Outcome in RCC Patients.

Renal cell carcinoma (RCC) is the most common form of kidney cancer, consisting of multiple distinct subtypes. RCC has the highest mortality rate amongst the urogenital cancers, with kidney renal clear cell carcinoma (KIRC), kidney renal papillary cell carcinoma (KIRP), and kidney chromophobe carcinoma (KICH) being the most common subtypes. The Paired-box (PAX) gene family encodes transcription factors, which orchestrate multiple processes in cell lineage determination during embryonic development and organogenesis. Several PAX genes have been shown to be expressed in RCC following its onset and progression. Here, we performed real-time quantitative polymerase chain reaction (RT-qPCR) analysis on a series of human RCC cell lines, revealing significant co-expression of PAX2, PAX6, and PAX8. Knockdown of PAX2 or PAX8 mRNA expression using RNA interference (RNAi) in the A498 RCC cell line resulted in inhibition of cell proliferation, which aligns with our previous research, although no reduction in cell proliferation was observed using a PAX2 small interfering RNA (siRNA). We downloaded publicly available RNA-sequencing data and clinical histories of RCC patients from The Cancer Genome Atlas (TCGA) database. Based on the expression levels of PAX2, PAX6, and PAX8, RCC patients were categorized into two PAX expression subtypes, PAXClusterA and PAXClusterB, exhibiting significant differences in clinical characteristics. We found that the PAXClusterA expression subgroup was associated with favorable clinical outcomes and better overall survival. These findings provide novel insights into the association between PAX gene expression levels and clinical outcomes in RCC patients, potentially contributing to improved treatment strategies for RCC.

Transcriptional and Epigenetic Regulation of Context-Dependent Plasticity in T-Helper Lineages.

Th cell lineage determination and functional specialization are tightly linked to the activation of lineage-determining transcription factors (TFs) that bind cis-regulatory elements. These lineage-determining TFs act in concert with multiple layers of transcriptional regulators to alter the epigenetic landscape, including DNA methylation, histone modification and three-dimensional chromosome architecture, in order to facilitate the specific Th gene expression programs that allow for phenotypic diversification. Accumulating evidence indicates that Th cell differentiation is not as rigid as classically held; rather, extensive phenotypic plasticity is an inherent feature of T cell lineages. Recent studies have begun to uncover the epigenetic programs that mechanistically govern T cell subset specification and immunological memory. Advances in next generation sequencing technologies have allowed global transcriptomic and epigenomic interrogation of CD4+ Th cells that extends previous findings focusing on individual loci. In this review, we provide an overview of recent genome-wide insights into the transcriptional and epigenetic regulation of CD4+ T cell-mediated adaptive immunity and discuss the implications for disease as well as immunotherapies.

Impaired Human Cardiac Cell Development due to NOTCH1 Deficiency.

NOTCH1 pathogenic variants are implicated in multiple types of congenital heart defects including hypoplastic left heart syndrome, where the left ventricle is underdeveloped. It is unknown how NOTCH1 regulates human cardiac cell lineage determination and cardiomyocyte proliferation. In addition, mechanisms by which NOTCH1 pathogenic variants lead to ventricular hypoplasia in hypoplastic left heart syndrome remain elusive. CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas9 genome editing was utilized to delete NOTCH1 in human induced pluripotent stem cells. Cardiac differentiation was carried out by sequential modulation of WNT signaling, and NOTCH1 knockout and wild-type differentiating cells were collected at day 0, 2, 5, 10, 14, and 30 for single-cell RNA-seq. Human NOTCH1 knockout induced pluripotent stem cells are able to generate functional cardiomyocytes and endothelial cells, suggesting that NOTCH1 is not required for mesoderm differentiation and cardiovascular development in vitro. However, disruption of NOTCH1 blocks human ventricular-like cardiomyocyte differentiation but promotes atrial-like cardiomyocyte generation through shortening the action potential duration. NOTCH1 deficiency leads to defective proliferation of early human cardiomyocytes, and transcriptomic analysis indicates that pathways involved in cell cycle progression and mitosis are downregulated in NOTCH1 knockout cardiomyocytes. Single-cell transcriptomic analysis reveals abnormal cell lineage determination of cardiac mesoderm, which is manifested by the biased differentiation toward epicardial and second heart field progenitors at the expense of first heart field progenitors in NOTCH1 knockout cell populations. NOTCH1 is essential for human ventricular-like cardiomyocyte differentiation and proliferation through balancing cell fate determination of cardiac mesoderm and modulating cell cycle progression. Because first heart field progenitors primarily contribute to the left ventricle, we speculate that pathogenic NOTCH1 variants lead to biased differentiation of first heart field progenitors, blocked ventricular-like cardiomyocyte differentiation, and defective cardiomyocyte proliferation, which collaboratively contribute to left ventricular hypoplasia in hypoplastic left heart syndrome.

The p53 Family Members p63 and p73 Roles in the Metastatic Dissemination: Interactions with microRNAs and TGFβ Pathway

TP53 (TP53), p73 (TP73), and p63 (TP63) are members of the p53 transcription factor family, which has many activities spanning from embryonic development through to tumor suppression. The utilization of two promoters and alternative mRNA splicing has been shown to yield numerous isoforms in p53, p63, and p73. TAp73 is thought to mediate apoptosis as a result of nuclear accumulation following chemotherapy-induced DNA damage, according to a number of studies. Overexpression of the nuclear ΔNp63 and ΔNp73 isoforms, on the other hand, suppresses TAp73's pro-apoptotic activity in human malignancies, potentially leading to metastatic spread or inhibition. Another well-known pathway that has been associated to metastatic spread is the TGF pathway. TGFs are a family of structurally related polypeptide growth factors that regulate a variety of cellular functions including cell proliferation, lineage determination, differentiation, motility, adhesion, and cell death, making them significant players in development, homeostasis, and wound repair. Various studies have already identified several interactions between the p53 protein family and the TGFb pathway in the context of tumor growth and metastatic spread, beginning to shed light on this enigmatic intricacy.

BCL7A‐containing SWI/SNF/BAF complexes modulate mitochondrial bioenergetics during neural progenitor differentiation

Mammalian SWI/SNF/BAF chromatin remodeling complexes influence cell lineage determination. While the contribution of these complexes to neural progenitor cell (NPC) proliferation and differentiation has been reported, little is known about the transcriptional profiles that determine neurogenesis or gliogenesis. Here, we report that BCL7A is a modulator of the SWI/SNF/BAF complex that stimulates the genome‐wide occupancy of the ATPase subunit BRG1. We demonstrate that BCL7A is dispensable for SWI/SNF/BAF complex integrity, whereas it is essential to regulate Notch/Wnt pathway signaling and mitochondrial bioenergetics in differentiating NPCs. Pharmacological stimulation of Wnt signaling restores mitochondrial respiration and attenuates the defective neurogenic patterns observed in NPCs lacking BCL7A. Consistently, treatment with an enhancer of mitochondrial biogenesis, pioglitazone, partially restores mitochondrial respiration and stimulates neuronal differentiation of BCL7A‐deficient NPCs. Using conditional BCL7A knockout mice, we reveal that BCL7A expression in NPCs and postmitotic neurons is required for neuronal plasticity and supports behavioral and cognitive performance. Together, our findings define the specific contribution of BCL7A‐containing SWI/SNF/BAF complexes to mitochondria‐driven NPC commitment, thereby providing a better understanding of the cell‐intrinsic transcriptional processes that connect metabolism, neuronal morphogenesis, and cognitive flexibility.