Partial reprogramming restores youthful gene expression through transient suppression of cell identity

Transient induction of pluripotent reprogramming factors has been reported to reverse some features of aging in mammalian cells and tissues. However, the impact of transient reprogramming on somatic cell identity programs and the necessity of individual pluripotency factors remain unknown. Here, we mapped trajectories of transient reprogramming in young and aged cells from multiple murine cell types using single cell transcriptomics to address these questions. We found that transient reprogramming restored youthful gene expression in adipocytes and mesenchymal stem cells but also temporarily suppressed somatic cell identity programs. We further screened Yamanaka Factor subsets and found that many combinations had an impact on aging gene expression and suppressed somatic identity, but that these effects were not tightly entangled. We also found that a transient reprogramming approach inspired by amphibian regeneration restored youthful gene expression in aged myogenic cells. Our results suggest that transient pluripotent reprogramming poses a neoplastic risk, but that restoration of youthful gene expression can be achieved with alternative strategies.

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Simultaneous quantification of protein-DNA contacts and transcriptomes in single cells

10.1101/529388 ◽

2019 ◽

Cited By ~ 1

Author(s):

Koos Rooijers ◽

Corina M. Markodimitraki ◽

Franka J. Rang ◽

Sandra S. de Vries ◽

Alex Chialastri ◽

...

Keyword(s):

Gene Expression ◽

Mammalian Cells ◽

Single Cells ◽

Critical Role ◽

Nuclear Lamina ◽

Cell Types ◽

Chromatin Accessibility ◽

Expression Variability ◽

The Impact

AbstractThe epigenome plays a critical role in regulating gene expression in mammalian cells. However, understanding how cell-to-cell heterogeneity in the epigenome influences gene expression variability remains a major challenge. Here we report a novel method for simultaneous single-cell quantification of protein-DNA contacts with DamID and transcriptomics (scDamID&T). This method enables quantifying the impact of protein-DNA contacts on gene expression from the same cell. By profiling lamina-associated domains (LADs) in human cells, we reveal different dependencies between genome-nuclear lamina (NL) association and gene expression in single cells. In addition, we introduce the E. coli methyltransferase, Dam, as an in vivo marker of chromatin accessibility in single cells and show that scDamID&T can be utilized as a general technology to identify cell types in silico while simultaneously determining the underlying gene-regulatory landscape. With this strategy the effect of chromatin states, transcription factor binding, and genome organization on the acquisition of cell-type specific transcriptional programs can be quantified.

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Optimizing expression quantitative trait locus mapping workflows for single-cell studies

Genome Biology ◽

10.1186/s13059-021-02407-x ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Anna S. E. Cuomo ◽

Giordano Alvari ◽

Christina B. Azodi ◽

Davis J. McCarthy ◽

Marc Jan Bonder ◽

...

Keyword(s):

Gene Expression ◽

Quantitative Trait Locus ◽

Single Cell ◽

Quantitative Trait ◽

Cell Types ◽

Expression Quantitative Trait Locus ◽

Eqtl Mapping ◽

Trait Locus ◽

The Impact

Abstract Background Single-cell RNA sequencing (scRNA-seq) has enabled the unbiased, high-throughput quantification of gene expression specific to cell types and states. With the cost of scRNA-seq decreasing and techniques for sample multiplexing improving, population-scale scRNA-seq, and thus single-cell expression quantitative trait locus (sc-eQTL) mapping, is increasingly feasible. Mapping of sc-eQTL provides additional resolution to study the regulatory role of common genetic variants on gene expression across a plethora of cell types and states and promises to improve our understanding of genetic regulation across tissues in both health and disease. Results While previously established methods for bulk eQTL mapping can, in principle, be applied to sc-eQTL mapping, there are a number of open questions about how best to process scRNA-seq data and adapt bulk methods to optimize sc-eQTL mapping. Here, we evaluate the role of different normalization and aggregation strategies, covariate adjustment techniques, and multiple testing correction methods to establish best practice guidelines. We use both real and simulated datasets across single-cell technologies to systematically assess the impact of these different statistical approaches. Conclusion We provide recommendations for future single-cell eQTL studies that can yield up to twice as many eQTL discoveries as default approaches ported from bulk studies.

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Defining cell identity beyond the premise of differential gene expression

Cell Regeneration ◽

10.1186/s13619-021-00083-7 ◽

2021 ◽

Vol 10 (1) ◽

Author(s):

Hani Jieun Kim ◽

Patrick P. L. Tam ◽

Pengyi Yang

Keyword(s):

Gene Expression ◽

Differential Expression ◽

Cell Fate ◽

Differential Gene Expression ◽

Cell Types ◽

Cell Identity ◽

Gene Transcripts ◽

Differential Gene ◽

Biological Attributes

AbstractIdentifying genes that define cell identity is a requisite step for characterising cell types and cell states and predicting cell fate choices. By far, the most widely used approach for this task is based on differential expression (DE) of genes, whereby the shift of mean expression are used as the primary statistics for identifying gene transcripts that are specific to cell types and states. While DE-based methods are useful for pinpointing genes that discriminate cell types, their reliance on measuring difference in mean expression may not reflect the biological attributes of cell identity genes. Here, we highlight the quest for non-DE methods and provide an overview of these methods and their applications to identify genes that define cell identity and functionality.

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Enhanced Transcriptome Maps from Multiple Mouse Tissues Reveal Evolutionary Constraint in Gene Expression for Thousands of Genes

10.1101/010884 ◽

2014 ◽

Cited By ~ 1

Author(s):

Dmitri Pervouchine ◽

Sarah Djebali ◽

Alessandra Breschi ◽

Carrie A Davis ◽

Pablo Prieto Barja ◽

...

Keyword(s):

Gene Expression ◽

Mammalian Cells ◽

Cellular Localization ◽

Cell Types ◽

Evolutionary Constraint ◽

Novel Transcript ◽

Transcriptional Regulatory ◽

Mouse Tissues ◽

Mouse Transcriptome ◽

Transcriptional Output

We characterized by RNA-seq the transcriptional profiles of a large and heterogeneous collection of mouse tissues, augmenting the mouse transcriptome with thousands of novel transcript candidates. Comparison with transcriptome profiles obtained in human cell lines reveals substantial conservation of transcriptional programs, and uncovers a distinct class of genes with levels of expression across cell types and species, that have been constrained early in vertebrate evolution. This core set of genes capture a substantial and constant fraction of the transcriptional output of mammalian cells, and participates in basic functional and structural housekeeping processes common to all cell types. Perturbation of these constrained genes is associated with significant phenotypes including embryonic lethality and cancer. Evolutionary constraint in gene expression levels is not reflected in the conservation of the genomic sequences, but it is associated with strong and conserved epigenetic marking, as well as to a characteristic post-transcriptional regulatory program in which sub-cellular localization and alternative splicing play comparatively large roles.

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Optimising expression quantitative trait locus mapping workflows for single-cell studies

10.1101/2021.01.20.427401 ◽

2021 ◽

Author(s):

Anna S.E. Cuomo ◽

Giordano Alvari ◽

Christina B. Azodi ◽

Davis J. McCarthy ◽

Marc Jan Bonder ◽

...

Keyword(s):

Gene Expression ◽

Quantitative Trait Locus ◽

Single Cell ◽

Quantitative Trait ◽

Cell Types ◽

Expression Quantitative Trait Locus ◽

Eqtl Mapping ◽

Trait Locus ◽

The Impact

AbstractSingle-cell RNA-sequencing (scRNA-seq) has enabled the unbiased, high-throughput quantification of gene expression specific to cell types and states. With the cost of scRNA-seq decreasing and techniques for sample multiplexing improving, population-scale scRNA-seq, and thus single-cell expression quantitative trait locus (sc-eQTL) mapping, is increasingly feasible. Mapping of sc-eQTL provides additional resolution to study the regulatory role of common genetic variants on gene expression across a plethora of cell types and states, and promises to improve our understanding of genetic regulation across tissues in both health and disease. While previously established methods for bulk eQTL mapping can, in principle, be applied to sc-eQTL mapping, there are a number of open questions about how best to process scRNA-seq data and adapt bulk methods to optimise sc-eQTL mapping. Here, we evaluate the role of different normalisation and aggregation strategies, covariate adjustment techniques, and multiple testing correction methods to establish best practice guidelines. We use both real and simulated datasets across single-cell technologies to systematically assess the impact of these different statistical approaches and provide recommendations for future single-cell eQTL studies that can yield up to twice as many eQTL discoveries as default approaches ported from bulk studies.

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Frequent and Transient Acquisition of Pluripotency During Somatic Cell Trans-differentiation with iPSC Reprogramming Factors

10.1101/008284 ◽

2014 ◽

Author(s):

Itay Maza ◽

Inbal Casoi ◽

Sergey Viukov ◽

Yoach Rais ◽

Asaf Zviran ◽

...

Keyword(s):

Somatic Cell ◽

Cell Types ◽

Lineage Tracing ◽

Functional Coupling ◽

Genetic Lineage ◽

New Paradigm ◽

Apply Genetic ◽

Lineage Differentiation ◽

Molecular Events ◽

Reprogramming Factors

Recent reports have proposed a new paradigm for obtaining mature somatic cell types from fibroblasts without going through a pluripotent state, by briefly expressing canonical iPSC reprogramming factors Oct4, Sox2, Klf4, c-Myc (abbreviated as OSKM) in cells expanded in lineage differentiation promoting conditions. Here we apply genetic lineage tracing for endogenous Nanog locus and X chromosome reactivation during OSKM induced trans-differentiation, as these molecular events mark final stages for acquisition of induced pluripotency. Remarkably, the majority of reprogrammed cardiomyocytes or neural stem cells derived from mouse fibroblasts via OSKM mediated trans-differentiation (~>90%), are attained after transient acquisition of pluripotency, and followed by rapid differentiation. Our findings underscore a molecular and functional coupling between inducing pluripotency and obtaining “trans-differentiated” somatic cells via OSKM induction, and have implications on defining molecular trajectories assumed during different cell reprogramming methods.

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Single-nuclei chromatin profiling of ventral midbrain reveals cell identity transcription factors and cell-type-specific gene regulatory variation

Epigenetics & Chromatin ◽

10.1186/s13072-021-00418-3 ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Yujuan Gui ◽

Kamil Grzyb ◽

Mélanie H. Thomas ◽

Jochen Ohnmacht ◽

Pierre Garcia ◽

...

Keyword(s):

Gene Expression ◽

Transcription Factors ◽

Cell Types ◽

Mouse Strains ◽

Cell Type ◽

Data Set ◽

Cell Identity ◽

Regulatory Variation ◽

Ventral Midbrain ◽

Cell Type Specific

Abstract Background Cell types in ventral midbrain are involved in diseases with variable genetic susceptibility, such as Parkinson’s disease and schizophrenia. Many genetic variants affect regulatory regions and alter gene expression in a cell-type-specific manner depending on the chromatin structure and accessibility. Results We report 20,658 single-nuclei chromatin accessibility profiles of ventral midbrain from two genetically and phenotypically distinct mouse strains. We distinguish ten cell types based on chromatin profiles and analysis of accessible regions controlling cell identity genes highlights cell-type-specific key transcription factors. Regulatory variation segregating the mouse strains manifests more on transcriptome than chromatin level. However, cell-type-level data reveals changes not captured at tissue level. To discover the scope and cell-type specificity of cis-acting variation in midbrain gene expression, we identify putative regulatory variants and show them to be enriched at differentially expressed loci. Finally, we find TCF7L2 to mediate trans-acting variation selectively in midbrain neurons. Conclusions Our data set provides an extensive resource to study gene regulation in mesencephalon and provides insights into control of cell identity in the midbrain and identifies cell-type-specific regulatory variation possibly underlying phenotypic and behavioural differences between mouse strains.

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Maternal Overnutrition Programs Central Inflammation and Addiction-Like Behavior in Offspring

BioMed Research International ◽

10.1155/2018/8061389 ◽

2018 ◽

Vol 2018 ◽

pp. 1-11 ◽

Cited By ~ 13

Author(s):

Larisa Montalvo-Martínez ◽

Roger Maldonado-Ruiz ◽

Marcela Cárdenas-Tueme ◽

Diana Reséndez-Pérez ◽

Alberto Camacho

Keyword(s):

Gene Expression ◽

Fetal Programming ◽

Gene Expression Pattern ◽

Cell Types ◽

Addictive Behavior ◽

Adaptive Responses ◽

Inflammatory Phenotype ◽

Pregnancy And Lactation ◽

Maternal Overnutrition ◽

The Impact

Obesity or maternal overnutrition during pregnancy and lactation might have long-term consequences in offspring health. Fetal programming is characterized by adaptive responses to specific environmental conditions during early life stages. Programming alters gene expression through epigenetic modifications leading to a transgenerational effect of behavioral phenotypes in the offspring. Maternal intake of hypercaloric diets during fetal development programs aberrant behaviors resembling addiction in offspring. Programming by hypercaloric surplus sets a gene expression pattern modulating axonal pruning, synaptic signaling, and synaptic plasticity in selective regions of the reward system. Likewise, fetal programming can promote an inflammatory phenotype in peripheral and central sites through different cell types such as microglia and T and B cells, which contribute to disrupted energy sensing and behavioral pathways. The molecular mechanism that regulates the central and peripheral immune cross-talk during fetal programming and its relevance on offspring’s addictive behavior susceptibility is still unclear. Here, we review the most relevant scientific reports about the impact of hypercaloric nutritional fetal programming on central and peripheral inflammation and its effects on addictive behavior of the offspring.

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A Synthetic Transcription Platform for Programmable Gene Expression in Mammalian Cells

10.1101/2020.12.11.420000 ◽

2020 ◽

Author(s):

William C.W. Chen ◽

Leonid Gaidukov ◽

Ming-Ru Wu ◽

Jicong Cao ◽

Gigi C.G. Choi ◽

...

Keyword(s):

Gene Expression ◽

Interferon Gamma ◽

Mammalian Cells ◽

Cell Types ◽

Regulatory Elements ◽

Genetic Circuits ◽

Control Of Gene Expression ◽

Transcription System ◽

Multiple Cell ◽

Cellular Phenotypes

Precise, scalable, and sustainable control of genetic and cellular activities in mammalian cells is key to developing precision therapeutics and smart biomanufacturing. We created a highly tunable, modular, versatile CRISPR-based synthetic transcription system for the programmable control of gene expression and cellular phenotypes in mammalian cells. Genetic circuits consisting of well-characterized libraries of guide RNAs, binding motifs of synthetic operators, transcriptional activators, and additional genetic regulatory elements expressed mammalian genes in a highly predictable and tunable manner. We demonstrated the programmable control of reporter genes episomally and chromosomally, with up to 25-fold more EF1[alpha]; promoter activity, in multiple cell types. We used these circuits to program secretion of human monoclonal antibodies and to control T cell effector function marked by interferon-[gamma] production. Antibody titers and interferon-[gamma]; concentrations were significantly correlated with synthetic promoter strengths, providing a platform for programming gene expression and cellular function for biological, biomanufacturing, and biomedical applications.

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131. SIRT3 IN OVARIAN CELLS IS ALTERED BY MATERNAL AGE AND OVARIAN RESERVE

Reproduction Fertility and Development ◽

10.1071/srb10abs131 ◽

2010 ◽

Vol 22 (9) ◽

pp. 49

Author(s):

L. Pacella ◽

D. Zander-Fox ◽

T. Hussein ◽

T. Fullston ◽

M. Lane

Keyword(s):

Gene Expression ◽

Ovarian Reserve ◽

Maternal Age ◽

Cumulus Cells ◽

Cell Types ◽

Advanced Maternal Age ◽

Cellular Functions ◽

Protein Levels ◽

Ovarian Cells ◽

The Impact

Maternal age and reduced AMH levels affect the follicular environment and consequently oocyte viability. The Sirtuin family of protein deacetylases are able to regulate various cellular functions involved in the ageing process in other tissues. In particular, SIRT3 is related to longevity in several cell types and regulates mitochondrial function, however, its presence and role in ovarian cells remains unknown. This study therefore, investigated the presence of SIRT3 in granulosa and cumulus cells, from patients undergoing IVF, and determined the impact of maternal age and low AMH on SIRT3 levels. Granulosa and cumulus cells were collected from women (n = 36), after informed consent, and classified into 3 groups; A (<35 years, normal AMH), B (>40 years (advanced maternal age), normal AMH) and C (<35 years, low AMH). The presence of SIRT3 was determined by q-PCR (expressed as fold-change) or immunohistochemistry. SIRT3 was present in the ovarian cells of all patients analysed. SIRT3 gene expression was reduced in granulosa cells from women with low AMH (0.67 ± 0.17) compared to women with normal AMH (1.00 ± 0.23; P < 0.05). In cumulus cells, levels were reduced with advanced maternal age (0.81 ± 0.08) compared to women <35 years (1.00 ± 0.22; P < 0.05). SIRT3 protein co-localised with mitochondria in the ovarian cells, confirming previous findings for other cell types. In comparison to women <35 years with normal AMH, image analysis determined that SIRT3 protein levels were significantly reduced in the granulosa and cumulus cells from women of advanced maternal age by 21.4% and 31.8% and in women with low AMH by 34.1% and 47.2% respectively. This is the first study to demonstrate SIRT3 presence in human ovarian cells. The observation that SIRT3 levels are altered by advanced maternal age or low AMH (reduced ovarian reserve) implicate its role in ovarian ageing and plausibly in the decrease in oocyte viability observed in these women.

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